KR20180112568A - Composition for preventing and treating a cancer comprising spatholobus suberectus dunn. - Google Patents

Abstract

The present invention relates to an anticancer activity of an extract of Spatholobus suberectus. Specifically, the extract of Spatholobus suberectus is confirmed to have cytotoxic effects on blood cancer cells, increase the intracellular ROS, and induce ER-stress reaction to induce apoptosis and remove cancer cells. Accordingly, the present invention relates to uses of the extract of Spatholobus suberectus as a composition for preventing or treating cancer, an anticancer adjuvant, and a food composition for preventing or alleviating cancer.

Description

TECHNICAL FIELD [0001] The present invention relates to a composition for preventing and treating cancers,

The present invention relates to the anticancer activity of extracts such as blood, blood and the like, and specifically relates to the cancer cell killing effect due to the ER-stress induction of extracts such as blood.

The smallest unit that makes up the human body is called a cell. Normal cells divide by the intracellular regulating function, grow, die and disappear, and maintain the balance of the cell number. When a cell is damaged for some reason, it is treated and acts as a normal cell. However, when the gene of a cell changes for various reasons, the cell is abnormally changed to incompletely mature, and the cell cycle is not regulated so that the cell division is continued, which is defined as cancer. Cancer is also characterized by its ability to invade the tissues and organs of the state and destroy them, as well as spread to other organs. The death rate due to cancer is the number one cause of death in Korea, and the number is increasing every year. Although there has been considerable progress in the medical treatment of certain cancers, the overall 5-year survival rate for all cancers has improved by only about 10% over the past 20 years. Cancer, or malignant tumors rapidly metastasize and grow in an uncontrolled manner, it is extremely difficult to detect and treat them on time.

Currently, surgery, radiation therapy, and chemotherapy are used to treat cancer. Of these, chemotherapy refers to a method of treating cancer using an anticancer agent. Today, about 60 kinds of various anticancer drugs are used. Recently, as knowledge of cancer development and characteristics of cancer cells is well known, researches on the development of new anticancer drugs are being actively carried out. Many patients in cancer chemotherapy suffer from the side effects of anticancer drugs, especially because of the toxicity of anticancer drugs. The anticancer substances used in clinical use have effects such as side effects and anticancer drug resistance, such as failure to treat the cancer cells as well as normal cells as well as repeated administration. A new type of anticancer drug is being studied to overcome the side effects of existing anticancer drugs caused by the diversity of the cancer itself and the variety of pathogenic mechanisms.

On the other hand, the ER (endoplasmic reticulum) consists of two types of membranous components, a rough endoplasmic reticulum with a ribosome attached thereto and a smooth endoplasmic reticulum without a ribosome have. Approximately one-third of intracellular proteins are transformed from mRNA to protein in post-translational modifications such as folding and assembly, glycation, and disulfide bonds. Resulting in an active protein structure. It is also a synthesis site of lipids and sterols and plays an important role in the regulation of intracellular calcium concentration by the calcium reservoir.

