Composition for inhibiting the proliferation of cancer cells

Abstract

To provide a material or component having antitumor effect using a plant raw material that has been eaten since a long time ago.SOLUTION: A composition for inhibiting the proliferation of cancer cells contains Fallopia japonica extract as an active ingredient. A composition for promoting the apoptosis of cancer cells contains Fallopia japonica extract as an active ingredient.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] According to the overview of vital statistics in 2020 (Reiwa 2) by the Ministry of Health, Labour and Welfare, cancer (malignant neoplasm, tumor) ranks first as the cause of death for both men and women, and the proportion in the total number of deaths has reached as high as 27.6%. This proportion has increased by 0.3% compared with the same statistical overview in 2019, and the development of active ingredients and treatment methods with high effectiveness and safety is still strongly desired.

[0002] From the perspective of safety, materials and components with antitumor effects have been screened from plant raw materials with food experience, and their application to antitumor agents has been studied. For example, in Patent Document 1, purified polysaccharides have been found as active ingredients of antitumor agents from crude drugs derived from plants of the Curcuma genus, such as zedoary (Rhizoma zedoariae) and turmeric (Curcuma longa).

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0004] In recent years in Japan, attention has been focused on preventive medicine in order to suppress the increasing medical costs, and the development of effective and safe food products, pharmaceuticals or quasi-drugs that can reduce the disease incidence risk is strongly desired.

[0005] Therefore, an object of the present invention is to provide materials and components with antitumor effects by using plant raw materials with food experience.

Means for Solving the Problems

[0007] In other words, the present invention provides the following compositions. [1] A composition for inhibiting the proliferation of cancer cells, containing an extract of Japanese knotweed (Fallopia japonica) as an active ingredient.

[0008] [2] A composition for promoting apoptosis of cancer cells, containing Japanese knotweed extract as an active ingredient.

[0009] [3] The composition for inhibiting the proliferation of cancer cells according to [1] or [2], wherein the extraction site of the Japanese knotweed extract is the leaves of the Japanese knotweed.

[0010] [4] The composition for inhibiting the proliferation of cancer cells according to any one of claims [1] to [3], wherein the Japanese knotweed extract contains at least one selected from the group consisting of neochlorogenic acid, piceatannol, rutin, and quercetin.

[0011] [5] The composition for inhibiting the proliferation of cancer cells according to any one of [1] to [4], wherein the Japanese knotweed extract contains an effective amount of neochlorogenic acid and rutin.

[0012] [6] The cancer cell proliferation inhibitory composition according to any one of [1] to [5], wherein the cancer cell is a colorectal cancer cell. [Effects of the Invention]

[0013] It is possible to provide a composition for suppressing the growth of cancer cells and promoting apoptosis of cancer cells with an extract derived from a plant that grows wild in Kochi Prefecture and other areas and has been eaten as wild vegetables and traditional cuisine.

Brief Description of the Drawings

[0014] [Figure 1] Figure 1 is a graph showing the determination results of cell viability by the WST-8 method in Test Example 1. [Figure 2] Figure 2 is a photographic image showing the morphological changes of Colon-26 cells in Test Example 2. [Figure 3] Figure 3 is an agarose electrophoresis photograph showing the results of evaluating the DNA fragmentation of Colon-26 cells by the Japanese knotweed extract in Test Example 3. [Figure 4] Figure 4 is a graph showing the results of evaluating apoptosis by caspase activity measurement in Test Example 4. [Figure 5] Figure 5 is a graph showing the results of evaluating apoptosis by caspase activity measurement in Test Example 4.

Modes for Carrying Out the Invention

[0015] [Composition for Suppressing the Growth of Cancer Cells] In the present invention, the composition for suppressing the growth of cancer cells contains a Japanese knotweed (Fallopia japonica) extract as an active ingredient.

