JP2006528174A - Aged kyoto extract prevents metastasis and cancer formation - Google Patents

Abstract

The present invention relates to an extract from a long-lived drug that has an anticancer effect and a cancer metastasis-inhibiting effect, and more particularly to an extract from a long-lived drug that has an inhibitory effect on cancer metastasis, cell adhesion, cell invasion, and cell proliferation. The long-life Doraj extract of the present invention suppresses tumor formation and cancer metastasis. Therefore, an anticancer agent containing the long-lived Doraj extract of the present invention as an active ingredient will be useful for the prevention and treatment of cancer.

[Selection] Figure 1

Description

  The present invention relates to an extract from a long-lived drug that has an anticancer effect and a cancer metastasis-inhibiting effect, and more particularly to an extract from a long-lived drug that has an inhibitory effect on cancer metastasis, cell adhesion, cell invasion, and cell proliferation.

  The roots of the asteraceae (Scientific name: Platycodon grandiflorum A.DC) have a main root length of about 10 to 15 cm, a root diameter of 1 to 3 cm, a brownish-white to milky white color, and the upper part branches irregularly. There is a part to be performed, and there are wrinkles in the longitudinal direction and shells and wrinkles in the transverse direction. In addition, they are hard but inherently non-fibrous, so they are easy to break, and have a slight aroma and bitterness (Luc, Chang-soo et al., “Modern Pharmacognosis”, Hakuchangsa, Seoul, p.460-461. 1993 (Ryuk, Chang-Soo, et al., Modern Pharmacognosy, Hackchangsa Co., Seoul, 460-461, 1993)).

  As the main pharmaceutical ingredient of fennel, there are triterpene saponins (platicodine A, C, D, D2 and D3), which constitute about 2% of the root. According to animal studies, these pharmaceutical ingredients have various effects such as antitussive and expectorant effects, central nervous system inhibitory effects (sedation, pain relief and antipyretic), anti-inflammatory action against acute chronic inflammation, anti-ulcer action, gastric juice It has been confirmed that it has a secretory inhibitory action, an anticholinergic action that lowers blood pressure by vasodilation, a blood glucose lowering action, and a cholesterol metabolism promoting action (“Chemical & Pharmaceutical Bulletin” No. 20, p. 1952, 1972; “Chemical & Pharmaceutical Bulletin” 23, p. 2965, 1975; “Journal of Chemical Society, Perkin Transaction I” p. 661, 1984; “Comprehensive Medicine” 3 1, 1951; "Journal of Pharmaceutical Society of Colli 19, 164, 1975 (Chem., Pharm. Bull., 20, 1952 (1972); Chem. Pharm. Bull., 23, 2965 (1975); J. Chem. Soc., Perkin trans I, 661 (1984); Sogoigaku, 3,1 (1951); J. Pharm., Soc. Kor., 19, 164 (1975))). Other pharmaceutical ingredients isolated from Roots include steroidal compounds such as α spinasterol, stigmast-7-enol, and α spinasterol glucoside, and polysaccharides such as inulin and betulin.

  In the present specification, the term “long-life doraji” is the name of the root of more than 20 years old, but despite its various pharmacological effects, it has been difficult to grow long-lived kyoto. However, a technology that enables the cultivation of long-grown doraji, which is more than 20 years old, has recently been developed (Korea Patent No. 045731, Lee, Sung-Ho), and the name of the invention “Method for Produce”. Sing Perennial Balloon Flowers (a method for producing perennial balloonflowers)), which has enabled mass production of long-lived drage. Such long-lived doraji is superior in pharmacological effect and physiological action as compared with ordinary root roots cultivated for 2 to 4 years.

  The research on the long-term Draj has so far been a study on anti-diabetic activity (Journal of the Korean Society of Food Science and Nutrition No. 25, p. 986, 1996 (J. Korean Soc. Food Sci. Nutri., 25, 986 (1996)), and studies on antihyperlipidemia (Journal of Pharmaceutical Society of Japan No. 93, 1973, p. 1188 to 1194 (J. Pharmaceutical Society of Japan 93 (1973), 1188-1194)), and research on the prevention and control of liver damage has also been conducted. Ethanol extract suppresses mold aplatoxin, inulin fraction has phagocytosis and potent anti-tumor activity against sarcoma, solid tumor, and kyoto root These 40% extracts are known to have an alcohol absorption inhibitory effect (“Mycopathologia”, 66, p. 16, 1979, “Biopharmaceutical Journal”, 40, p. 367, 1986). "Biopharmaceutical Journal" No. 40, p. 375, 1986, Japanese Unexamined Patent Publication No. 60-89427, 1985 (Mycopathologia, 66, 16 (1979), Shoyakugaku Zasshi, 40, 367 (1986), Shoyakugaku Zasshi, 40, 375 (1986), Japanese Patent No. 60-89427 (1985)).

