WO2020122476A1 - Pharmaceutical composition comprising lactobacillus sakei wikim30 as active ingredient for prevention or treatment of cancer - Google Patents

Pharmaceutical composition comprising lactobacillus sakei wikim30 as active ingredient for prevention or treatment of cancer Download PDF

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Publication number
WO2020122476A1
WO2020122476A1 PCT/KR2019/016627 KR2019016627W WO2020122476A1 WO 2020122476 A1 WO2020122476 A1 WO 2020122476A1 KR 2019016627 W KR2019016627 W KR 2019016627W WO 2020122476 A1 WO2020122476 A1 WO 2020122476A1
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Prior art keywords
cancer
wikim30
composition
lactobacillus
prevention
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PCT/KR2019/016627
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French (fr)
Korean (ko)
Inventor
최학종
윤미선
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한국식품연구원
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Priority claimed from KR1020190082026A external-priority patent/KR102136772B1/en
Application filed by 한국식품연구원 filed Critical 한국식품연구원
Priority to EP19896691.3A priority Critical patent/EP3895719A4/en
Priority to CN201980081668.4A priority patent/CN113164536A/en
Priority to AU2019396986A priority patent/AU2019396986A1/en
Priority to SG11202106163RA priority patent/SG11202106163RA/en
Priority to CA3122124A priority patent/CA3122124A1/en
Priority to US17/312,698 priority patent/US20220062355A1/en
Priority to JP2021532434A priority patent/JP2022534150A/en
Publication of WO2020122476A1 publication Critical patent/WO2020122476A1/en

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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K10/00—Animal feeding-stuffs
    • A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
    • A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
    • A23K10/18—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions of live microorganisms
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A61K35/741—Probiotics
    • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
    • A61K35/747—Lactobacilli, e.g. L. acidophilus or L. brevis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the present invention relates to a composition for preventing or treating cancer comprising Lactobacillus sakei (Accession No. KCCM11878P) isolated from kimchi as an active ingredient.
  • Cancer has a high mortality rate worldwide and is the most common cause of death after cardiovascular disease in Western societies.
  • colonization, breast cancer, and prostate cancer are continuously increasing due to the rapid increase in environmental pollutants and increased alcohol consumption due to the westernization of eating habits, and the smoking population along with the aging of the population.
  • Lung cancer is increasing due to the increase in and air pollution.
  • lactic acid bacteria are widely distributed in humans and animals such as oral, intestinal, vaginal, fecal, and fermented foods such as kimchi, and are closely related to human and animal health. Lactic acid bacteria show various health promoting effects such as intestinal action, suppression of harmful bacteria, immune regulation, lowering of blood cholesterol, and anti-cancer.
  • Korean Patent Publication No. 10-2015-0068061 discloses anticancer activity of Lactobacillus plantarum PNU (KCCM11352P) or Lactobacillus mesenteroides PNU (KCCM11353P), and registered Korean patent No. 10-1287120 discloses a pharmaceutical composition for treating cancer containing Lactobacillus plantarium DSR CK10 [Accession No.: KFCC-11433P] or Lactobacillus plantarium DSR M2 [Accession No.: KFCC-11432P] as an active ingredient.
  • KFCC-11433P Accesion No.: KFCC-11433P
  • Lactobacillus plantarium DSR M2 Accesion No.: KFCC-11432P
  • An object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of cancer comprising the Lactobacillus Sakeai strain as an active ingredient.
  • another object of the present invention is to provide a food composition or a food additive composition for the prevention or improvement of cancer comprising the novel kimchi lactic acid bacteria Lactobacillus Sakeai strain as an active ingredient.
  • Another object of the present invention is to provide a feed composition or feed additive composition for the prevention or improvement of cancer, including the novel kimchi lactic acid bacteria Lactobacillus Sakeai strain as an active ingredient.
  • Another object of the present invention is to provide a method for preventing or treating cancer, wherein the composition is administered to an individual other than a human.
  • the present inventors tried to find a lactic acid bacteria strain that shows the prevention and treatment effect of cancer while showing excellent effects as a probiotic from kimchi, and isolated and identified Lactobacillus sakei, WIKIM30, Lactobacillus sakei, WIKIM30, a lactic acid bacteria strain having anticancer activity.
  • the present invention has been completed.
  • the present invention provides a food composition or food additive composition for preventing or improving cancer comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a culture thereof as an active ingredient.
  • the present invention provides a feed composition or feed additive composition for preventing or improving cancer in livestock, comprising Lactobacillus sakei WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a culture thereof as an active ingredient.
  • another object of the present invention provides a method for preventing or treating cancer, wherein the composition is administered to an individual other than a human.
  • Lactobacillus Sakeai WIKIM30 exhibits excellent anticancer activity in animal models and cancer cell lines in which cancer cells are transplanted, it can be usefully used as a composition for the treatment, prevention or improvement of cancer in humans or animals Can be.
  • FIG. 1 is a photograph of a tumor portion of a mouse for observing a change in the size of a tumor cell over time after intravenous injection of a composition according to the present invention to a CT26 cell transplanted mouse once (Single) or three times (Serial) It is a diagram showing.
  • FIG. 2 is a view showing a graph of measuring the change in volume of tumor cells over time after a single (or single) or three (Serial) intravenous injection of CT26 cell-grafted mice according to the present invention.
  • FIG. 3 is a view showing a photograph of a tumor portion of a mouse for observing that the size of tumor cells changes over time after intravenous injection of a composition according to the present invention into a B16F10 skin melanoma mouse.
  • FIG. 4 is a graph showing a change in the volume of tumor cells over time after intravenous injection of a composition according to the present invention into a B16F10 skin melanoma mouse.
  • FIG. 5 is a graph showing the composition of the present invention treated with Lactobacillus Sakeai WIKIM30 in mouse-derived colorectal cancer CT26 cell line, human-derived colorectal cancer SW620 cell line, and HCT116 cell line, and measuring cell viability assay.
  • FIG. 6 is a graph showing the measurement of cell viability assay by treating Lactobacillus Sakeai WIKIM30 in a human-derived non-small lung cancer cell H1650 cell line.
  • FIG. 7 is a graph showing the treatment of Lactobacillus Sakeai WIKIM30 on human-derived pancreatic cancer ASPC1 cell line and PANC1 cell line and measuring cell viability assay.