[0016] Japanese knotweed is a perennial plant of the genus Reynoutria in the family Polygonaceae. In addition to the whole of Japan, it is also distributed overseas in Taiwan, China, etc. It grows to 1.5 m to 2 m in a short period and grows in groups, so it may affect the vegetation of other plant species or cause damage by breaking through concrete and asphalt.

[0017] However, on the one hand, it is also called "Hu Zhang" from the tiger-striped pattern on the stem and is used in traditional Chinese medicine as a crude drug using the rhizome. In traditional Chinese medicine, the dried root is used as a laxative and for joint pain and cough suppression.

[0018] As described above, although Itadori grows wild throughout Japan, there are few regions where it is consumed as wild vegetables or traditional vegetables. Kochi Prefecture has been using Itadori as a wild vegetable since ancient times, and currently ranks first in the country in terms of the shipment volume of Itadori. Itadori is used in various dishes such as stir-fried and simmered dishes, tempura, and white dressing.

[0019] In the present invention, the applicable parts of Itadori are not limited as long as the effects of the present invention are achieved. For example, at least one selected from the group consisting of leaves, stems, and roots can be mentioned. Leaves and / or stems are preferable, and leaves are more preferable.

[0020] The production area of Itadori is not limited as long as the effects of the present invention are achieved. For example, Kochi Prefecture, Hokkaido, Akita Prefecture, etc. can be mentioned. It is preferably produced in Kochi Prefecture, Hokkaido, or Akita Prefecture, more preferably produced in Kochi Prefecture or Hokkaido, and particularly preferably produced in Kochi Prefecture.

[0021] Although not limited, from the viewpoint of significantly achieving the effects of the present invention, in the case of Itadori produced in Kochi Prefecture, for example, the harvesting period of Itadori is preferably from April to October, and more preferably from July to September. Although not limited, for example, in the case of Itadori produced in Kochi Prefecture, by harvesting from July to September, the leaves become 50 mm or more in size, which is preferable from the viewpoint of significantly achieving the effects of the present invention.

[0022] The growth period of Itadori is, for example, preferably from 2 months to 5 years, more preferably from 3 months to 4 years, still more preferably from 4 months to 3 years, particularly preferably from 5 months to 2 years, and most preferably from 6 months to 1 year.

[0023] The extract of Itadori is prepared by extracting with water, hydrous ethanol, an organic solvent, or other extraction solvents by a conventional method as long as the effects of the present invention are achieved. It may be an extract or a dried product thereof.

[0024] While not limited to these methods, it is possible to prepare an extract of Japanese knotweed by, for example, immersing it in an extraction solvent in its fresh, dried, or ground state and extracting it under optimal temperature and time conditions.

[0025] When using dried Japanese knotweed, it is preferable to dry it at 25°C to 80°C, although this is not limited to that temperature, in order to suppress the reduction of active ingredients. For example, when drying Japanese knotweed at 25°C to 45°C, it falls within the temperature range used by general low-temperature dryers, and can therefore be easily manufactured using known low-temperature dryers.

[0026] Dried Japanese knotweed can be frozen before being dried using a drying device. Freezing allows for long-term storage in a frozen state and also enables efficient extraction of active ingredients.

[0027] As described in the examples below, it has been found that the extract of Japanese knotweed contains at least five components: neochlorogenic acid, chlorogenic acid, piceatannol, rutin, and quercetin.

[0028] The temperature conditions in the extraction described above are not limited as long as they achieve the effects of the present invention, but for example, they may be at room temperature or heated. When extraction is performed by heating, the extraction temperature is preferably 30°C to 100°C, more preferably 40°C to 100°C, even more preferably 50°C to 90°C, particularly preferably 50°C to 80°C, and most preferably 60°C to 80°C.

[0029] The time conditions in the above extraction are not limited insofar as they achieve the effects of the present invention, but the extraction time is preferably 10 minutes to 24 hours, more preferably 30 minutes to 10 hours, even more preferably 1 hour to 6 hours, and particularly preferably 2 hours to 4 hours.