ER stress refers to the infiltration of immature proteins into the endoplasmic reticulum beyond the ability of the endoplasmic reticulum to be treated by physiological or pathological conditions, or the depletion of calcium in the endoplasmic reticulum, resulting in a failure of the endoplasmic reticulum. Which means that the ER protein folding load has been exceeded. When an endoplasmic reticulum stress occurs, the cell has a defense mechanism to survive, which is called an ER stress response. The endoplasmic reticulum stress response is mediated by three signaling pathways, PERK (pancreatic ER kinase), IRE-1α / XBP-1 (inositol-requiring 1α / X-box binding protein 1) and ATF6 (activating transcription factor) Lt; / RTI > The ER stress response occurs in four ways. The first response is translational attenuation of translation from mRNA to protein in the ribosome, reducing the entry of new proteins into the endoplasmic reticulum. Specifically, protein synthesis is reduced as a result of reduced eIF2 (eukaryotic translation initiation factor 2) polymer activity, which normally collects tRNAs with initiation methionyl on the 40S ribosomal subunit. Recently, more than two mechanisms for reducing protein folding load have been described, including IRE1 (also known as ERN1) -mediated ER-material mRNA degradation mechanism (Hollien and Weissman, 2006), or p58 ^IPK (Rutkowski et al., 2007; Rutkowski et al., 2007). This is the mechanism by which proteins that go to the ER are synthesized and decomposed simultaneously by enzymes (also known as DNAJC3). The second response is to enhance the folding ability of the endoplasmic reticulum by inducing the expression of an endoplasmic chaperone, such as Bip (also known as HSPA5), which is required to fold the protein. Transcription of the gene encoding the epidermal chaperone, rather than the cytoplasmic chaperone, is activated through a signal transduction pathway known as the unfolded protein response (UPR). In mammals, IRE1, PERK and ATF6 are transmembrane transmembrane proteins that sense the presence of misfolded proteins in the endoplasmic reticulum lumen. Under normal conditions, these signaling molecules remain inactive through the interaction of the lumenal region with the protein chaperone, Bip. Bip is an ER material member of the Hsp70 protein chaperone assembly that interacts with unfolded polypeptide chains. When the endoplasmic reticulum is exposed to stress, the Bip binds to the unfolded protein and is secreted from these stress sensors. Secretion of IRE1 from Bip activates protein kinase and endoribonuclease activity to initiate the junction of XBP1 that generates the major transcription factor of UPR gene expression. When secreted from Bip, PERK is activated, phosphorylating the alpha subunit of eIF2 and weakening protein synthesis. Paradoxically, eIF2a phosphorylation is promoted simultaneously with translation of selective mRNAs. For example, the high translational activity of the transcription factor ATF4 by eIF2a phosphorylation results in about 33% induction of the UPR gene. Finally, Bip secretion activates the transcription of the UPR gene, which contains a molecular chaperone such as Bip and Grp94 (also known as HSP90B1) and the ER internal folding enzyme, which translocates ATF6 to the Golgi and breaks down into the cytoplasmic segments that migrate from the Golgi to the nucleus. The third response is the ER stress-asoociated degradation (ERAD), which is the process by which the unfolded or misfolded proteins in the endoplasmic reticulum are degraded and removed through the cytoplasmic ubiquitin-proteasome system. Finally, when the endoplasmic reticulum is unable to recover from its function (ATF4 and CHOP expression), the apoptosis pathway is activated and the damaged cells are removed.

Avoiding apoptosis is a typical feature of malignant cells, and cell resistance to apoptosis is an important clinical problem (Jia et al. , 2012; Lopez-Beltran et al. , 2007). Accordingly, the inventors of the present invention have made intensive studies to find natural products with less side effects that can prevent and treat cancer by controlling the stress of the endoplasmic reticulum. As a result, it has been confirmed that the extracts of blood and blood can induce ER-stress and effectively kill cells in cancer cells Thus completing the present invention.

In the present invention, it has been confirmed in the present invention that the blood-extractive extract has a cytotoxic effect on cancer cells, in particular induces ER-stress and induces the death of cancer cells. Thus, it is possible to provide a composition for preventing or treating cancer, Or a food composition for improvement.

In order to accomplish the above object, the present invention provides a pharmaceutical composition for preventing or treating cancer comprising an extract of blood-brain and the like as an active ingredient.

In addition, the present invention provides an anticancer adjuvant comprising an extract of blood-brain-extract etc. as an active ingredient.

In addition, the present invention provides a food composition for preventing or ameliorating a cancer containing a blood-brain-like extract.

According to the present invention, the blood-brain extracts have cytotoxic effects on blood cancer cells, increase intracellular ROS, induce ER-stress reaction and induce apoptosis, thereby eliminating cancer cells.

FIG. 1 is a view showing the cell survival rate of multiple myeloma cell line U266 by treatment with blood extract and the like.
FIG. 2 is a graph showing the degree of ROS production in a multiple myeloma cell line U266 by treatment with a blood-brain-light extract.
FIG. 3 is a graph showing the degree of expression of apoptosis-related proteins in the multiple myeloma cell line U266 by treatment with blood extract.
FIG. 4 is a graph showing the expression levels of ER-stress-related cell death inducing proteins in the multiple myeloma cell line U266 by treatment with blood extract.

Hereinafter, the present invention will be described in detail with reference to embodiments of the present invention. It should be understood, however, that the invention is not limited thereto and that various changes and modifications may be made therein without departing from the spirit and scope of the invention as defined by the appended claims and their equivalents. .