  • FIG. 8 is a diagram showing a graph of measuring Lactobacillus Sakeai WIKIM30 in a human-derived liver cancer HepG2 cell line and measuring cell viability assay.
  • FIG. 9 is a view showing a graph of measuring Lactobacillus Sakeai WIKIM30 in a human-derived bladder cancer T24 cell line and measuring cell viability assay.
  • FIG. 10 is a diagram showing a graph obtained by treating Lactobacillus Sakeai WIKIM30 on a human-derived normal skin CCD-986-sk cell line and measuring cell viability assay.
  • composition comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) or a culture thereof as an active ingredient of the present invention has an effect of preventing or treating cancer and can be used as a pharmaceutical composition.
  • Lactobacillus sakei is a lactic acid bacteria strain derived from kimchi. Although Lactobacillus sakei (WIKIM30) of Lactobacillus sake in the present invention was separated and identified from kimchi, the route for obtaining it is not limited thereto.
  • the microorganism of the present invention having the 16S rRNA nucleotide sequence of SEQ ID NO: 1 was named Lactobacillus sake WIKIM30 (Lactobacillus sakei), and deposited with the Korea Microbiological Conservation Center on September 10, 2015 (Accession No. KCCM11878P).
  • Lactobacillus sakeai WIKIM30 of the present invention is a probiotic, and has a general suit effect and an immune enhancing effect of lactic acid bacteria. It is well known that lactic acid bacteria in the genus Lactobacillus have a formal effect immune enhancing effect.
  • probiotics in the present invention, is understood to mean a living microorganism that has a beneficial effect on the health of the host by improving the intestinal microbial environment of the host in the gastrointestinal tract of animals, including humans.
  • Probiotics which are living microorganisms with probiotic activity, may have a beneficial effect on the intestinal microflora of a host when fed in the form of dried or fermented products to humans or animals in the form of single or multiple strains.
  • the cancer may include bladder cancer, breast cancer, melanoma, thyroid cancer, parathyroid cancer, rectal cancer, throat cancer, larynx cancer, esophageal cancer, pancreatic cancer, stomach cancer, snow cancer, skin cancer, brain tumor, uterine cancer, bile duct cancer, oral cancer, colon cancer, anal cancer, liver cancer, lung cancer, and It may be any one selected from the group consisting of colorectal cancer, preferably colorectal cancer, skin cancer, lung cancer, pancreatic cancer, liver cancer or bladder cancer, but is not limited thereto.
  • the Lactobacillus Sakeai WIKIM30 included in the composition according to the present invention may exist as a living cell or a living cell, and may also exist in a dried or lyophilized form. Forms and formulation methods of lactic acid bacteria suitable for inclusion in various compositions are well known to those skilled in the art.
  • composition can be administered orally or parenterally.
  • parenteral administration intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, endothelial administration, topical administration, intranasal administration, intrapulmonary administration, and rectal administration may be administered. It may be administered, but is not limited thereto.
  • Suitable dosages of the composition can be variously prescribed by factors such as the method of formulation, the mode of administration, the patient's age, weight, sex, morbidity, food, time of administration, route of administration, rate of excretion, and response sensitivity.
  • the pharmaceutical composition of the present invention may be prepared using a pharmaceutically acceptable and physiologically acceptable adjuvant in addition to the active ingredient, and the adjuvant includes excipients, boron Release agents, sweeteners, binders, coating agents, expanding agents, lubricants, lubricants or flavoring agents may be used.
  • the pharmaceutical composition may be preferably formulated into a pharmaceutical composition by including one or more pharmaceutically acceptable carriers in addition to the active ingredients described above for administration.
  • the active ingredient can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like.
  • suitable binders, lubricants, disintegrants and colorants can also be included in the mixture.
  • suitable binders include, but are not limited to, natural sugars such as starch, gelatin, glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, trachocanth or sodium oleate, sodium stearate, magnesium stearate, sodium benzo Eight, sodium acetate, sodium chloride, and the like.
  • Disintegrants include, but are not limited to, starch, methylcellulose, agar, bentonite, xanthan gum, and the like.
  • a composition formulated as a liquid solution sterile and biocompatible, saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol and
  • saline sterile water
  • Ringer's solution buffered saline
  • albumin injection solution dextrose solution
  • maltodextrin solution glycerol
  • ethanol ethanol
  • One or more of these components can be mixed and used, and other conventional additives such as antioxidants, buffers and bacteriostatic agents can be added as necessary.
  • diluents such as aqueous solutions, suspensions, emulsions, pills, capsules, granules or tablets.
  • composition comprising the Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) of the present invention or a culture thereof as an active ingredient may be used as a food composition or a food additive composition for preventing or improving cancer.
  • the food composition may be in the form of a health functional food.
  • the “health functional food” refers to food manufactured and processed using ingredients or ingredients with useful functionality in accordance with the Act on Health Functional Food (Article 3, No. 1), and “functional” is It means to control the nutrients for the structure and function of the human body or to obtain useful effects for health purposes such as physiological action (Article 2).
  • the food composition may further include a food additive, and whether or not it is suitable as a "food additive" is the standard for the relevant item and the standard and general test method of the food additives approved by the Korea Food and Drug Administration unless otherwise specified. Judging by the criteria.
  • Items listed in the "Food Additives Fair” include, for example, chemical compounds such as ketones, glycine, potassium citrate, nicotinic acid, and cinnamon acid, natural additives such as chromosomes, licorice extract, crystalline cellulose, and guar gum, L- And mixed preparations such as sodium glutamate, noodle-added alkalis, preservatives, and tar colorants.
  • chemical compounds such as ketones, glycine, potassium citrate, nicotinic acid, and cinnamon acid
  • natural additives such as chromosomes, licorice extract, crystalline cellulose, and guar gum
  • L- And mixed preparations such as sodium glutamate, noodle-added alkalis, preservatives, and tar colorants.