[0030] [Application] In the present invention, the inhibitory effect on cancer cell proliferation can be evaluated by the method described in the examples below. While not limited to these methods, the effect on cancer cells can be evaluated using indicators such as cancer cell viability, morphological changes in cancer cells, DNA fragmentation of cancer cells, or caspase activity.

[0031] Furthermore, as described in the examples below, it has been shown that Japanese knotweed extract induces apoptosis in cancer cells by fragmenting DNA fragments in cancer cells and activating caspase 3 and caspase 8, which are regulators of apoptosis. Therefore, in another embodiment, the present invention can also be used as a composition for promoting apoptosis in cancer cells, containing Japanese knotweed extract as an active ingredient. In this embodiment, the raw materials, extraction method, dosage form, etc., of Japanese knotweed are the same as those used when it is used as a composition for inhibiting the proliferation of cancer cells.

[0032] The types of cancer (cancer species) are not particularly limited, and include brain tumors, head and neck cancers, salivary gland cancers, thyroid cancers, lung cancers, small cell lung cancers, breast cancers, mesotheliomas, pancreatic cancers, liver cancers, biliary tract cancers, esophageal cancers, stomach cancers, GISTs, small intestine cancers, colorectal cancers, kidney cancers, renal pelvis cancers, ureteral cancers, bladder cancers, prostate cancers, cervical cancers, ovarian cancers, uterine sarcomas, malignant lymphomas, leukemias, chronic lymphocytic leukemia (CLL), multiple myelomas, skin cancers, melanomas (malignant melanomas), sarcomas, etc. Although not limited, from the viewpoint of significantly demonstrating the effects of the present invention, it is preferable that the present invention be applied to at least one type of digestive system cancer selected from the group consisting of colorectal cancer, small intestine cancer, esophageal cancer, stomach cancer, and GISTs, more preferably to colorectal cancer and / or stomach cancer, and even more preferably to colorectal cancer.

[0033] The compositions of the present invention can be prepared and processed as pharmaceuticals, quasi-drugs, cosmetics, food and beverages, animal feed, or pet food, but are not limited to these.

[0034] For example, in the food and beverage sector, these can be used as health foods, functional foods, foods for the sick, and foods for specified health uses. Such foods and beverages may be marketed to suppress the onset of cancer, inhibit its progression in cancer prevention, or make similar claims, even if they do not lead to the development of a disease. Furthermore, these foods and beverages may be offered as products aimed at individuals concerned about developing cancer or those who wish to suppress cancer cells.

[0035] Pharmaceuticals, health foods, functional foods, foods for the sick, and foods for specified health uses can be used in various formulations, such as solid preparations (tablets, granules, fine granules, powders, capsules (soft capsules, hard capsules, seamless capsules, etc.), chewable tablets, etc.), liquid preparations (syrups, suspensions), and liquid foods. Foods in formulation form can be manufactured in the same way as known pharmaceutical preparations, by mixing the active ingredient with a carrier acceptable as a pharmaceutical or food, such as a suitable excipient, and then manufacturing it using conventional methods.

[0036] For example, tablets can be prepared by mixing a powdered active ingredient with a pharmaceutically acceptable carrier component (such as an excipient) and then compression molding the mixture. Confectionery tablets, such as candies, may also be prepared by injecting the mixture into a mold. Tablets may be coated with sugar or film. Furthermore, tablets may be single-layer tablets or multi-layer tablets such as double-layer tablets.

[0037] Granules and other powdered materials may be prepared by various granulation methods (extrusion granulation, crushing granulation, dry compaction granulation, fluid bed granulation, rolling granulation, high-speed stirring granulation, etc.), and tablets can be prepared by appropriately combining the above granulation methods and tableting methods (wet tableting, direct tableting, etc.).

[0038] Capsules can be prepared by conventional methods, which involve filling capsules with powders (such as powders or granules) or liquids.

