KR20080056462A - Pharmaceutical composition for prevention, treatment and inhibition of cancer comprising prunella vulgaris extract as an effective component - Google Patents

Abstract

A composition comprising an extract of Prunella vulgaris is provided to enhance the activity of immune system such as macrophages, spleen lymphocytes, natural killer cells or lymphokine activated killer cells and show the inhibitory effect on growth of cancer cells and metastasis, thereby being used to prevent, treat and inhibit cancer diseases excellently with minimized side effects. A composition for preventing, treating and inhibiting cancer diseases comprises an extract of Prunella vulgaris as an effective ingredient. A pharmaceutical formulation comprises the composition, an excipient, a diluent and other crude drugs. A functional food for preventing, improving and inhibiting the cancer diseases comprises the composition and a sitologically acceptable additive. Further, the cancer is ascites carcinoma, solid cancer or lung cancer.

Description

Pharmaceutical composition for prevention, treatment and inhibition of cancer comprising Prunella vulgaris extract as an effective component

Figure 1 is a graph measuring the cytotoxicity in macrophages according to the concentration of the extract.

Figure 2 is a graph showing the phagocytic activity of macrophages in accordance with the concentration of the extract.

Figure 3 is a graph showing the amount of Nitric oxide (NO) production of macrophages according to the concentration of the extract.

Figure 4 is a graph showing the proliferative capacity of spleen lymphocytes according to the concentration of Hachocho extract.

5 is a graph showing the proliferative capacity of splenic B lymphocytes according to the concentration of Hachocho extract.

Figure 6 is a graph showing the proliferative capacity of splenic T lymphocytes according to the concentration of Hachocho extract.

Figure 7 is a graph showing the cancer cell killing ability of natural killer cells (NK cells) according to the concentration of the vinegar extract.

Figure 8 is a graph showing the cancer cell killing ability of lymphokine active killer cells according to the concentration of the extract.

Field of technology

The present invention relates to a composition for preventing, treating and inhibiting cancer diseases comprising the extract of Hagocho as an active ingredient, and in particular, exhibits the effect of inhibiting the growth or inhibiting cancer metastasis of cancer cells, macrophages, spleen lymphocytes, natural killer cells Or it is characterized by enhancing the immune system activity of lymphokine active killer cells.

Prior art

Unlike most other organs, the immune system is not present in any part of the body, and the host's organs (thymus, spleen, bone marrow, lymph nodes, etc.) and immune-related cells (granulocytes) resist the invasion of various foreign objects and pathogens. , Macrophages, lymphocytes, natural killer cells, etc.), and a variety of factors (complement, interferon, etc.) are very complex lines [Playfair, JH, Essays Fundam Immunol., 1: 44-56, 1973].

The spleen, a peripheral lymphoid organ, is a lymphoid tissue formed on the side of the stomach at 5 weeks of birth. Except for red blood cells in the spleen, spleen cells are called lymphocytes (T cells, B cells), macrophages, and white blood cells. Etc.

T cells are involved in delayed hypersensitivity reactions, resistance to bacterial or viral infections, rejection of organ transplants, and immune responses to tumors. Another function of T cells is to regulate immune responses directed by other cells, such as B cells and macrophages (Ritzmann et al., Ann Allergy, 3: 109-125, 1973). T cells have surface markers and two subtypes that function differently from each other. Among them, CD4 + T cells are helper T cells (Th cells) and act as cofactors for the production of antibodies by B cells and the generation and proliferation of cytotoxic T cells (Tc cells). As described above, helper T cells promote the activation of other lymphocytes and produce various cell activators, and Th1 cells cause delayed hypersensitivity or TN responses depending on the type of cell activator produced. selectively produce -γ and IL-2, and Th2 cells produce IL-4, IL-5, IL-6 and the like, thereby promoting the response of B cells to antigens [Feldman and Globerson, Curr Top Dev Biol] , 8: 1-40, 1974. CD8 + T cells, on the other hand, are cytotoxic T cells that are known to act directly on cells such as rejection of allograft antigens, destruction of virally infected cells, and cancer cell injury.