  • Foods containing the active ingredient of the present invention include breads, rice cakes, dried fruits, candy, chocolates, chewing gum, confectionary ice creams such as confections, ice creams, ice cream products such as ice cream powders, low fat milks, lactose milk, Processed milk, goat milk, fermented milk, butter oil, concentrated milk, milk cream, butter oil, natural cheese, processed cheese, milk powder, dairy products such as whey, meat processed products, egg processed products, meat products such as hamburger fish cake, ham, Fish products such as sausages, bacon, and other processed meat products such as ramen, dried noodles, raw noodles, fried noodles, luxurious dried noodles, improved noodles, frozen noodles, pasta, fruit fruits, vegetable drinks, carbonated drinks, soy milk, Beverages such as lactic acid bacteria drinks such as yogurt, soy sauce, miso, red pepper paste, chunjang, cheonggukjang, mixed vinegar, sauces, tomato ketchup, curry, seasoning foods such as curry, dressing, margarine, shorten
  • composition of the present invention includes various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, And carbonated agents used in carbonated beverages.
  • composition of the present invention may include flesh for the preparation of natural fruit juice, fruit juice beverages and vegetable beverages. These ingredients can be used independently or in combination.
  • the beverage composition containing the active ingredient of the present invention is not particularly limited to other ingredients, and may contain various flavoring agents or natural carbohydrates, etc. as additional ingredients, such as ordinary drinks.
  • natural carbohydrates described above include monosaccharides (eg, glucose, fructose, etc.); Disaccharides (eg maltose, sucrose, etc.); And polysaccharides, common sugars such as (eg, dextrin, cyclodextrin, etc.), and sugar alcohols such as xylitol, sorbitol, and erythritol.
  • natural flavoring agents such as taumatin and stevia extract (for example, rebaudioside A, glycyrrhizine)
  • synthetic flavoring agents sacharin, aspartame, etc.
  • composition comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) or a culture thereof as an active ingredient of the present invention may be used as a feed composition or feed additive composition for preventing or improving cancer in livestock. .
  • the composition When the composition is prepared as a feed additive, the composition may be 20 to 90% highly concentrated or may be prepared in powder or granular form.
  • the feed additives include organic acids such as citric acid, humic acid, adipic acid, lactic acid, and malic acid, phosphates such as sodium phosphate, potassium phosphate, acid pyrophosphate, polyphosphate (polymeric phosphate), polyphenols, catechins, alpha-tocopherols, and rosemary Extract, vitamin C, green tea extract, licorice extract, chitosan, tannic acid, phytinic acid, etc. may further include any one or more of natural antioxidants.
  • the composition When prepared as a feed, the composition may be formulated in a conventional feed form, and may include common feed ingredients.
  • the feed and feed additives include grains, such as crushed or crushed wheat, oats, barley, corn and rice; Vegetable protein feeds, such as feeds based on rape, soybeans, and sunflower; Animal protein feeds, such as blood meal, meat meal, bone meal and fish meal; Sugars and dairy products, for example, may further include dry ingredients made of various milk powders and whey powders, and may further include nutritional supplements, digestion and absorption enhancers, growth promoters, and the like.
  • grains such as crushed or crushed wheat, oats, barley, corn and rice
  • Vegetable protein feeds such as feeds based on rape, soybeans, and sunflower
  • Animal protein feeds such as blood meal, meat meal, bone meal and fish meal
  • Sugars and dairy products for example, may further include dry ingredients made of various milk powders and whey powders, and may further include nutritional supplements, digestion and absorption enhancers, growth promoters, and the like.
  • the feed additive may be administered to animals alone or in combination with other feed additives in an edible carrier.
  • the feed additives can be easily administered to animals as topdressing or by mixing them directly into animal feed or in a separate oral formulation from feed.
  • a pharmaceutically acceptable edible carrier can be solid or liquid, for example corn starch, lactose, sucrose, soy flakes, peanut oil, olive oil, sesame oil and propylene glycol.
  • the feed additive can be a tablet, capsule, powder, troche or sugar-coated tablet or a top dressing in undispersed form.
  • the feed additive may be a gelatin soft capsule, or a syrup or suspension, emulsion, or solution formulation.
  • the feed and feed additives may contain adjuvants, such as preservatives, stabilizers, wetting or emulsifying agents, solution accelerators, and the like.
  • the feed additive may be used by adding to animal feed by acupuncture, spraying, or mixing.
  • the feed or feed additive of the present invention is applicable to a number of animal diets including mammals, poultry and fish.
  • the mammal As the mammal, it can be used for pigs, cows, horses, sheep, rabbits, goats, rodents and experimental animals such as rats, hamsters, guinea pigs, pets (eg dogs, cats), and chickens as the poultry. It can also be used for turkey, duck, goose, pheasant, quail, and the like, and may be used as carp, crucian carp and trout, but is not limited thereto.
  • composition comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) or a culture thereof as an active ingredient of the present invention may provide a method for preventing or treating cancer administered to an individual except humans. .
  • Lactobacillus Sakeai WIKIM30 (Accession No. KCCM11878P) was pre-sale from the World Kimchi Research Institute Microbial Gene Bank.
  • a single colony of pre-sold Lactobacillus Sakeai WIKIM30 was passaged in MRS agar medium for 48 hours at 30° C., and the cultured single colony was inoculated in 10 ml of MRS liquid medium and shake cultured at 200 rpm for 24 hours at 37° C. . After incubation, the cells were centrifuged at 8,000 rpm for 5 minutes to remove the culture medium, and washed with PBS (Phosphate Buffered Saline) solution three times to remove the remaining medium components.
  • PBS Phosphate Buffered Saline
  • the experimental animals used in the experiment were supplied with a male 5 week old BALB/c mouse and a C57BL/6 mouse (Orient Bio, Korea), and an animal breeding room in an SPF environment where the room temperature was maintained at 20 ⁇ 2°C and humidity 55 ⁇ 15%. After a 1-week stabilization period, they were bred during the experimental period. The feed was fed with a general pellet feed without antibiotics, and water was allowed to be consumed from time to time. All animal breeding, experiments and euthanasia were conducted according to the protocol approved by the Animal Experimental Ethics Committee of the World Kimchi Research Institute. The observation of tumor size change was performed by measuring the volume of the tumor (mm 3 ) using the formula 3.14 x (length x height x width)/6.
  • CT26 mouse colon cancer cells (Korea Cell Line Bank, Korea) were purchased and used. CT26 mouse colon cancer cells were cultured in 5% CO 2 and 37° C. conditions using DMEM medium (Hyclone, USA) containing 10% fetal calf serum and 1% penicillin-streptomycin.