[0039] The liquid formulation can be prepared by dissolving or dispersing each component in an aqueous medium (such as purified water or purified water containing ethanol) that serves as a carrier component, filtering or sterilizing as necessary, filling it into a designated container, and sterilizing it.

[0040] Common methods for manufacturing soft capsules include the plate method, the rotary method, and the seamless method.

[0041] In rotary (punching) manufacturing, a sheet-like capsule coating sandwiches the flowing filling material and forms a capsule shape along the holes of a rotating cylindrical mold. On the other hand, in seamless (dropping) manufacturing, the capsule coating composition and the contents are simultaneously discharged from multiple concentric nozzles, forming a seamless capsule shape.

[0042] The base material for the soft capsule coating is not particularly limited, but starch, pullulan, cellulose, polyvinyl alcohol, gelatin, succinylated gelatin, etc., can be used, with starch, gelatin, and succinylated gelatin being preferred, and gelatin and succinylated gelatin being even more preferred. These may be used individually or in combination of two or more.

[0043] The preferred dosage form of the solid formulation of the present invention is a capsule or a tablet, more preferably a soft capsule or a seamless capsule, and even more preferably a soft capsule.

[0044] Furthermore, the composition of the present invention can be used to produce liquid beverages such as soups, juices, fruit juices, milk, dairy beverages, whey beverages, lactic acid bacteria beverages, tea beverages, alcoholic beverages, coffee beverages, carbonated beverages, soft drinks, water beverages, cocoa beverages, jelly beverages, sports drinks, and diet drinks, as well as semi-solid foods such as puddings and yogurts, noodles, confectionery, and spreads.

[0045] When preparing the composition of the present invention as a food or beverage, various food additives may be added. Examples of food additives include antioxidants, colorants, flavorings, seasonings, sweeteners, acidulants, pH adjusters, quality stabilizers, and preservatives.

[0046] When preparing the composition of the present invention as a pharmaceutical or quasi-drug, it can be prepared as a formulation containing an extract of Japanese knotweed, which can serve as an active ingredient, and preferably a pharmaceutically acceptable carrier. A pharmaceutically acceptable carrier generally refers to an inert, non-toxic, solid or liquid filler, diluent, or encapsulating material that does not react with the active ingredient, and examples include water, ethanol, polyols, suitable mixtures thereof, solvents or dispersion media such as vegetable oils.

[0047] Pharmaceuticals and quasi-drugs are administered orally or parenterally, for example, into the oral cavity, gastrointestinal tract, or nasal cavity. Oral formulations include solid formulations (tablets, granules, fine granules, powders, capsules, chewable tablets, etc.) and liquid formulations (syrups, suspensions, inhalants, etc.). Parenteral formulations include intravenous infusions, nasal drops, and injections.

[0048] Pharmaceuticals and quasi-drugs may further contain additives commonly used in the pharmaceutical field. Such additives include, for example, excipients, binders, disintegrants, lubricants, antioxidants, colorants, and flavoring agents, and may be used as appropriate. To enable sustained release for longer-lasting action, they may also be coated with known retarders, etc. Pharmaceuticals and quasi-drugs may further contain other additives or agents, such as antacids and gastric mucosal protectants, as needed.

[0049] Pharmaceuticals and quasi-drugs can be applied in forms such as oral compositions and oral compositions. Furthermore, pharmaceuticals and quasi-drugs may be used therapeutically or non-therapeutically.

[0050] The daily oral intake or dosage of Japanese knotweed extract for adults can be appropriately determined based on the individual's condition, weight, sex, age, activity of the material, route of intake or administration, intake or administration schedule, formulation form, or other factors. For example, the daily oral intake or dosage of Japanese knotweed extract for adults is preferably 100 mg / day or more, more preferably 200 mg / day or more, even more preferably 300 mg / day or more, particularly preferably 400 mg / day or more, and most preferably 500 mg / day or more, based on dry solids.