B cells are distributed in lymphoid tissues such as blood, bone marrow and spleen and lymph nodes and are involved in humoral immune responses (Weissman et al., Ser Haematol., 4: 482-504, 1974). B cells have immunoglobulins on their cell surface and differentiate into plasma cells by antigen, producing and secreting antibodies such as IgM, IgG, IgA, IgD, and IgE [Kramer TT, Am J Vet Res., 4: 492-493, 1975.

Natural killer cells (NK cells) are morphologically large granular lymphocytes and are found in blood and lymphoid tissues [Melchers and Corbel, Ann. Immunol., P 63-73, 1983]. These cells are capable of recognizing and killing tumor cells and virus-infected cells without being stimulated and secreting several cytokines when activated [Unanue et al., Am. J. Pathol., 2: 465-478, 1976.

In addition, the reticuloendothelial system (RES) is a general term of a phagocytic system that can detect a microorganism, a pigment, or a particle when a foreign substance invades in a living body, and is a macrophage system or a mononuclear phagocytic system, MPS) is synonymous with this [Kamo and Friedman, Adv. Cancer Res., 25: 271-321, 1977.

Macrophage (macrophage) is called by various names because there is a difference in the cell type according to the type of organ or tissue that the cell is distributed. Examples include monocytes in the blood, histocytes in connective tissue, kuffer cells in the liver, alveolar macrophage in the lungs, macrophages in lymphoid tissues, and osteoclasts in bone tissues. et al., Semin. Hematol., 4: 252-282, 1979]. Macrophages function to exclude foreign bodies [Van Rooijen N., Res. Immunol., 4: 328-330, 1991], but also plays an important role in the establishment and expression of immunity [Barr, R.D., Scott. Med. J., 4: 267-272, 1979. The role of macrophages in immune response is strongly activated by the role of antigen presenting cells (APCs) in lymphocytes and by lymphocytes, the products of T cells in cellular immune responses. Humphrey, JH, Ciba. Found. Symp., 71: 287-298, 1980] and secretion of soluble factors (monokine) that affect lymphocyte growth and function [Kraal G., Int Rev Cytol., 132: 31-74, 1992].

Cancer is a disease that is hard to cure and excessively expensive medical cost is one of the various diseases that occur when the immune function is reduced as described above, it will be the most effective way to prevent cancer by enhancing the immune activity to prevent cancer in advance.

Cancers generally involve a very complex carcinogenic process of initiation, promotion, progression and metastasis. Among them, cancer metastasis is known to be the fatal cause of cancer mortality rate among the human disease. The cancer metastasis process involves metastatic malignant cancer cells infiltrating other new tissues at the first site of cancer, forming secondary tumors. There are three main stages: adhesion, invasion, and angiogenesis. Consists of. Angiogenesis, the most important factor influencing tumor growth and metastasis, involves processes of degradation of the basement membrane and extracellular matrix, migration, proliferation, substrateization and maturation of endothelial cells, and the balance between angiogenesis and inhibitors. Is adjusted. These factors directly stimulate the secretion of enzymes that act on endothelial cells or promote angiogenesis, and indirectly stimulate enzymes that dissolve extracellular matrix by stimulating stromal cells.

Although many developed countries have invested enormous research funds to study the mechanism and treatment of cancer, cancer remains a refractory disease and various treatments have been shown to cause side effects [Goodman et al., Cancer Res., 9: 2295-2304, 1987. Accordingly, the United States and Japan have recently made great efforts in research and development of new drugs to suppress cancer, and in particular, attention has been focused on the development of anticancer substances in natural products with fewer side effects.