  • CT26 cell transplant animal model 6-week-old BALB/c mice (18-21 g) were used in the experiment.
  • Cultured mouse colon cancer cells CT26 were harvested 1x10 5 cells, resuspended in 50 ⁇ l PBS, and injected under the right thigh of the mouse.
  • the tail of a CT26 cell transplanted mouse with tumors formed in a volume of about 80-100 mm 3 was intravenously injected with Lactobacillus Sakeai WIKIM30.
  • a composition containing Lactobacillus Sakeai WIKIM30 in the CT26 cell transplanted mouse was prepared using PBS to prepare 1 ⁇ 10 9 CFU/0.1ml and 5 ⁇ 10 8 CFU/0.1ml.
  • the tumor-implanted mice were injected intravenously once or three times, and PBS was administered to the negative control group.
  • the results of visually confirming the change in tumor size of colorectal cancer in CT26 cell transplanted mice over time are shown in FIG. 1.
  • the results of measuring the volume change of tumor size in CT26 cell transplanted mice over time are shown in FIG. 2.
  • Example 5 Tumor size change of mouse skin melanoma B16F10 cells
  • B16F10 mouse melanoma cells (Korea Cell Line Bank, Korea) were purchased and used. B16F10 mouse melanoma cells were cultured at 5% CO 2 and 37° C. using DMEM medium (Hyclone, USA) containing 10% fetal calf serum and 1% penicillin-streptomycin.
  • mice For the production of the B16F10 cell transplant animal model, 6-week-old C57BL/6 mice (18-21 g) were used in the experiment.
  • the cultured mouse melanoma cells B16F10 were harvested from each 1x10 5 cell, resuspended in 50 ⁇ l PBS, and injected under the right thigh of the mouse.
  • the tail of a mouse transplanted with B16F10 cells formed with a tumor volume of about 80-100 mm 3 was intravenously injected with Lactobacillus Sakeai WIKIM30.
  • Lactobacillus Sakeai WIKIM30 was prepared by quantifying 1 ⁇ 10 10 CFU/mL bacteria using PBS, 0.1 mL (1 ⁇ 10 9 CFU) was injected intravenously into the experimental animals, and PBS was administered to the negative control group.
  • Fig. 3 shows the results of visually confirming the change in colorectal cancer tumor size of B16F10 cell transplanted mice over time.
  • Cell viability assay was measured using Cell counting Kit-8.
  • Cells used for the anticancer activity efficacy test include mouse-derived colon cancer CT26 cell line, human-derived colon cancer SW620 and HCT116 cell lines, human-derived non-small cell lung cancer H1650 cell line, human-derived pancreatic cancer ASPC1 and PANC1 cell lines, human-derived liver cancer HepG2 cell line, and human-derived bladder cancer The T24 cell line and the normal skin cell line CCD-986-sk cell line were used.
  • the mouse-derived colorectal cancer CT26 cell line was passaged at 5% CO 2 and 37° C. using DMEM medium containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin. It was tested using.
  • the cultured B16F10 cells were treated with 0.25% trypsin-EDTA in the incubator at 37°C for 3 minutes to detach the cells, washed twice with DPBS, and prepared at 1 X 10 4 cells/well.
  • SW620, HCT116, H1650, ASPC1, PANC1, HepG2 and T24 cell lines are 5% CO 2 using RPMI medium containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin. , it was tested using the growth phase cells subcultured at 37 °C condition. Cultured SW620, HCT116, H1650, ASPC1, PANC1, HepG2 and T24 cells were treated with 0.25% trypsin-EDTA for 3 minutes at 37°C in the incubator, detached and washed twice with DPBS, then 1 X 10 4 cells/well Prepared.
  • FBS fetal bovine serum
  • Human-derived normal skin cell CCD-986-sk cell line was passaged at 5% CO 2 at 37°C using RPMI medium containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin. Cultured and tested using cells in the growth phase. The cultured CCD-986-na cells were treated with 0.25% trypsin-EDTA in the incubator at 37°C for 3 minutes to detach the cells, washed twice with DPBS, and prepared at 1 X 10 4 cells/well.
  • FBS fetal bovine serum
  • the cell growth rate (cell viability assay) was treated with MOI1 concentration, and then cultured for 72 hours. After incubation, the cell counting Kit-8 reagent was treated and absorbance (OD 450 nm) was measured. Based on the absorbance of cells not treated with Lactobacillus Sakeai WIKIM30 (comtrol), the growth rate of cells treated with Lactobacillus Sakeai WIKIM30 was calculated, and the experimental results are described below.
  • a cell growth rate of 59.1% was measured in a human-derived non-small cell lung cancer H1650 cell line treated with Lactobacillus Sakeai WIKIM30. From the above results, it can be confirmed that in the case of the lung cancer cell line treated with Lactobacillus Sakeai WIKIM30, since the cell growth rate is significantly inhibited, it has excellent anticancer activity efficacy against lung cancer.
  • a cell growth rate of 18.8% was measured in a human-derived liver cancer HepG2 cell line treated with Lactobacillus Sakeai WIKIM30. From the above results, it can be confirmed that in the case of a liver cancer cell line treated with Lactobacillus Sakeai WIKIM30, cell growth rate is remarkably inhibited and thus has excellent anticancer activity efficacy against liver cancer.
  • a cell growth rate of 53.9% was measured in a bladder cancer T24 cell line derived from humans treated with Lactobacillus Sakeai WIKIM30. From the above results, it can be seen that in the case of the bladder cancer cell line treated with Lactobacillus Sakeai WIKIM30, the cell growth rate is significantly inhibited, and thus it has excellent anticancer activity efficacy against bladder cancer.
  • the two-way t-test determined statistically significant differences in tumor growth between the negative control group and the Lactobacillus Sakeai WIKIM30 treatment group. P ⁇ 0.05 was considered significantly for all tumor growth graph analysis.

Abstract

The present invention relates to novel Lactobacillus sakei WIKIM30 (accession number KCCM11878P) isolated from Kimchi, and a composition comprising same as an active ingredient. Exhibiting excellent anticancer activity in animal models into which cancer cells have been transplanted and cancer cell lines, Lactobacillus sakei WIKIM30 according to the present invention can be advantageously used as a composition for treatment, prevention, or alleviation of cancer in humans or animals.