In one aspect, the present invention provides a method of treating a blood- suberectus Dunn.) extract as an active ingredient. The present invention also relates to a pharmaceutical composition for preventing or treating cancer.

In one embodiment, the extract may be extracted with water, an alcohol having 1 to 4 carbon atoms or a mixed solvent thereof, more preferably ethanol.

The term "extract " used in the present invention means a preparation which is obtained by squeezing a herbal medicine with an appropriate leaching solution and concentrating the liquid by evaporating the leaching solution. The extract is not limited thereto, but may be a diluent or concentrate of the extract, A dried product obtained by drying the extract, a controlled preparation thereof, or a purified product thereof. The blood-brain extract can be prepared by a common extraction method, separation and purification methods known in the art. The extraction method may be, but not limited to, hot water extraction, hot water extraction, cold extraction, reflux cooling extraction, or ultrasonic extraction.

In the present invention, the extract may be prepared by extracting with an extraction solvent or extracting with an extraction solvent, followed by fractionation with a fraction solvent. The organic solvent may be an alcohol having 1 to 4 carbon atoms, a polar solvent such as ethyl acetate or acetone, a solvent such as hexane or dichloro, or an organic solvent such as dichloromethane, Methane, or a mixed solvent thereof may be used. Further, water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof may be preferably used, and more preferably ethanol can be used. In one embodiment of the present invention, the extract was extracted with 100% ethanol as a solvent and then concentrated under reduced pressure.

In one embodiment, the cancer is selected from the group consisting of: colorectal cancer, breast cancer, cervical cancer, cervical cancer, ovarian cancer, prostate cancer, brain tumor, head and neck carcinoma, melanoma, myeloma, leukemia, lymphoma, stomach cancer, pancreatic cancer, Renal cell carcinoma, kidney cell carcinoma, renal pelvic carcinoma, bone cancer, skin cancer, head cancer, endometrial carcinoma, endometrial carcinoma, vulvar carcinoma, Hodgkin's disease, bladder cancer, kidney cancer, Cancer of the central nervous system (CNS), primary CNS lymphoma, spinal cord tumor, cervical cancer, endometrial carcinoma, endometrial carcinoma, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, Brain giant glioma, and pituitary adenoma. It is more preferable that the myeloma is multiple myeloma.

In one embodiment, a blood sample extract may be included at a concentration of 10-100 μg / ml.

In one embodiment, the prevention or treatment of cancer can be accomplished by apoptosis due to endoplasmic reticulum (ER) stress.

In one embodiment of the present invention, it was confirmed that the death of multiple myeloma cells by the treatment with blood extract was increased, and in order to confirm the anticancer activity mechanism, expression of the apoptosis-related proteins and ER-stress related apoptosis- As a result, it was confirmed that Pro-PARP and procaspase-3 related to apoptosis were decreased and Bax expression was increased by treatment with extracts such as blood, etc. (Fig. 3), and ERK-induced apoptosis (FIG. 4), the expression of IRE1α, ATF4, Elf2α, XBP-1 and CHOP (FIG. 4) was confirmed to be indicative of the anticancer activity by inducing ER-stress and inducing apoptosis Respectively.

As used herein, the term "apoptosis" is also referred to as apoptosis or apoptosis, and is a kind of programmed cell death (PCD), which is characterized by abnormal cells, damaged cells, aged It is a mechanism that allows cells to maintain their overall physical health by killing themselves and killing themselves.

In the present invention, the term "prevention" means any action which inhibits or delays the development, spread and recurrence of cancer by the administration of the pharmaceutical composition according to the present invention, and "treatment" Suspicion of cancer and all the behaviors that alleviate or ameliorate symptoms of the onset individuals.

As used herein, the term "treatment" means any action that improves or alters the death of a cancer cell or the symptom of cancer by administration of a composition including blood-brain or the like of the present invention. Those skilled in the art will be able to ascertain the precise criteria of the disease for which the composition of the present invention is effective by referring to the data presented by the Korean Medical Association, will be.