  Malignant tumors (cancer) that threaten human health are the first or second cause of death in the world and the most common cause of death after cardiovascular disease in Western society. In modern society, lung cancer is increasing due to aging, increasing smoking population and air pollution. In addition, colon cancer, breast cancer, prostate cancer, and the like are also increasing due to the intake of high-fat foods, an increase in environmental pollutants, increased alcohol consumption, and the like of Western diets. Therefore, there is an urgent need to develop anticancer substances that enable early prevention and treatment of cancer, thereby helping to promote human health, improve the quality of healthy life, and promote human health management.

  Drugs developed so far for cancer treatment are systemically treated by injection or oral administration through the bloodstream to the whole body, so the level of specificity for cancer cells is low. Therefore, the administration of such drugs has systemic side effects, and has the disadvantages of showing stronger side effects than surgery or radiation therapy. Existing anticancer drugs are administered at intervals of 3-4 weeks every half year to one year for 3-5 days each week, which causes various side effects due to anticancer drugs and often results in discontinuation of treatment. ing. Existing chemotherapy shows a medical effect by directly attacking cancer cells, but has very strong side effects and is nonspecific. Despite these shortcomings, innovative new drugs have not appeared on the market in recent years.

  An object of the present invention is to provide a therapeutic agent that suppresses cancer cell metastasis and tumor formation, and the therapeutic agent contains an extract from Nagase Doraj, a safe and non-toxic natural substance, as an active ingredient. .

  In order to achieve the above-mentioned object, the present invention provides a drug that suppresses tumor formation and cancer metastasis, and the drug contains an extract from Chosei Doraj as an active ingredient.

  Hereinafter, the present invention will be described in detail.

  The present inventors have conducted extensive research to develop drugs that suppress cancer metastasis and tumor formation using safe and non-toxic natural substances, and as a result, extracts from long-lived doraj have been extracted from cancer cells. (Melanoma) was found to suppress lung metastasis, cell adhesion, cell invasion and cell proliferation. Based on this discovery, the present invention has been completed.

  The long-grown doraj extract according to the present invention is produced by any method conventionally used in the art. The long-life Doraj extract is preferably a hot water extract or an organic solvent extract, which can be prepared alone or mixed with a pharmaceutically acceptable herbal material. When extracting the long-life drage, it is preferable to use the long-life draj powder, but the powder is dried to a moisture content of less than 5%, and the dried material is powdered to obtain a particle size. Is a powder prepared by sorting out powders of less than 0.6 mm.

  The hot water extract is preferably prepared by adding about 5 to 10 times the volume of water to the long-life Doraj powder and extracting the solution twice at a temperature of 90-100 ° C. for 4-6 hours. . The resulting filtrate is used alone or mixed with one selected from acceptable herbal materials or herbal extracts.

  The organic solvent extract is preferably added about 3 times the volume of organic solvent to the long-life Doraj powder, the solution is extracted twice at room temperature, and the resulting filtrate is concentrated under reduced pressure and then acceptable A selection of fresh herbal materials or herbal extracts is prepared by adding to this concentrate. As organic solvent, it may be preferred to use an alcohol, preferably methanol or ethanol.

  The long-life Doraj extract of the present invention may be formulated by known methods using food hygiene acceptable or pharmaceutically acceptable excipients, thereby preparing a pharmaceutical formulation. When preparing a pharmaceutical preparation, the long-lived Doradi extract as an active ingredient is preferably mixed or diluted with a carrier, or enclosed in a container-like carrier. When a carrier is used as a diluent, the carrier may be a solid, semi-solid or liquid material that acts as a vehicle, excipient or medium. As such, the formulation may be in the form of tablets, pills, powders, elixirs, suspensions, emulsions, syrups, aerosols, soft or hard capsules, sterile injections, sterile powders and the like. Examples of suitable carriers, excipients or diluents include lactose, glucose, sucrose, sorbitol, mannitol, calcium silicate, cellulose, methylcellulose, amorphous cellulose, polyvinylpyrrolidone, water, methyl hydroxybenzoate, hydroxybenzoic acid Examples include propyl, talc, magnesium stearate, and mineral oil. The preparation may contain fillers, anti-aggregating agents, lubricants, wetting agents, fragrances, emulsifiers, preservatives and the like. An angiogenesis inhibitor comprising the long-lived Doraj extract of the present invention can be formulated by any method well known in the art to release an active ingredient rapidly, continuously or delayed after administration to a mammal. is there.