[0051] The daily oral intake or dosage of Japanese knotweed extract for adults is preferably 10 g / day or less, more preferably 8 g / day or less, even more preferably 6 g / day or less, particularly preferably 4 g / day or less, and most preferably 2 g / day or less.

[0052] The daily oral intake or dosage of Japanese knotweed extract for adults is preferably 100 mg to 10 g / day, more preferably 200 mg to 8 g / day, even more preferably 300 mg to 6 g / day, particularly preferably 400 mg to 4 g / day, and most preferably 500 mg to 2 g / day.

[0053] The amount of Japanese knotweed extract contained may be the amount corresponding to the above-mentioned intake or dosage. The daily oral intake or dosage for adults may be divided according to the dosage form; for example, if it is a capsule, it may be taken in 1 to 6 capsules, 1 to 4 capsules, 1 to 3 capsules, or 1 to 2 capsules.

[0054] The composition of the present invention can be taken or administered once to several times a day, usually 1 to 6 times a day, 1 to 3 times a day, 1 to 2 times a day, or at any period and interval. [Examples]

[0055] Next, the present invention will be specifically described with reference to examples, but the present invention is not limited to the following examples. In addition, unless otherwise specified, the amounts of extracts shown in the examples are on a dry solids basis.

[0056] [Sample preparation] 1. Preparation of 80% ethanol extract of Japanese knotweed leaves It is made from Japanese knotweed leaves harvested in July from Kochi Prefecture. 287 g of cold-dried Japanese knotweed leaves were crushed and stirred overnight in 13 L of 80% ethanol. The mixture was then centrifuged at 3000 rpm for 10 minutes, and the supernatant was collected as the extract. The extract was filtered and concentrated using an evaporator. The concentrated extract was freeze-dried to obtain 102.67 g of extract. 2. Preparation of other samples The reagents used were neochlorogenic acid (manufactured by Tokyo Chemical Industry Co., Ltd.), piceatannol (manufactured by Tokyo Chemical Industry Co., Ltd.), rutin (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.), and quercetin (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.).

[0057] [Investigation of the inhibitory effect of Japanese knotweed extract on the proliferation of colorectal cancer cells] The samples, cancer cell models, culture media, and culture conditions used are as follows: Additives: Dried Japanese knotweed leaf ethanol extract, neochlorogenic acid, piceatannol, rutin, quercetin Cancer cell model: Colon-26, a mouse-derived colorectal cancer cell line. Culture medium: RPMI 1640 medium containing 10% neonatal bovine serum and antibiotics. Culture conditions: 37℃, CO2 concentration 5%

[0058] [Test Example 1. Determination of cell viability using the WST-8 method] 2 x 10 in a 96-well plate 4Colon-26 cells were seeded and cultured for 48 hours. After 48 hours, Japanese knotweed extract, neochlorogenic acid, piceatannol, rutin, and quercetin were added to the culture medium to reach the specified concentration. Culture was then continued for another 20 hours. During this time, dimethyl sulfoxide (DMSO) was used as the dilution solution for the extract, ensuring that the DMSO concentration in the culture medium remained at 0.5%. After 20 hours, 10 μl of Cell Counting Kit-8 (DOJINDO) was added and reacted for 4 hours. After 4 hours, the absorbance at 450 nm was measured using a microplate reader. The absorbance was compared to the control group (no additives) using the Dunnett method (the same method was used in the following test examples).

[0059] The results are shown in Figure 1. The viability of Colon-26 cells decreased in a concentration-dependent manner in the presence of Japanese knotweed extract (Figure 1). A decrease in cell viability was observed from the addition of 500 μg / ml, and the IC50 value was 1.18 mg / ml. At the same time, the proliferation of Colon-26 cells decreased in a concentration-dependent manner in the presence of various polyphenols. The IC50 values ​​for proliferation inhibition were 7.37 mg / mL for neochlorogenic acid, 5.87 mg / mL for piceatannol, 7.41 mg / mL for rutin, and 6.46 mg / mL for quercetin.