The term "therapeutically effective amount " used in combination with the active ingredient in the present invention means the amount of a pharmaceutically acceptable salt of an extract of the blood-brain extract effective for preventing or treating a target disease, The effective amount may vary depending on a variety of factors, such as the mode of administration, the site of administration, the condition of the patient, and the like. Therefore, when used in the human body, the dosage should be determined in consideration of safety and efficacy. It is also possible to estimate the amount used in humans from the effective amount determined through animal experiments. Such considerations in determining the effective amount are described, for example, in Hardman and Limbird, eds., Goodman and Gilman ' s Pharmacological Basis of Therapeutics, 10th ed. (2001), Pergamon Press; And E.W. Martin ed., Remington ' s Pharmaceutical Sciences, 18th ed. (1990), Mack Publishing Co.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. As used herein, the term "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment and not causing side effects, Factors well known in the art and other medical disciplines including health status, type of cancer, severity, activity of the drug, sensitivity to the drug, method of administration, time of administration, route of administration and rate of release, duration of treatment, ≪ / RTI > The composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered singly or multiply. Taking all of the above factors into consideration, it is important to administer an amount that can achieve the maximum effect in a minimal amount without side effects, which can be easily determined by those skilled in the art.

The pharmaceutical compositions of the present invention may include carriers, diluents, excipients, or a combination of two or more thereof commonly used in biological formulations. As used herein, the term "pharmaceutically acceptable" means that the composition is free of toxicity to cells or humans exposed to the composition. The carrier is not particularly limited as long as the composition is suitable for in vivo delivery, for example, Merck Index, 13th ed., Merck & Inc. A buffered saline solution, a buffer solution, a dextrose solution, a maltodextrin solution, glycerol, ethanol, and one or more of these components may be mixed and used, and if necessary, an antioxidant, a buffer, Conventional additives may be added. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to formulate into main dosage forms such as aqueous solutions, suspensions, emulsions, etc., pills, capsules, granules or tablets. Further, it can be suitably formulated according to each disease or ingredient, using the method disclosed in Remington's Pharmaceutical Science (Mack Publishing Company, Easton PA, 18th, 1990) in a suitable manner in the art.

The pharmaceutical composition of the present invention may further comprise a pharmaceutically acceptable additive, wherein the pharmaceutically acceptable additives include starch, gelatinized starch, microcrystalline cellulose, lactose, povidone, colloidal silicon dioxide, calcium hydrogen phosphate, Wherein the starch is selected from the group consisting of lactose, mannitol, sugar, arabic gum, pregelatinized starch, cornstarch, powdered cellulose, hydroxypropyl cellulose, opaques, sodium starch glycolate, carnauba wax, synthetic aluminum silicate, stearic acid, magnesium stearate, Calcium, white sugar, dextrose, sorbitol and talc may be used. The pharmaceutically acceptable additive according to the present invention is preferably included in the composition in an amount of 0.1 part by weight to 90 parts by weight, but is not limited thereto.

The pharmaceutical composition of the present invention may further comprise a known anticancer agent in addition to an extract of blood or blood as an active ingredient and may be used in combination with other known treatments for the treatment of these diseases. Other treatments include, but are not limited to, chemotherapy, radiation therapy, hormone therapy, bone marrow transplantation, stem cell replacement therapy, other biological therapies, immunotherapy, and the like.

Examples of anticancer agents that can be included in the pharmaceutical composition of the present invention include DNA alkylating agents such as mechloethamine, chlorambucil, phenylalanine, mustard, cyclophosphate, Cyclophosphamide, ifosfamide, carmustine (BCNU), lomustine (CCNU), streptozotocin, busulfan, thiotepa, cisplatin cisplatin and carboplatin; The anticancer antibiotics include dactinomycin (actinomycin D), doxorubicin (adriamycin), daunorubicin, idarubicin, mitoxantrone, Plicamycin, mitomycin C, and bleomycin; And plant alkaloids such as vincristine, vinblastine, paclitaxel, docetaxel, etoposide, teniposide, topotecan, And iridotecan, but are not limited thereto.

In one aspect, the present invention relates to a method of preventing or treating cancer, comprising administering an extract, such as a blood-brain extract, to a subject in need thereof.