Description

락토바실러스 사케아이 WIKIM30을 유효성분으로 포함하는 암의 예방 또는 치료용 약학 조성물Lactobacillus Sakeai WIKIM30 as an active ingredient for the prevention or treatment of cancer pharmaceutical composition
본 발명은 김치로부터 분리된 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, 기탁번호 KCCM11878P)을 유효성분으로 포함하는 암의 예방 또는 치료용 조성물에 관한 것이다.The present invention relates to a composition for preventing or treating cancer comprising Lactobacillus sakei (Accession No. KCCM11878P) isolated from kimchi as an active ingredient.
암은 세계적으로 높은 사망률을 보이며, 서구 사회에서는 심혈관 질환 다음으로 가장 일반적인 사망 원인이다. 특히, 식생활이 서구화되어 고지방식의 섭취가 일반화되고, 환경 오염 물질의 급격한 증가, 음주량의 증가 등으로 대장암, 유방암, 전립선암 등이 지속적으로 증가하는 추세에 있으며, 인구의 고령화와 더불어 흡연 인구의 증가 및 대기 오염으로 인해 폐암이 증가하고 있는 실정이다. 이러한 실정에서 암의 조기 예방 및 치료를 가능하게 하여 인간 건강의 증진, 건강한 삶의 질 향상 및 인류 보건 증진에 기여할 수 있는 항암 물질의 창출이 절실히 요구되고 있다.Cancer has a high mortality rate worldwide and is the most common cause of death after cardiovascular disease in Western societies. In particular, colonization, breast cancer, and prostate cancer are continuously increasing due to the rapid increase in environmental pollutants and increased alcohol consumption due to the westernization of eating habits, and the smoking population along with the aging of the population. Lung cancer is increasing due to the increase in and air pollution. In this situation, there is an urgent need for the creation of anti-cancer substances that can contribute to the promotion of human health, the improvement of healthy quality of life, and the promotion of human health by enabling the early prevention and treatment of cancer.
한편, 유산균은 인간과 동물의 구강, 장, 질, 분변, 그리고 김치와 같은 발효식품 등에 널리 분포하면서 사람과 동물의 건강과 밀접한 관련을 맺고 있다. 유산균은 정장작용, 유해균 억제, 면역조절, 혈중 콜레스테롤 저하, 항암 등 다양한 건강증진 효과를 나타내고 있다.On the other hand, lactic acid bacteria are widely distributed in humans and animals such as oral, intestinal, vaginal, fecal, and fermented foods such as kimchi, and are closely related to human and animal health. Lactic acid bacteria show various health promoting effects such as intestinal action, suppression of harmful bacteria, immune regulation, lowering of blood cholesterol, and anti-cancer.
현재, 한국공개특허 제10-2015-0068061호에서는 락토바실러스 플란타룸(Lactobacillus plantarum) PNU (KCCM11352P) 또는 락토바실러스 메센테로이드(Lactobacillus mesenteroides) PNU (KCCM11353P)의 항암 활성을 개시하고 있고, 한국등록특허 제10-1287120호에서는 락토바실러스 플란타륨 DSR CK10[기탁번호: KFCC-11433P] 또는 락토바실러스 플란타륨 DSR M2[기탁번호: KFCC-11432P]를 유효성분으로 함유하는 암 치료용 약학 조성물을 개시하고 있으나, 락토바실러스 사케아이에 관한 항암 활성 및 우수성에 대해서는 보고된 바 없다.Currently, Korean Patent Publication No. 10-2015-0068061 discloses anticancer activity of Lactobacillus plantarum PNU (KCCM11352P) or Lactobacillus mesenteroides PNU (KCCM11353P), and registered Korean patent No. 10-1287120 discloses a pharmaceutical composition for treating cancer containing Lactobacillus plantarium DSR CK10 [Accession No.: KFCC-11433P] or Lactobacillus plantarium DSR M2 [Accession No.: KFCC-11432P] as an active ingredient. However, no anti-cancer activity and superiority of Lactobacillus Sakeai have been reported.
본 발명의 목적은 락토바실러스 사케아이 균주를 유효성분으로 포함하는 암의 예방 또는 치료용 약학 조성물을 제공하기 위한 것이다.An object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of cancer comprising the Lactobacillus Sakeai strain as an active ingredient.
또한 본 발명의 다른 목적은 신규한 김치 유산균 락토바실러스 사케아이 균주를 유효성분으로 포함하는 암의 예방 또는 개선용 식품 조성물 또는 식품 첨가제 조성물을 제공하기 위한 것이다.In addition, another object of the present invention is to provide a food composition or a food additive composition for the prevention or improvement of cancer comprising the novel kimchi lactic acid bacteria Lactobacillus Sakeai strain as an active ingredient.
또한 본 발명의 또 다른 목적은 신규한 김치 유산균 락토바실러스 사케아이 균주를 유효성분으로 포함하는 암의 예방 또는 개선용 사료 조성물 또는 사료 첨가제 조성물을 제공하기 위한 것이다.In addition, another object of the present invention is to provide a feed composition or feed additive composition for the prevention or improvement of cancer, including the novel kimchi lactic acid bacteria Lactobacillus Sakeai strain as an active ingredient.
또한 본 발명의 또 다른 목적은 상기 조성물을 인간을 제외한 개체에 투여하는 암의 예방 또는 치료 방법을 제공하기 위한 것이다.Another object of the present invention is to provide a method for preventing or treating cancer, wherein the composition is administered to an individual other than a human.
이에, 본 발명자들은 김치로부터 프로바이오틱스로서 우수한 효과를 나타내면서 암의 예방 및 치료 효과를 나타내는 유산균 균주를 찾고자 노력한 결과, 항암 활성을 갖는 유산균 균주인 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30)을 분리, 동정하여 본 발명을 완성하게 되었다.Thus, the present inventors tried to find a lactic acid bacteria strain that shows the prevention and treatment effect of cancer while showing excellent effects as a probiotic from kimchi, and isolated and identified Lactobacillus sakei, WIKIM30, Lactobacillus sakei, WIKIM30, a lactic acid bacteria strain having anticancer activity. Thus, the present invention has been completed.