[0060] [Experiment Example 2. Observation of Cell Morphology] 2 x 10 on a 24-hole plate 5 Cell Colon-26 cells were seeded and pre-cultured for 24 hours. After 24 hours, Japanese knotweed extract was added to the culture medium to achieve a concentration of 500 μg / ml or 1000 μg / ml. The control group was cultured in a medium containing 0.5% DMSO. Cell morphology was observed immediately after the addition of Japanese knotweed extract and 4 hours later.

[0061] The results are shown in Figure 2. Morphological changes in Colon-26 cells were observed upon addition of Japanese knotweed extract, resulting in cell atrophy and deformation (Figure 2). In the presence of 500 μg / ml of Japanese knotweed extract, significant cell contraction was observed 4 hours after addition. In the presence of 1000 μg / ml of Japanese knotweed extract, the morphological changes were even more pronounced, with aggregation and deformation observed.

[0062] [Test Example 3. Evaluation of DNA fragmentation by Japanese knotweed extract using agarose electrophoresis] 2 x 10 in a petri dish 6 Colon-26 cells from Cell were seeded and pre-cultured for 24 hours. After 24 hours, Japanese knotweed extract was added to the culture medium to achieve concentrations of 250 μg / ml, 500 μg / ml, or 1000 μg / ml of neochlorogenic acid, piceatannol, rutin, and quercetin, and the cells were cultured for 2 hours. The control group was cultured in a medium containing 0.5% DMSO. After 2 hours of culture, 2 × 10⁶ cells were cultured. 6 Cells were collected and DNA was isolated. The Apototic DNA-Ladder Kit (Roche) was used for DNA isolation. The isolated DNA was subjected to electrophoresis on a 3% agarose gel containing 1 μg / mL of ethylenedium bromide, and then irradiated at 254 nm to evaluate fragmentation.

[0063] The results are shown in Figure 3. Agarose electrophoresis revealed clear bands at bp, 3,000 bp, and 900 bp in DNA from normal Colon-26 cells (Figure 3). In the presence of 250 μg / ml of Japanese knotweed extract, the 900 bp band was enhanced, and new bands appeared below 900 bp, confirming DNA fragmentation. Furthermore, in the presence of 500 μg / ml of Japanese knotweed extract, the bands below 900 bp became even clearer. On the other hand, in the presence of 1000 μg / ml of Japanese knotweed extract, bands other than 3000 bp became fainter.

[0064] [Test Example 4. Evaluation of Apoptosis by Measurement of Caspase Activity] 1 x 10 in a 96-well plate 4Colon-26 cells were seeded and pre-cultured for 24 hours. Subsequently, Japanese knotweed extract and various polyphenols (neochlorogenic acid, piceatannol, rutin, quercetin) were added to the culture medium to a concentration of 250 μg / ml and cultured for 24 hours. After 24 hours, Caspase-Glo® 3 / 7 Reagent (Promega) was added and reacted at room temperature for 1 hour. The control group was subjected to the same procedure without the addition of Japanese knotweed extract and various polyphenols. The luminescence after 1 hour was measured using GloMax (Promega), and the relative value to the control group was evaluated.

[0065] The results are shown in Figures 4 and 5. Compared to the control, the addition of Japanese knotweed extract significantly increased caspase activity, reaching 1.55 times the activity (Figure 4). When polyphenols contained in Japanese knotweed leaves were added, neochlorogenic acid, rutin, and quercetin showed significantly increased activity, with neochlorogenic acid increasing 3.31 times, rutin 3.41 times, and quercetin 1.81 times (Figure 5).