  The angiogenesis inhibitor contained as an active ingredient in the long-life Doraj extract of the present invention can be administered via various routes, oral, transdermal, subcutaneous, intravenous, and intramuscular routes depending on the purpose. . For humans, the daily dose of the extract of the present invention may be usually 10 to 1,000 mg / kg body weight, preferably 10 to 100 mg / kg body weight, and administered once or several times. Also good. However, the actual dosage of the extract of the present invention should be determined in view of various factors including the route of administration, patient age, gender and weight, health status, disease severity and the like.

  Hereinafter, the present invention will be described in more detail by way of examples and test examples. However, it should be understood that these examples are presented for the purpose of illustrating a preferred embodiment of the invention and are not intended to limit the scope of the invention.

Example 1 Production of Hot Water Extract from Chosei Doraji 2 liters of distilled water was added to 0.2 kg of Chosei Doraji powder and the solution was extracted twice at 90-95 ° C. for 5 hours and filtered. The filtrate was lyophilized to obtain 100 g of long-life Doraj extract.

  The following tests were conducted to examine the inhibitory effect of the drug of the present invention on lung metastasis, cell adhesion, cell invasion, and cancer cell proliferation of melanoma cancer cells.

Test animals and treatment Reagents The following reagents were purchased and used.
B16F10 cell line was purchased from Korean Cell Line Bank, and Matrigel, Eosin, Gelatin, EMEM medium, and fetal bovine serum were purchased from Sigma Chemical Co.

2. Animals As test animals, 8 weeks old C57BL / 6 mice weighing 25-30 g were used, and the mice were allowed to freely feed (Purina, Korea) and water. The animals were housed under a cycle of temperature 21-24 ° C., relative humidity 40-80%, 12 hours light / 12 hours dark.

Test Example 1: Inhibitory effect of long-life Doraj extract on lung metastasis of melanoma cancer cell line B16F10 in C57BL / 6 mice Test method In order to examine the inhibitory effect of Nagasei Doraj extract on cancer metastasis, metastatic melanoma cancer cells B16-F10 were injected into the tail vein of C57BL / 6 test animals. Next, the long-life Doraj extract was orally administered to the test animals, and the number of colonies of melanoma cancer cells B16-F10 that had subsequently metastasized to the lungs was counted by hematoxylin-eosin staining. The results are shown in Table 1 below.

Table 1

  FIG. 1 is a graph showing the number of colonies that have metastasized to the lungs according to the treatment concentration of the long-life Doraj extract. The upper part of FIG. 2 is a photograph showing a colony of melanoma cancer cells B16-F10 that has metastasized to the lung, taken after staining with hematoxylin-eosin. The lower part of FIG. 2 is a photograph showing the result of staining melanoma cell tissue with hematoxylin-eosin after embedding in paraffin. As can be seen from Table 1 and FIG. 1, melanoma cells B16F10 were administered to C57BL / 6 mice, long-lived Doraj extract of each concentration was administered to the mice for 14 days, and the number of melanoma B16F10 cells that had metastasized to the lungs was determined. As a result of the counting, it was shown that the number of colonies that had metastasized to the lung decreased according to the treatment concentration of the long-life Doraj extract.

Test Example 2: Inhibitory effect of long-life Doraj extract on cell adhesion of melanoma cancer cell B16F10 Test method In order to examine the cell adhesion inhibitory effect of long-life Doraj extract, metastatic melanoma cancer cells B16F10 were subcultured in a 48-well plate coated with gelatin, and then each concentration of sample was placed in a well plate. It was. After 18 hours, cell adhesion was observed under a microscope, whereby the number of adhered cells was counted. The result is shown in FIG.

  FIG. 3 is a graph showing cell adhesion suppression according to the treatment concentration of the long-life Doraj extract. As shown in FIG. 3, cell adhesion inhibition increased with the treatment concentration of the long-life Doraj extract.

Test Example 3: Inhibitory effect of long-life Doraj extract on cell infiltration of melanoma cancer cell B16F10 Test method In order to examine the cell infiltration inhibitory effect of the long-life Doraj extract, metastatic melanoma cancer cells B16F10 were subcultured in a 48-well plate coated with matrigel, and then each concentration of sample was placed in a well plate. . After 18 hours, cell invasion was observed under a microscope and the number of infiltrated cells was counted. The result is shown in FIG.

  FIG. 4 is a graph showing the number of infiltrated cells according to the treatment concentration of the long-life Doraj extract. As shown in FIG. 4, the number of infiltrated cells decreased according to the treatment concentration of the long-life Doraj extract, suggesting that the long-life Draj extract has an inhibitory effect on the invasion of the melanoma cancer cell line B16F10. .