Meanwhile, Prunellae Herba refers to the herbaceous herb of Prunella vulgaris L. var. Lilacina Nakai, a perennial herb belonging to Labiatae. It has many bracts and calyxes on flower axis, almost cylindrical, 3 ~ 6cm long, 10 ~ 15mm in diameter, and greyish brown on the outside. There is a corolla (花 있으며) on the upper part, a stem on the lower part, and 4 branches in the calyx. Bracts are heart-to-eccentric with white hairs on calyx and veins. It is produced in Korea, China, and Japan, and is a herbal medicine used for blue liver, diuresis, anti-inflammatory, small-species, fever, and blood pressure lowering.

The main components of the vinegar are triterpenoid, saponin, ursolic acid, rutin, hyperoxide, tannin, caffeic acid, alkaloid, vitamin B1, vitamin C, vitamin K, potassium salt, etc.The pharmacological action is potassium salt, triterpenoid, ursolic acid, etc. Tannins are astringent, alkaloids are anti-hemostatic [Ju Kim, et al., Korean Journal of Pharmacognosy, 4: 416-420, 2000]. However, the anticancer effects of hachocho have not been reported until now.

The present invention has been made to solve the problems of the prior art as described above, the object of the present invention is to provide an anticancer composition with excellent anticancer effect and minimized side effects as a natural product extract.

In order to achieve the above technical problem, the present invention provides a composition for the prevention, treatment and inhibition of cancer diseases comprising the extract of Hagocho as an active ingredient.

In detail, the present invention is characterized by inhibiting the growth of cancer cells or inhibiting cancer metastasis, and enhances the immune system activity of macrophages, spleen lymphocytes, natural killer cells or lymphokine activated killer cells.

In addition, excipients, extenders, and other herbal medicines may be mixed with the compositions of the present invention according to conventional methods to form pharmaceutically acceptable formulations such as tablets, capsules, syrups, oral dosage forms, injections, and the like.

In addition, the composition may be added to food materials such as beverages, teas, spices, gums, confections, or the like, including food acceptable food additives, or may be prepared as foods such as capsules, powders, and suspensions.

Hereinafter, the present invention will be described in detail by describing preferred embodiments of the present invention.

Production Example  : Hagocho  Preparation of Extract

The extracts of Hagocho were harvested at the end of July, when the flowers grown in Yangcheon Village, Baekjeon-myeon, Hamyang-gun, Gyeongnam Province, were collected. Sun dried, cut into 2cm size, and extracted with hot water at 100 ℃ for 7 hours. After concentration in vacuo, lyophilization and grinding were used as samples.

Lyophilization yield was 14.38%.

Example  0: Cell culture and experimental animal

1. Reagent

The following reagent was purchased and used.

RPMI 1640, DMEM (Dulbecco's modified Eagle's medium), HBSS (Hank's Balanced Salt Solution), fetal bovine serum (FBS), 100 U / ml penicillin, 50 µg / ml streptomycin are all GIBCO (Gaethersburg, MD, USA) Purchased from Trypsin was purchased from Sigma chemical co.

2. Cell Culture

YAC-1 (KCLB, # 40160) cells, mouse lymphoma cells, were passaged in RPMI 1640 medium containing 10% fetal bovine serum. Raw 264.7 (ATCC, # TIB-71), a macrophage, was used. Cells were passaged in DMEM medium containing 10% FBS. In addition, sarcoma 180 (Korean Cell Line Bank: KCLB, # 40066, Korea) cells, which are mouse sarcoma cells, were passaged in RPMI 1640 medium containing 10% fetal bovine serum and B16, a mouse melanoma cell. -F1 (American Type Culture Collection (ATCC, # CRL-6323, USA) and B16-F10 (American Type Culture Collection: ATCC, # CRL-6475, USA) cells were passaged in DMEM medium containing 10% FBS.

3. Experimental Animal

The experimental animals were purchased from Orient Bio Western Branch of 32-34 g ICR male 7 weeks old, 21-23 g C57BL / 6 male mice 7 weeks old, and 21-23 g Balb / c male mice 7 weeks old. . The feed (Purina Korea) and water were freely ingested, and the temperature in the kennel was maintained at 21 ~ 24 ℃ and relative humidity 40 ~ 80%. In addition, the light in the kennel was adjusted to repeat day and night every 12 hours.