[0066] [Discussion regarding the above results] Cell viability measurements using the WST-8 method showed that the addition of high concentrations of Japanese knotweed extract reduced the viability of Colon-26 cancer cells. In addition, cell morphology contracted in the presence of 50 μg / ml of Japanese knotweed extract, and cells aggregated, deformed, and decreased in number in the presence of 100 μg / ml, suggesting that Japanese knotweed extract has an effect of inducing cell death in cancer cells.

[0067] Cell death can be broadly divided into two categories: accidental / accidental cell death and programmed cell death. Accidental cell death includes necrosis caused by infection or injury. On the other hand, programmed cell death refers to cell death that is managed and controlled within the body, such as eliminating abnormal cells to maintain the individual in a better state, and is also called apoptosis. Therefore, if the cell death caused by the Japanese knotweed extract was necrosis, it would mean that it was toxic to the cells. Conversely, if it was apoptosis, it could be said that it promoted physiological functions. Therefore, apoptosis was evaluated using a DNA ladder assay and caspase activity measurement.

[0068] Apoptosis is a pre-programmed form of cell death in living organisms, and therefore its process is regular, with regularity also observed in the DNA cleavage that occurs during this process. As a result, DNA fragments of a certain size are observed. In necrosis, however, uncontrolled cell death progresses due to physiological functions, resulting in random DNA cleavage, or even cleavage so severe that DNA detection is difficult. The DNA ladder assay is a method for confirming the presence or absence of these DNA fragments.

[0069] DNA ladder assay results showed that in the presence of 25 μg / ml of Japanese knotweed extract, large DNA molecules decreased, bands representing small DNA molecules were enhanced, and new bands appeared. Furthermore, the detection of these new bands was enhanced in the presence of a higher concentration of 50 μg / ml, suggesting that apoptosis was induced and promoted.

[0070] Caspases are a family of proteases that play a central role in numerous processes, including cell death and inflammation, during the progression of apoptosis. Caspase-3 plays a crucial role in initiating apoptosis within cells. Caspase-3 and caspase-7 share the same target substrate sequence, making it difficult to distinguish their cleavage activity in vitro. Therefore, we measure the activity of either Caspase-3 or Caspase-7 by adding common substrates for both. This assay system ensures the specific measurement of Caspase-3 or Caspase-7 activity in apoptotic samples. Other caspases and nonspecific proteases are not detected.

[0071] Measurements of caspase activity showed that the addition of 25 μg / ml of Japanese knotweed extract increased caspase activity by 1.5 times, suggesting that the extract induces apoptosis. Activity measurements using polyphenols contained in the Japanese knotweed extract suggested that neochlorogenic acid and rutin particularly strongly activate caspases. Since neochlorogenic acid and rutin are also abundant in the polyphenol content of Japanese knotweed leaf extract, it is suggested that these two polyphenols are the main components responsible for apoptosis induction by the extract. However, the effects of these two components alone cannot explain the phenomenon, and since Japanese knotweed leaf extract contains various components, the contribution of other unidentified components is also expected.

Claims

1. A composition for inhibiting the proliferation of cancer cells, containing an extract of the leaves of Japanese knotweed (Fallopia japonica) as an active ingredient, A composition in which the extract of the leaves of Japanese knotweed is an extract of water or aqueous ethanol.

2. A composition for promoting apoptosis of cancer cells, containing an extract of Japanese knotweed leaves as an active ingredient, A composition for promoting apoptosis of cancer cells, wherein the extract of the leaves of Japanese knotweed is an extract with water or aqueous ethanol.

3. The composition according to claim 1 or 2, wherein the extract of the leaves of Japanese knotweed is an extract of 80% ethanol.

4. The composition according to any one of claims 1 to 3, wherein the extract of the leaves of Japanese knotweed contains at least one selected from the group consisting of piceatannol, rutin, and quercetin.

5. The composition according to any one of claims 1 to 4, wherein the extract of the leaves of Japanese knotweed contains neochlorogenic acid and rutin.

6. The composition according to any one of claims 1 to 5, wherein the cancer cells are colon cancer cells.

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