상기 목적을 달성하기 위하여, 락토바실러스 사케아이 WIKIM30을 유효성분으로 포함하는 암의 예방 또는 치료용 약학 조성물을 제공한다.In order to achieve the above object, to provide a pharmaceutical composition for preventing or treating cancer comprising Lactobacillus Sakeai WIKIM30 as an active ingredient.
또한, 본 발명은 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 암의 예방 또는 개선용 식품 조성물 또는 식품 첨가제 조성물을 제공한다.In addition, the present invention provides a food composition or food additive composition for preventing or improving cancer comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a culture thereof as an active ingredient.
또한, 본 발명은 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 가축의 암 예방 또는 개선용 사료 조성물 또는 사료 첨가제 조성물을 제공한다.In addition, the present invention provides a feed composition or feed additive composition for preventing or improving cancer in livestock, comprising Lactobacillus sakei WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a culture thereof as an active ingredient.
또한 본 발명의 또 다른 목적은 상기 조성물을 인간을 제외한 개체에 투여하는 암의 예방 또는 치료 방법을 제공한다.In addition, another object of the present invention provides a method for preventing or treating cancer, wherein the composition is administered to an individual other than a human.
본 발명에 따른 락토바실러스 사케아이 WIKIM30은 암 세포가 이식된 동물모델 및 암 세포주에서 우수한 항암 활성을 나타내므로, 사람 또는 동물의 암의 치료, 예방 또는 개선 등의 용도를 위한 조성물로서 유용하게 이용될 수 있다.Since Lactobacillus Sakeai WIKIM30 according to the present invention exhibits excellent anticancer activity in animal models and cancer cell lines in which cancer cells are transplanted, it can be usefully used as a composition for the treatment, prevention or improvement of cancer in humans or animals Can be.
도 1은 본 발명에 따른 조성물을 CT26 세포 이식 마우스에 1회(Single) 또는 3회(Serial) 정맥 주사한 후 시간이 경과함에 따라 종양 세포의 크기가 변화하는 것을 관찰하기 위한 마우스의 종양 부분 사진을 나타낸 도이다. 1 is a photograph of a tumor portion of a mouse for observing a change in the size of a tumor cell over time after intravenous injection of a composition according to the present invention to a CT26 cell transplanted mouse once (Single) or three times (Serial) It is a diagram showing.
도 2는 본 발명에 따른 조성물을 CT26 세포 이식 마우스에 1회(Single) 또는 3회(Serial) 정맥 주사한 후 시간이 경과함에 따라 종양 세포의 부피 변화를 측정한 그래프를 나타낸 도이다. 2 is a view showing a graph of measuring the change in volume of tumor cells over time after a single (or single) or three (Serial) intravenous injection of CT26 cell-grafted mice according to the present invention.
도 3은 본 발명에 따른 조성물을 B16F10 피부 흑색종 마우스에 정맥 주사한 후 시간이 경과함에 따라 종양 세포의 크기가 변화하는 것을 관찰하기 위한 마우스의 종양 부분 사진을 나타낸 도이다. 3 is a view showing a photograph of a tumor portion of a mouse for observing that the size of tumor cells changes over time after intravenous injection of a composition according to the present invention into a B16F10 skin melanoma mouse.
도 4는 본 발명에 따른 조성물을 B16F10 피부 흑색종 마우스에 정맥 주사한 후 시간이 경과함에 따라 종양 세포의 부피 변화를 측정한 그래프를 나타낸 도이다. FIG. 4 is a graph showing a change in the volume of tumor cells over time after intravenous injection of a composition according to the present invention into a B16F10 skin melanoma mouse.
도 5는 마우스 유래 대장암 CT26 세포주, 사람 유래 대장암 SW620 세포주 및 HCT116 세포주에 본 발명에 따른 조성물을 락토바실러스 사케아이 WIKIM30을 처리하고 세포 성장률(cell viability assay)을 측정한 그래프를 나타낸 도이다.FIG. 5 is a graph showing the composition of the present invention treated with Lactobacillus Sakeai WIKIM30 in mouse-derived colorectal cancer CT26 cell line, human-derived colorectal cancer SW620 cell line, and HCT116 cell line, and measuring cell viability assay.
도 6은 사람 유래 비소폐암세포 H1650 세포주에 락토바실러스 사케아이 WIKIM30을 처리하고 세포 성장률(cell viability assay)을 측정한 그래프를 나타낸 도이다.6 is a graph showing the measurement of cell viability assay by treating Lactobacillus Sakeai WIKIM30 in a human-derived non-small lung cancer cell H1650 cell line.
도 7은 사람 유래 췌장암 ASPC1 세포주 및 PANC1 세포주에 락토바실러스 사케아이 WIKIM30을 처리하고 세포 성장률(cell viability assay)을 측정한 그래프를 나타낸 도이다.FIG. 7 is a graph showing the treatment of Lactobacillus Sakeai WIKIM30 on human-derived pancreatic cancer ASPC1 cell line and PANC1 cell line and measuring cell viability assay.
도 8은 사람 유래 간암 HepG2 세포주에 락토바실러스 사케아이 WIKIM30을 처리하고 세포 성장률(cell viability assay)을 측정한 그래프를 나타낸 도이다.8 is a diagram showing a graph of measuring Lactobacillus Sakeai WIKIM30 in a human-derived liver cancer HepG2 cell line and measuring cell viability assay.
도 9는 사람 유래 방광암 T24 세포주에 락토바실러스 사케아이 WIKIM30을 처리하고 세포 성장률(cell viability assay)을 측정한 그래프를 나타낸 도이다.9 is a view showing a graph of measuring Lactobacillus Sakeai WIKIM30 in a human-derived bladder cancer T24 cell line and measuring cell viability assay.
도 10은 사람 유래 정상피부 CCD-986-sk 세포주에 락토바실러스 사케아이 WIKIM30을 처리하고 세포 성장률(cell viability assay)을 측정한 그래프를 나타낸 도이다.FIG. 10 is a diagram showing a graph obtained by treating Lactobacillus Sakeai WIKIM30 on a human-derived normal skin CCD-986-sk cell line and measuring cell viability assay.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하고자 한다. 이들 실시예는 오로지 본 발명을 보다 구체적으로 설명하기 위한 것으로서, 본 발명의 요지에 따라 본 발명의 범위가 이들 실시예에 의해 제한되지 않는다는 것은 당업계에서 통상의 지식을 가진 자에게 있어서 자명할 것이다.Hereinafter, the present invention will be described in more detail through examples. These examples are only for explaining the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples according to the gist of the present invention. .