Example  One : Hagocho  Experiment on Cytotoxicity of Extracts

To determine the non-toxic concentration of the Prunella vulgaris extract, 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide: MTT (Sigma, USA) was applied to mouse macrophage Raw 264.7 cells. The cytotoxicity was confirmed using the method. The MTT method is a sensitive method that can measure the viability, proliferation and activity of the cell, using the ability of mitochondrial dehydrogenase to convert yellow MTT into insoluble formazan.

Dispense the raw 264.7 cells into a 96-well plate at 1 × 10 ⁶ cells / ml, treat the vinegar extract with 1, 10, 50, 100 μg / ml, and then after 24 hours, 96 mg of 2 mg / ml MTT. The plate was treated with 100 μl. After 4 hours, the supernatant was discarded, and 100 μl of dimethyl sulfoxide (DMSO, Merck) solution was added to the wells, mixed uniformly, and absorbed at 540 nm using a Thermocycler ELISA Reader. The absorbances of formazan crystals of the experimental group were calculated as follows.

Cell viabilaty (%)

The results of measuring cytotoxicity of the extract of Hagocho are shown in FIG. As a result of measuring the cytotoxicity of the extract, the cytotoxicity was not found in the 1-100 μg / ml treatment group of the vinegar extract compared to the control group (VH). Thus, the concentration of 1-100 μg / ml Hachocho extract was used for cell culture experiments.

Example  2: effect of increasing phagocytosis of macrophages

To investigate the effects of macrophages on phagocytosis, macrophage raw 264.7 (ATCC, # TIB-71) cells were placed on a 48-well culture plate in 1 × 10 cells. ^The cells were aliquoted at ⁶ cells / well and treated with 10, 50, and 100 μg / ml of Hagocho extract and incubated for 24 hours.

The amount of fluorescein isothiocyanate (FITC) -labelled latex bead introduced into the cell by phagocytosis after the culture medium was changed and washed with PBS for 90 minutes with 1 × 10 ⁹ particles / ml of fluorescein isothiocyanate (FITC) -labelled latex bead. Was measured by fluorescence spectrophotometer (FL600, Bio-tek) at excitation 490 nm, emission 530 nm, and LPS (lipopolysaccride) was used as a positive control.

Figure 2 shows the effect of increasing the phagocytic activity of macrophages in accordance with the concentration of the extract. The phagocytic activity of macrophages by the positive control group (LDS) was increased 3.2 times compared to the negative control group (VH), 2 times in the 10 μg / ml group, 3 times in the 50 μg / ml group, and 100 μg / ml group Showed a four-fold increase in phagocytosis. That is, it can be seen that the effect of increasing the phagocytic activity in a concentration-dependent significant effect on the extract.

Example  3: macrophage Nitric oxide ( NO ) Increase increase effect

To investigate the effects of macrophages on nitric oxide (NO) by macrophages extract, mouse macrophage Raw 264.7 cells were dispensed at 5 × 10 ⁵ cells / well on a 48-well culture plate. The extracts were treated with 10, 50, 100 μg / ml and incubated for 24 hours.

In order to measure the amount of NO produced in the culture solution, 90 μl of the culture supernatant and 90 μl of the reaction solution I (1% sulfanyl amide, 5% phosphoric acid) were mixed and reacted at room temperature for 10 minutes, followed by reaction solution II (0.1% NED). 90 μl of the mixture was reacted for 10 minutes, and absorbance (Thermo electron corporation, VARIOSCAN) was measured at 550 nm. LPS was used as a positive control.

Figure 3 is a diagram showing the NO production amount of macrophages according to the concentration concentration of Haechocho extract. In the positive control group (LPS), the NO production of macrophages was increased by 8.2 times compared to the negative control group (VH), 3.3 times in the 10 μg / ml group, 5.4 times in the 50 μg / ml group, and 100 μg in the group treated with P. 7 times in the / ml group. That is, it can be seen that the production of NO significantly increased depending on the concentration of hachocho extract.