The term "individual" used in the present invention means a monkey, a horse, a sheep, a pig, a chicken, a turkey, a quail, a cat, a dog, a mouse, a rat, a rabbit Refers to all animals including guinea pigs, and the diseases can be effectively prevented or treated by administering the pharmaceutical composition of the present invention to an individual. The pharmaceutical composition of the present invention can be administered in parallel with existing therapeutic agents.

The term "administering" as used herein means providing a predetermined substance to a patient in any appropriate manner, and may be administered orally or parenterally (for example, by intravenous, subcutaneous, The dosage may vary depending on the patient's body weight, age, sex, health condition, diet, administration time, administration method, excretion rate, and severity of the disease. Examples of the liquid preparation for oral administration of the composition of the present invention include suspensions, solutions, emulsions, syrups, and the like. In addition to water and liquid paraffin which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, Etc. may be included together. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried preparations, suppositories, and the like. The pharmaceutical composition of the present invention may be administered by any device capable of moving the active substance to the target cell. The preferred modes of administration and formulations are intravenous, subcutaneous, intradermal, intramuscular, and drip injections. The injectable solution may be a non-aqueous solvent such as an aqueous solvent such as a physiological saline solution or a ring gel solution, a vegetable oil, a higher fatty acid ester (e.g., oleic acid), an alcohol (e.g., ethanol, benzyl alcohol, propylene glycol, glycerin, etc.) (For example, ascorbic acid, sodium hydrogen sulfite, sodium pyrophosphate, BHA, tocopherol, EDTA and the like), an emulsifier, a buffer for pH control, a microbial growth inhibitor And a pharmaceutical carrier such as a preservative (e.g., mercury nitrate, thimerosal, benzalkonium chloride, phenol, cresol, benzyl alcohol, etc.).

In one aspect, the present invention provides a method of treating a blood- suberectus Dunn.) extract as an active ingredient.

In one aspect, the present invention provides a method of treating a blood- suberectus Dunn.) extract of the present invention.

When the composition of the present invention is used as a food composition, the above-mentioned blood-brain extract etc. can be used as it is, or it can be used together with other foods or food ingredients, and can be suitably used according to ordinary methods. The composition may contain a food-acceptable food-aid additive in addition to the active ingredient, and the amount of the active ingredient to be mixed may be suitably determined according to the intended use (prevention, health or therapeutic treatment).

As used herein, the term "food-aid additive " refers to a component that can be added to foods in a supplementary manner, and is appropriately selected and used by those skilled in the art as added to produce health functional foods of each formulation. Examples of food-aid additives include flavors such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, colorants and fillers, pectic acid and its salts, alginic acid and its salts, organic acids, , a pH adjusting agent, a stabilizer, a preservative, a glycerin, an alcohol, and a carbonating agent used in a carbonated drink. However, the types of the food auxiliary additives of the present invention are not limited by these examples.

A health functional food may be included in the food composition of the present invention. The term "health functional food " as used in the present invention refers to a food prepared and processed in the form of tablets, capsules, powders, granules, liquids and rings using raw materials and components having useful functions in the human body. Here, 'functional' refers to the structure and function of the human body to obtain nutritional effects and obtain useful effects for health use such as physiological action. The health functional food of the present invention can be prepared by a method commonly used in the art, and can be prepared by adding raw materials and components which are usually added in the conventional technical fields. In addition, the formulations of the above health functional foods may also be manufactured without limitations as long as they are acceptable as health functional foods. The composition for food of the present invention can be manufactured in various forms, and unlike general pharmaceuticals, it has the advantage that there is no side effect that may occur when a drug is used for a long period of time, and is excellent in portability, Can be ingested as an adjuvant to enhance the effectiveness of anticancer drugs.

There is no limitation on the kind of health food to which the composition of the present invention can be used. In addition, the composition containing the blood extract of the present invention as an active ingredient may be prepared by mixing other suitable auxiliary ingredients, which may be contained in health functional foods, with known additives according to the selection of a person skilled in the art. Examples of foods that can be added include dairy products, such as meat, sausage, bread, chocolates, candies, snacks, confectionery, pizza, ramen, other noodles, gums, ice cream, various soups, drinks, tea, Vitamin complex, and the like, and can be prepared by adding to the juice, tea, jelly, and juice prepared from the extract of the present invention as a main component.