본 발명의 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 조성물은 암의 예방 또는 치료 효과를 가지며, 약제학적 조성물로 이용될 수 있다.The composition comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) or a culture thereof as an active ingredient of the present invention has an effect of preventing or treating cancer and can be used as a pharmaceutical composition.
락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30)은 김치 유래의 유산균 균주이다. 비록 본 발명에서의 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30)을 김치에서 분리, 동정하기는 했으나, 이의 입수 경로가 이에 한정되는 것은 아니다.Lactobacillus sakei (WIKIM30) is a lactic acid bacteria strain derived from kimchi. Although Lactobacillus sakei (WIKIM30) of Lactobacillus sake in the present invention was separated and identified from kimchi, the route for obtaining it is not limited thereto.
상기 락토바실러스 사케아이 WIKIM30의 동정 및 분류를 위한 16S rRNA 염기서열 분석 결과, 서열번호 1(SEQ ID NO: 1)의 핵산서열을 갖는 것으로 나타났다.As a result of sequencing of 16S rRNA for identification and classification of the Lactobacillus Sakeai WIKIM30, it was found to have the nucleic acid sequence of SEQ ID NO: 1 (SEQ ID NO: 1).
따라서, 서열번호 1(SEQ ID NO: 1)의 16S rRNA 염기서열을 갖는 본 발명의 미생물을 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei)로 명명하였으며, 한국미생물보존센터에 2015년 9월 10일자로 기탁하였다(기탁번호 KCCM11878P).Therefore, the microorganism of the present invention having the 16S rRNA nucleotide sequence of SEQ ID NO: 1 (SEQ ID NO: 1) was named Lactobacillus sake WIKIM30 (Lactobacillus sakei), and deposited with the Korea Microbiological Conservation Center on September 10, 2015 (Accession No. KCCM11878P).
본 발명의 락토바실러스 사케아이 WIKIM30은 프로바이오틱스로서, 유산균의 일반적인 정장 효과 및 면역 증강 효과를 갖는다. 락토바실러스 속의 유산균이 정장 효과 면역 증강 효과를 갖는 것은 잘 알려진 사실이다.Lactobacillus sakeai WIKIM30 of the present invention is a probiotic, and has a general suit effect and an immune enhancing effect of lactic acid bacteria. It is well known that lactic acid bacteria in the genus Lactobacillus have a formal effect immune enhancing effect.
본 발명에 있어서, '프로바이오틱스(probiotics)'는 사람을 포함한 동물의 위장관 내에서 숙주의 장내 미생물 환경을 개선하여 숙주의 건강에 유익한 영향을 주는 살아있는 미생물'이라는 의미로 이해된다. 프로바이오틱스는 프로바이오틱 활성을 갖는 살아있는 미생물로 단일 또는 복합균주 형태로 사람이나 동물에 건조된 세포 형태나 발효산물 형태로 급여될 경우, 숙주의 장내 균총에 유익한 영향을 미칠 수 있다.In the present invention,'probiotics (probiotics)' is understood to mean a living microorganism that has a beneficial effect on the health of the host by improving the intestinal microbial environment of the host in the gastrointestinal tract of animals, including humans. Probiotics, which are living microorganisms with probiotic activity, may have a beneficial effect on the intestinal microflora of a host when fed in the form of dried or fermented products to humans or animals in the form of single or multiple strains.
상기 암은 방광암, 유방암, 흑색종양, 갑상선암, 부갑상선암, 직장암, 인후암, 후두암, 식도암, 췌장암, 위암, 설암, 피부암, 뇌종양, 자궁암, 두담낭암, 구강암, 결장암, 항문 부근암, 간암, 폐암 및 대장암으로 이루어지는 군으로부터 선택되는 어느 하나의 암일 수 있으며, 바람직하게는 대장암, 피부암, 폐암, 췌장암, 간암 또는 방광암일 수 있으나, 이에 한정되지 않는다.The cancer may include bladder cancer, breast cancer, melanoma, thyroid cancer, parathyroid cancer, rectal cancer, throat cancer, larynx cancer, esophageal cancer, pancreatic cancer, stomach cancer, snow cancer, skin cancer, brain tumor, uterine cancer, bile duct cancer, oral cancer, colon cancer, anal cancer, liver cancer, lung cancer, and It may be any one selected from the group consisting of colorectal cancer, preferably colorectal cancer, skin cancer, lung cancer, pancreatic cancer, liver cancer or bladder cancer, but is not limited thereto.
본 발명에 따른 조성물에 포함되는 락토바실러스 사케아이 WIKIM30은 생균체 또는 사균체로서 존재할 수 있으며, 또한 건조 또는 동결건조된 형태로 존재할 수도 있다. 다양한 조성물 내에 포함시키기 적합한 유산균의 형태 및 제제화 방법은 당업자에게 잘 알려져 있다.The Lactobacillus Sakeai WIKIM30 included in the composition according to the present invention may exist as a living cell or a living cell, and may also exist in a dried or lyophilized form. Forms and formulation methods of lactic acid bacteria suitable for inclusion in various compositions are well known to those skilled in the art.
상기 조성물은 경구 또는 비경구로 투여할 수 있다. 비경구 투여인 경우에는 정맥 내 주입, 피하 주입, 근육 주입, 복강 주입, 내피 투여, 국소 투여, 비내 투여, 폐 내 투여 및 직장 내 투여 등으로 투여할 수 있으며, 바람직하게는 정맥 내 주입방법으로 투여할 수 있으나, 이에 한정되지 않는다.The composition can be administered orally or parenterally. In the case of parenteral administration, intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, endothelial administration, topical administration, intranasal administration, intrapulmonary administration, and rectal administration may be administered. It may be administered, but is not limited thereto.
상기 조성물의 적합한 투여량은 제제화 방법, 투여 방식, 환자의 연령, 체중, 성, 병적 상태, 음식, 투여시간, 투여경로, 배설 속도 및 반응 감응성과 같은 요인들에 의해 다양하게 처방될 수 있다.Suitable dosages of the composition can be variously prescribed by factors such as the method of formulation, the mode of administration, the patient's age, weight, sex, morbidity, food, time of administration, route of administration, rate of excretion, and response sensitivity.