Example  4: spleen Lymphocyte  Proliferative effect

In order to separate splenocytes, Balb / c mice were sacrificed by cervical dislocation, and the abdomen was excised to remove spleen and subdivided using a 1 ml syringe. The cells were suspended in HBSS and then transferred to a 50 ml centrifuge tube (corning, USA) and centrifuged for 5 minutes at 1,000 rpm. The cell precipitate was suspended in 5 ml of ACK lysis buffer (Tris-buffered ammonium chloride: 0.16 M NH ₂ Cl, 0.17 M Tris, pH 7.2) per spleen, left for 1 minute, and washed twice with HBSS to remove red blood cells. After counting the cells with RPMI 1640 medium containing 10% fetal bovine serum, dispense the cells into 96-well plates at 1.5 × 10 ⁶ cells / ml, and process the extracts of Hagocho extract at 10, 50, 100 μg / ml After the time was put WST-1 Kit (Roche, 1 644 807) in 10μl / well and reacted in a CO ₂ incubator for 30 minutes and then measured the absorbance at 450nm to investigate the proliferative capacity of splenic lymphocytes by Hachocho extract.

Figure 4 shows the proliferation results of spleen lymphocytes according to the concentration of the extract. As a result of measuring the proliferative ability of lymphocytes by the haejucho extract, significant increase of 2 fold in the 50 μg / ml administration group and 2.5 times in the 100 μg / ml administration group was observed, compared to the control group (VH).

Example  5: spleen B Lymphocyte  Proliferative effect

Splenocytes were aliquoted into 96-well plates at 1.5 × 10 ⁶ cells / ml. And L lymphocyte B mitogen (mitogen) LPS was treated all 1μg / ml except negative control group (VH) and the extract was treated with 0, 10, 50, 100μg / ml, respectively, 72 After hours, WST-1 Kit (Roche, 1 644 807) was added to 10 μl / well and reacted in a CO ₂ incubator for 30 minutes, and then absorbance was measured at 450 nm to determine splenic B lymphocyte proliferative capacity.

The results of measuring the proliferative capacity of splenic B lymphocytes by concentrations of the extract of Hagocho are shown in FIG.

As a result of measuring the B lymphocyte proliferation ability by the Pleurotus eryngii extract, the LPS-only group increased 2.2 times compared to the control group (VH) without any treatment, and the proliferative capacity of B lymphocytes activated by LPS was 50μg / ml It was significantly increased to 3 times in the administration group and 3.7 times in the 100 μg / ml administration group.

Example  6: spleen T Lymphocyte  Proliferative effect

Splenocytes were dispensed in a 96-well plate at a concentration of 1.5 × 10 ⁶ cells / ml, and Con T (concanavaline A), a T lymphocyte mitogen, was 8 ng / ml except for the negative control group (VH). After all the treatments, and the Gochocho extract were treated with 0, 10, 50 and 100 μg / ml, and after 72 hours, the WST-1 Kit (Roche, 1 644 807) was placed in 10 μl / well and the CO ₂ incubator for 30 minutes. After the reaction, the absorbance was measured at 450 nm, and the splenic T lymphocyte proliferation ability by the extract of Hachocho was measured.

As a result of measuring the proliferative capacity of the spleen T lymphocytes according to the concentration of the extract of Hagocho. As a result of measuring T lymphocyte proliferative ability by P. chinensis extract, the proliferative capacity of Con A only group was increased by 2 times compared to the control group (VH) without any treatment. Significant increase was found to be 2.8 fold in the 50 μg / ml dose group and 3.5 fold in the 100 μg / ml dose group.