Since the blood-extractive extract of the present invention is a raw material of natural plant, it can be used safely in pharmaceutical compositions and health functional foods because it can be less harmful than general synthetic compounds even when used as a pharmaceutical composition or a food composition .

The present invention will be described in more detail with reference to the following examples. However, the following examples are only for the purpose of illustrating the present invention, and thus the present invention is not limited thereto.

Example  One. Bloodbath ( Spatholobus suberectus Dunn .) Preparation of extract

Blood and blood were extracted with 100% ethanol and concentrated under reduced pressure to prepare ethanol extracts such as blood.

Experimental Example  One. Bloodbath  Cytotoxicity of extracts to multiple myeloma

To confirm the anticancer activity against the multiple myeloma of the blood red blood extract prepared in Example 1, 50 μl of U266 cell line (Korean Cell Line Bank), which is a multiple myeloma cell line, was added to 96 well-plate at 2 × 10 ⁴ cells / well The cells were cultured in RPMI medium supplemented with 10% inactivated fetal bovine serum and 1% penicillin-streptomycin in a 5% CO ₂ incubator (MCO-15AC, Sanyo, Osaka, Japan) at 37 ° C. The cell line U266 was treated with 50 μl of each of the blood extracts prepared in Example 1 at concentrations of 0, 12.5, 25, 50, 100 and 200 μg / ml, respectively, followed by incubation for one day. Cells incubated in the dark state were treated with 10 [mu] l of EZ-Cytox reagent (DoGen) and incubated for 30 minutes to 4 hours, and cell viability was confirmed at 450 nm absorbance of the microplate reader (Bio-rad Model 680).

As a result, it was found that toxicities were observed in multiple myeloma depending on the concentrations of extracts such as blood and blood (Fig. 1).

Experimental Example  2. In multiple myeloma Bloodbath  Determination of active oxygen inducing effect of extract

4 × 10 ⁵ U266 cell lines, multiple myeloma cell lines, were washed with PBS and then stained with DCFDA Cellular Reactive Oxygen Species Detection Assay kit (abcam) to confirm the effect of the blood extract of Example 1 on multiple myeloma. Was incubated at 37 [deg.] C for 30 minutes in a dark state, and then washed again with PBS. Subsequently, 2 x 10 ⁴ cells (50 μl) were dispensed into 96-well plates, and 50 μl of the blood-red light extract prepared in Example 1 was treated with 0, 10 and 20 μg / ml, ≪ / RTI > for a period of time. After that, the absorbance was measured at 485/535 nm using a microplate reader.

As a result, it was found that ROS production was increased in multiple myeloma cells when 10 or 20 μg / ml of blood was administered (FIG. 2).

Experimental Example  3. Bloodbath  Identification of cell death-inducing effects of multiple myeloma by extract

In order to examine the effect of the blood extract of Example 1 on apoptosis in multiple myeloma, PARP (Poly ADP ribose polymerase) and caspase-3, which are known to be cleaved during apoptosis, The expression level of Bax (bcl-2-like protein 4), which is known to be known, was confirmed by Western blot analysis. Specifically, the U266 cell line, which is a multiple myeloma cell line, was treated with 0, 10 and 20 / / ml of the blood extracts prepared in Example 1, respectively, and incubated in a 5% CO ₂ incubator at 37 캜 for 24 hours. Cells were then harvested and winked with PBS and resuspended in cell disruption buffer RIPA (20 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1 mM NaEDTA, 1 mM EGTA, 1% NP-40, 1% sodium pyrpphophate, 1 mM beta-glycerophosphate , 1 mM Na ₃ VO ₄ And 1 ug / ml leupeptin (Cell signaling)) was added and cells were disrupted at 4 캜 for 30 minutes. Cell lysates were centrifuged at 15,520 × g for 20 minutes to remove cell membrane components. Proteins were quantified using RC DC ™ Protein Assay Kit II (Bio-Rad, USA) 0, 10 and 20 [mu] g / ml treated groups). 5 × loading dye was added to the prepared protein sample and the protein was denatured by heating at 95 ° C for 5 minutes. Thereafter, the sample was loaded on an 8 to 10% SDS-PAGE gel and electrophoresed at 90 to 100 V for 1 hour and 30 minutes to 3 hours. Proteins separated from the gel were incubated at 200 mA to 300 mA for 1 hour to 4 hours And transferred to the membrane. The transferred membrane was blocked with TBST (tris buffered saline, pH 7.5) solution containing 5% skim milk for 2 hours at room temperature, followed by primary antibody anti-Pro-PARP (cell signaling), anti- signaling or anti-Bax (cell signaling) were diluted 1: 500, respectively, and reacted with the blocked membrane at 4 ° C for 1 hour to overnight. After washing 3 times with TBST, anti-rabbit IgG conjugated with horseradish peroxidase was diluted 1: 2,000, reacted with membrane at room temperature for 1 hour, and washed three times with TBST. The washed membrane was reacted with a chemiluminescent reagent for 1 to 3 minutes and photographed on a film to visualize the protein band.