Test Example 4: Inhibitory effect of long-life Doraj extract on proliferation of melanoma cancer cell B16F10 Test Method In order to examine the inhibitory effect on the cancer cell proliferation of the long-life Doraj extract, metastatic melanoma cancer cells B16F10 were subcultured in a 48-well plate, and samples of each concentration were placed in the well plate. After 24 hours, the number of proliferated cells was counted by the MTT method. In the MTT method, the concentration of formazan is measured based on the fact that mitochondrial enzymes in the cells reduce MTT to produce formazan. The result is shown in FIG.

  FIG. 5 is a graph showing suppression of cancer cell growth according to the treatment concentration of the long-life Doraj extract. As shown in FIG. 5, cell proliferation of the melanoma cancer cell line B16F10 was suppressed according to the treatment concentration of the long-life Doraj extract.

  As described above, the effect of suppressing the pulmonary metastasis of the long-life Doraj extract of the present invention was examined in an in vivo animal test of C57BL / 6 mice in which melanoma cancer cell line B16F10 was administered via the tail vein to induce pulmonary metastasis. . As a result, it was shown that lung metastasis decreased according to the treatment concentration of the long-life Doraj extract. In addition, the inhibitory effect of the long-life Draj extract on cell adhesion was also examined in an in vivo test using the melanoma cancer cell line B16F10. As a result, the cell adhesion of the melanoma cancer cell line B16F10 depends on the treatment concentration of the long-life Draj extract. It was shown that it decreased. Furthermore, in order to examine the suppression mechanism of the long-lived Doraj extract on cell adhesion and invasion of the melanoma cancer cell line B16F10, the proliferation of the melanoma cancer cell line B16F10 against the long-lived Doraj extract was examined. Decreased according to the treatment concentration of Chosei Doraj extract.

Test Example 5: Acute toxicity test Oral administration 50 ICR mice weighing 25 g were divided into 5 groups of 10 mice each. The long-life Doraj extract of the present invention was orally administered to the groups of 5 animals at 1, 5, 25, 125 and 625 mg / kg, respectively. Thereafter, toxicity was observed over 2 weeks. The results showed that none of the 5 groups died.

2. Abdominal administration 40 ICR mice weighing 25 g were divided into 4 groups of 10 mice each. The long-life Doraj extract of the present invention was administered to the abdominal cavity of the four animal groups at 0.2, 2, 20, and 200 mg / kg, respectively. Thereafter, toxicity was observed over 24 hours. The results showed that none of the four groups died.

  As a result, it was confirmed that the long-life Doraj extract of the present invention has little or no acute toxicity.

  In the following formulation examples, a formulation of an anticancer agent containing the long-life Doraj extract of the present invention as an active ingredient will be described. However, as a matter of course, the anticancer agent of the present invention is not limited by these preparation examples.

Formulation Example 1: Tablet preparation Chosei Doraji extract 50mg
Lactose 20mg
Starch 20mg
Magnesium stearate appropriate amount The above-mentioned composition containing the long-grown doradi extract of the present invention as an active ingredient was tableted by a conventional tableting method, and a tablet was prepared.

Formulation Example 2: Preparation of capsule preparation Nagai Doraji extract 50 mg
Lactose 20mg
Starch 20mg
Talc 1mg
Magnesium stearate appropriate amount Gelatin capsules were filled with the above-mentioned composition containing the long-grown Doradi extract of the present invention as an active ingredient by a conventional method to prepare a capsule preparation.

  As described above, cancer metastasis and tumor formation can be suppressed by the therapeutic agent of the present invention containing, as an active ingredient, the long-life Doraj extract of the present invention, which is a safe and non-toxic natural substance. Therefore, the therapeutic agent of the present invention will be clinically useful for cancer treatment and prevention.

  As described above, the composition containing the long-lived Doraj extract of the present invention that suppresses cancer metastasis and tumor formation will be useful as a functional food and a pharmaceutical product.

It is a graph which shows the colony number which metastasized to the lungs according to the treatment density | concentration of a long life Doraj extract. It is a photograph which shows the colony of the melanoma cancer cell B16-F10 which metastasized to the lung after dyeing | staining with hematoxylin-eosin. It is a graph which shows cell-adhesion suppression according to the process density | concentration of Nagasei Doraj extract. It is a graph which shows the cell number which infiltrated according to the process density | concentration of long-life Doraj extract. It is a graph which shows cancer cell growth suppression according to the process density | concentration of a long-life Doraj extract.

Claims (2)

  A drug that suppresses tumor formation and cancer metastasis, comprising an extract from Nagasei Doraj as an active ingredient. The extract according to claim 1, wherein the extract from the long-life doraj is a hot water extract or an organic solvent extract.