Example  7: cancer cells of natural killer cells Killing ability  Increase effect

In order to investigate the effect of natural killer cells (NK cells) on cancer cell killing ability by the vinegar extract, 10% bovine YAC-1 cells, mouse lymphoma cells, were used as target cells. Subcultures were performed using RPMI 1640 medium containing fetal serum. In addition, 1 × 10 ⁶ YAC-1 cells, which are target cells, were added to 15 μM of a fluorescent substance, calcin-AM (invitrogen, C1430), and reacted for 30 minutes in a CO ₂ incubator at 37 ° C. for fluorescent labeling. ) And washed three times with HBSS.

The effector cell / target cell ratio (E / T ratio) of natural killer cells as effector cells and YAC-1 cells as target cells was 10/1, 25/1, 50/1 1 × 10 ⁴ target cells were added to the culture medium, and 50 μl were dispensed into 96 well-plates, and the vinegar extract was treated with 10, 50, and 100 μg / ml for 4 hours. After centrifugation of the 96 well plate, 50 μl of the supernatant was measured by fluorescence spectrophotometer (FL600, Bio-tek) at excitation 485 nm and emission 530 nm to determine cytotoxicity against YAC-1 cells, which are target cells of natural killer cells. It is expressed in%.

Figure 7 is the result of measuring the cancer cell killing ability of natural killer cells (NK cells) according to the concentration of the vinegar extract. As shown in FIG. 7, the ratio of natural killer cells as effector cells and YAC-1 cells as target cells was 10/1, 25/1, and 50/1 compared to the control group (VH) without any treatment as shown in FIG. 7. Fluorescence was increased due to the increase of cancer cell killing ability of the killer cells, and the fluorescence was increased depending on the concentration of the hydrangea extract, and the extract was found to increase the cell killing ability of natural killer cells.

Example  8: cancer cell of lymphokine activated killer cell Killing ability  Increase effect

In order to investigate the killing effect of haejucho extract on cancer cells not killed by natural killer cells (NK cells), lymphocytes were treated with IL-2 (Interleukin-2) at 10 U / ml for 3 days. It induced the production of lymphokine activated killer cells (LAK cells) that can culture and kill cancer cells that are not killed by natural killer cells. 1 × 10 ⁶ YAC-1 cells, target cells, were added to 15 μM fluorescent calcin-AM (invitrogen, C1430) and reacted for 30 minutes in a CO ₂ incubator at 37 ° C. for fluorescent labeling, followed by HBSS Washed three times.

The effector cell / target cell ratio (E / T ratio) of lymphokine activated killer cells as effector cells and YAC-1 cells as target cells was 10/1, 25/1 and 50/1, respectively. Three experiments were carried out, and 1 × 10 ⁴ target cells were added to the culture medium, and 50 μl were dispensed into 96 well-plates, and the vinegar extract was treated with 10, 50, and 100 μg / ml for 4 hours. After centrifugation of 96well-plate, 50 μl of the supernatant was measured by fluorescence spectrophotometer (FL600, Bio-tek) at excitation 485 nm and emission 530 nm, and cytotoxicity against YAC-1 cells, which are target cells of lymphokine activating cells. (cytotoxicity) is expressed in%.

As a result of confirming the cancer cell killing ability of lymphokine activating killer cells by the haechocho extract as shown in Fig. 8 compared to the control group that did not process anything, effector cells lymphokine activating killer cells and target cells YAC-1 cells The fluorescence was increased in the group of 10/1, 25/1, 50/1, and fluorescence was increased depending on the concentration of P. vulgaris extract. It was confirmed that the ability to increase.

Example  9: In revenge cancer  Survival Rate Increase Effect

In order to investigate the effect of increasing the survival rate on the ascites cancer of P. vulgaris extract, the P. vulgaris extract was dissolved in physiological saline at 2 and 5 mg / kg, respectively, and administered orally to two experimental groups (10 animals per group) for 2 days each. It was. And saline was administered to the ICR mouse of the control group.

Subsequently, Sarcoma 180 cells, which are mouse sarcoma cells, were passaged in RPMI 1640 medium containing 10% fetal bovine serum and washed with HBSS, and then the number of cells was measured to measure 1 × 10 ⁶ cells / 0.1 ml. ICR mice were implanted intraperitoneally to cause ascites cancer.