As a result, Pro-PARP and procaspase-3 decreased depending on the concentration of blood and blood, and the cleavage of these was improved by the cerebellum, etc., and the expression of Bax increased in proportion to the treatment concentration of the blood, (Fig. 3). The results are shown in Fig.

Experimental Example  4. Bloodbath  Identification of the ER-stress-related apoptosis inducing effect of multiple myeloma by extract

In order to investigate whether ERK-induced apolipoprotein (PMK) -induced cell death is due to ER-stress and its apoptotic cell death, PERK (PRKR-like endoplasmic reticulum kinase), IRE1α 6), Elf2α (eukaryotic translation initiation factor 2α), XBP-1 (X-box-binding protein 1) and CHOP (C / EBP homologous protein) Specifically, the U266 cell line, which is a multiple myeloma cell line, was treated with 0, 10, and 20 / / ml of the blood extract of Example 1, incubated for 24 hours in a 5% CO ₂ incubator at 37 캜, Cells were disrupted in the same manner as in Experimental Example 3, electrophoresed, and transferred to the membrane. The transferred membrane is treated with primary antibodies such as anti-PERK (BIOSS antibodies), anti-IRE1? (Cell signaling), anti-ATF4 (cell signaling), anti-Elf2? (Cell signaling), anti- Western blot analysis was performed using anti-rabbit IgG and anti-mouse IgG (secondary antibody against anti-CHOP) as a secondary antibody and reacted with -CHOP (cell signaling).

As a result, expression of PERK, IRE1α, ATF4, Elf2α, XBP-1 and CHOP, which are proteins that induce ER-stress related cell death in multiple myeloma cells, was increased by treatment with extracts such as blood, (Fig. 4).

Claims (8)

Blood gases ( Spatholobus suberectus Dunn.) extract as an active ingredient. The pharmaceutical composition for preventing or treating cancer according to claim 1, wherein the extract is extracted with water, an alcohol having 1 to 4 carbon atoms or a mixed solvent thereof. The method of claim 1, wherein the cancer is selected from the group consisting of colon cancer, breast cancer, uterine cancer, cervical cancer, ovarian cancer, prostate cancer, brain tumor, head and neck carcinoma, melanoma, myeloma, leukemia, lymphoma, gastric cancer, lung cancer, Renal cancer, renal cell carcinoma, renal pelvic carcinoma, bone cancer, skin cancer, head cancer, endometrial carcinoma, endometrial carcinoma, vulvar carcinoma, Hodgkin's disease, bladder cancer, kidney cancer, Cancer of the central nervous system (CNS), primary CNS lymphoma, spinal cord tumor, endometrial carcinoma, thyroid cancer, pituitary cancer, adenocarcinoma, soft tissue sarcoma, urethral cancer, , Brain stem glioma, and pituitary adenoma. ≪ Desc / Clms Page number 19 > The pharmaceutical composition according to claim 1, wherein the cancer is multiple myeloma. The pharmaceutical composition for preventing or treating cancer according to claim 1, wherein the blood extract is contained at a concentration of 10 to 100 μg / ml. The pharmaceutical composition according to claim 1, wherein the prevention or treatment of cancer is accomplished by apoptosis caused by an endoplasmic reticulum (ER) stress. Anticancer supplements containing extracts such as blood and blood as active ingredients. A composition for preventing or ameliorating a cancer containing an extract such as blood, blood and the like.

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