After transplantation, the same method was used for 10 days at the same concentration. Only vinegar extract or saline solution was orally administered to the experimental and control groups, and the average survival and prolongation of survival were determined 30 days after tumor transplantation.

The prolongation of survival days for ascites cancers was calculated as follows, and the effect of increasing the survival rate for the plural cancers of Hachocho extract was investigated.

<Table 1> Survival rate increase effect of asiatica extract for ascites cancer

 Classification Dose (mg / kg)  Dosing method Average number of days living (day) % Survival extension 30 Day Survival (Marine)  Control  saline  Oral administration  20.3  0  0/10 Hagocho extract treatment group 1  2  Oral administration  28  37.9  2/10 Hagocho extract treatment group 2  5  Oral administration  30.7  51.2  3/10

As shown in Table 1, there were no mice surviving for 30 days in the control group, but in the group administered 2 mg / kg of vinegar extract, 2 out of 10 mice survived for 30 days, and the group administered 5 mg / kg of vinegar extract. Three out of 10 mice survived for 30 days, and the surviving number of ascites cancers was increased by the hyacinth extract.

In addition, although the average survival day of the control group was 20.3 days, the survival rate for the multi-cancerous cancer was 51.2% (37.9%, 28 days in the 2 mg / kg administration group of Hachocho extract, and 30.7 days in the 5 mg / kg administration group). It has been confirmed that the extracts of Prunella chinensis increased the survival rate for ascites cancer.

Example  10: Solid rock  Formation inhibitory effect

In order to investigate the inhibitory effect of the Prunella vulgaris extract on solid cancers, the Prunella vulgaris extract was dissolved in physiological saline at 2 and 5 mg / kg, respectively, and pre-administered to two experimental groups of C57BL / 6 mice (10 per group) for 2 days. . In addition, saline was administered to C57BL / 6 mice in the control group.

Subsequently, B16-F1 cells, which were mouse melanoma cells, were passaged in RPMI 1640 medium containing 10% fetal bovine serum. After washing with HBSS, the cell number was measured, and 2 × 10 ⁵ cells / 0.1 ml of cells were implanted subcutaneously into the C57BL / 6 mouse right inguinal tract to induce solid cancer. After transplantation, the haecho extract in the experimental group was administered orally for 10 days at the same concentration only in the control group, and solid tumors generated after 21 days were measured and weighed, and the inhibition of tumor formation was determined as an index of anticancer effect.

Tumor formation inhibition rate was calculated as follows to investigate the inhibition rate of solid cancer formation.

<Table 2> Inhibitory effect of Hachocho extract against solid cancer

Classification Dose (mg / kg) Dosing method Tumor weight (g) Tumor Inhibition Rate (%)  Control  Saline  Oral administration  3.99 ± 1.82  0 Hagocho extract treatment group 1  2  Oral administration  1.46 ± 0.79  63.4 Hachocho extract treatment group 2  5  Oral administration  0.54 ± 0.25  86.5

As shown in Table 2, the average weight of solid tumors formed in the control group was 3.99 g, and the inhibitory rate of tumor formation was 1.46 g in 6 mg / kg administration group and 63.4% in 5 mg / kg group, and 0.54 g in 5 mg / kg group. It was confirmed that 86.5% and the extract of the vinegar inhibited the tumor formation rate of solid cancer.

Example  11: inhibitory effect of lung cancer metastasis

In order to investigate the inhibitory effect on the lung cancer metastasis of the extract, the extract was dissolved in physiological saline at 2 and 5 mg / kg, respectively, and pre-administered orally to two experimental groups of C57BL / 6 mice (10 mice per group) for 2 days. In addition, saline was administered to C57BL / 6 mice in the control group.

B16-F10 cells, mouse melanoma cells, were passaged in RPMI 1640 medium containing 10% fetal bovine serum. After washing with HBSS, the number of cells was measured and lung cancer metastasis was induced by intravenously injecting 2 × 10 ⁵ cells / 0.1ml cells into the tail of C57BL / 6 mice. After transplantation, only Hachocho extract or physiological saline was orally administered to the experimental group and the control group for 14 days at the same concentration, and the number of colonies formed in the lungs generated after 14 days was measured. .

Lung cancer metastasis inhibition rate was calculated as follows to investigate the inhibition rate of lung cancer metastasis of hachocho extract.

<Table 3> Anticancer Effects of Pulmonary Tumor Extracts on Lung Tumor Metastasis of B16-F10 Tumor Cells

Classification Dose (mg / kg) Dosing method Tumor colony count (dog) Tumor metastasis inhibition rate (%)  Control  Saline  Oral administration  26.6 ± 3.54  0 Hagocho extract treatment group 1  2  Oral administration  15.4 ± 8.82  42.1 Hagocho extract treatment group 2  5  Oral administration  8.6 ± 2.28  67.7

As shown in Table 3, the average number of colonies in the control group was 26.6, and the ultrahigh extract 2 mg / kg group was 15.4, showing 42.1% inhibition of lung cancer metastasis compared to the control group, and the 5 mg / kg group was 8.6, 67.7%. Inhibition of lung cancer metastasis was shown and administration of the herb extract increased the inhibition of lung cancer metastasis.

Pharmaceutical Example

Formulation Example 1 Preparation of Tablet

100 mg of Hagocho extract was prepared according to a conventional tablet preparation method by mixing 100 mg of corn starch, 100 mg of lactose and 2 mg of magnesium stearate.

Formulation Example 2 Preparation of Capsule

500 mg of Hagocho extract was filled into soft gelatin capsules to prepare capsules.

Formulation Example 3 Preparation of Syrup

100 ml of syrup was prepared according to the conventional method of preparing a liquid formulation with a content of 1 g of Hagocho extract, 10 g of isomerized sugar, 5 g of mannitol, and an appropriate amount of purified water.

Formulation Example 4 Preparation of Injection

200 mg of the vinegar extract was heat-dissolved in 200 mg of physiological saline containing polyoxyethylene hydrogenated castro oil to prepare an injection containing the mixed extract at a concentration of 0.1%.

Functional Food Example : Preparation of Functional Beverages

After dissolving 200 mg of Gochocho extract in 96 ml of water, 500 mg of vitamin C as an adjuvant, 1 g of citric acid and oligosaccharide as an adjuvant, and 0.05 g of sodium benzoate as preservatives were added, followed by addition of purified water to 100 ml. Made functional drinks.

As described above, the composition containing the haejucho extract according to the present invention as an active ingredient is effective in enhancing immune system activity such as macrophages, spleen lymphocytes, natural killer cells or lymphokine active killer cells and cancer cell growth inhibition or cancer metastasis. It appears to have an inhibitory effect. It also appears to be safe and nontoxic.

That is, the present invention provides a composition for the prevention, treatment and suppression of cancer diseases with excellent anticancer effect and minimized side effects as a natural product extract.

Claims (6)

A composition for the prevention, treatment and inhibition of cancer diseases comprising the Hagocho extract as an active ingredient. In claim 1, The composition is a composition for the prevention, treatment and inhibition of cancer diseases, characterized in that it has the effect of inhibiting the growth or cancer metastasis of cancer cells. In claim 2, The cancer is ascites cancer, solid cancer or lung cancer, the composition for the prevention, treatment and inhibition of cancer diseases. In claim 1, The composition is a composition for the prevention, treatment and inhibition of cancer diseases, characterized in that to enhance the immune system activity of macrophages, spleen lymphocytes, natural killer cells or lymphokine activating killer cells. A pharmaceutical preparation for preventing, treating and inhibiting cancer diseases comprising the composition of any one of claims 1 to 4. Functional food for the prevention, improvement and suppression of cancer diseases comprising the composition of any one of claims 1 to 4.

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