본 발명의 조성물이 약제학적 조성물로 활용될 경우, 본 발명의 약제학적 조성물은 상기 유효성분 이외에 약제학적으로 적합하고 생리학적으로 허용되는 보조제를 사용하여 제조될 수 있으며, 상기 보조제로는 부형제, 붕해제, 감미제, 결합제, 피복제, 팽창제, 윤활제, 활택제 또는 향미제 등을 사용할 수 있다.When the composition of the present invention is utilized as a pharmaceutical composition, the pharmaceutical composition of the present invention may be prepared using a pharmaceutically acceptable and physiologically acceptable adjuvant in addition to the active ingredient, and the adjuvant includes excipients, boron Release agents, sweeteners, binders, coating agents, expanding agents, lubricants, lubricants or flavoring agents may be used.
상기 약제학적 조성물은 투여를 위해서 상기 기재한 유효성분 이외에 추가로 약제학적으로 허용가능한 담체를 1종 이상 포함하여 약제학적 조성물로 바람직하게 제제화할 수 있다.The pharmaceutical composition may be preferably formulated into a pharmaceutical composition by including one or more pharmaceutically acceptable carriers in addition to the active ingredients described above for administration.
예를 들어, 정제 또는 캡슐제의 형태로의 제제화를 위해, 유효성분은 에탄올, 글리세롤, 물 등과 같은 경구, 무독성의 약제학적으로 허용가능한 불활성 담체와 결합될 수 있다. 또한, 원하거나 필요한 경우, 적합한 결합제, 윤활제, 붕해제 및 발색제 또한 혼합물로 포함될 수 있다. 적합한 결합제는 이에 제한되는 것은 아니나, 녹말, 젤라틴, 글루코스 또는 베타-락토오스와 같은 천연당, 옥수수감미제, 아카시아, 트래커캔스 또는 소듐올레이트와 같은 천연 및 합성검, 소듐 스테아레이트, 마그네슘 스테아레이트, 소듐 벤조에이트, 소듐아세테이트, 소듐클로라이드 등을 포함한다. 붕해제는 이에 제한되는 것은 아니나, 녹말, 메틸셀룰로스, 아가, 벤토니트, 잔탄검 등을 포함한다. 액상 용액으로 제제화되는 조성물에 있어서 허용가능한 약제학적 담체로는, 멸균 및 생체에 적합한 것으로서, 식염수, 멸균수, 링거액, 완충식염수, 알부민 주사 용액, 덱스트로즈 용액, 말토덱스트린 용액, 글리세롤, 에탄올 및 이들 성분 중 1 성분이상을 혼합하여 사용할 수 있으며, 필요에 따라 항산화제, 완충액, 정균제 등 다른 통상의 첨가제를 첨가할 수 있다. 또한 희석제, 분산제, 계면활성제, 결합제 및 윤활제를 부가적으로 첨가하여 수용액, 현탁액, 유탁액 등과 같은 주사용 제형, 환약, 캡슐, 과립 또는 정제로 제제화할 수 있다.For example, for formulation in the form of tablets or capsules, the active ingredient can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. In addition, if desired or necessary, suitable binders, lubricants, disintegrants and colorants can also be included in the mixture. Suitable binders include, but are not limited to, natural sugars such as starch, gelatin, glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, trachocanth or sodium oleate, sodium stearate, magnesium stearate, sodium benzo Eight, sodium acetate, sodium chloride, and the like. Disintegrants include, but are not limited to, starch, methylcellulose, agar, bentonite, xanthan gum, and the like. As an acceptable pharmaceutical carrier in a composition formulated as a liquid solution, sterile and biocompatible, saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol and One or more of these components can be mixed and used, and other conventional additives such as antioxidants, buffers and bacteriostatic agents can be added as necessary. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to formulate into injectable formulations such as aqueous solutions, suspensions, emulsions, pills, capsules, granules or tablets.
나아가 해당 분야의 적절한 방법으로 Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA에 개시되어 있는 방법을 이용하여 각 질환에 따라 또는 성분에 따라 바람직하게 제제화할 수 있다.Furthermore, it can be preferably formulated according to each disease or component using methods disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA by appropriate methods in the field.
본 발명의 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 조성물은 암의 예방 또는 개선용 식품 조성물 또는 식품 첨가제 조성물로 이용될 수 있다.The composition comprising the Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) of the present invention or a culture thereof as an active ingredient may be used as a food composition or a food additive composition for preventing or improving cancer.
상기 식품 조성물은 건강기능식품의 형태일 수 있다.The food composition may be in the form of a health functional food.
상기 "건강기능식품"은 건강기능식품에 관한 법률에 따라 인체에 유용한 기능성을 가진 원료나 성분을 사용하여 제조 및 가공한 식품을 의미(제3조 제1호)하며, "기능성"이라 함은 인체의 구조 및 기능에 대하여 영양소를 조절하거나 생리학적 작용 등과 같은 보건 용도에 유용한 효과를 얻는 것(동조 제2호)을 의미한다.The "health functional food" refers to food manufactured and processed using ingredients or ingredients with useful functionality in accordance with the Act on Health Functional Food (Article 3, No. 1), and "functional" is It means to control the nutrients for the structure and function of the human body or to obtain useful effects for health purposes such as physiological action (Article 2).
상기 식품 조성물은 식품 첨가물을 추가로 포함할 수 있으며, "식품첨가물"로서의 적합여부는 다른 규정이 없는 한 식품의약품안정청에 승인된 식품첨가물공전의 총칙 및 일반시험법 등에 따라 해당 품목에 관한 규격 및 기준에 의하여 판정한다.The food composition may further include a food additive, and whether or not it is suitable as a "food additive" is the standard for the relevant item and the standard and general test method of the food additives approved by the Korea Food and Drug Administration unless otherwise specified. Judging by the criteria.
상기 "식품첨가물공전"에 수재된 품목으로 예를 들어, 케톤류, 글리신, 구연산칼륨, 니코틴산, 계피산 등의 화학적 합성품, 감색소, 감초추출물, 결정셀롤로오스, 구아검 등의 천연첨가물, L-글루타민산나트륨제제, 면류첨가알칼리제, 보존료제제, 타르색소제제 등의 혼합 제제류 들을 들 수 있다.Items listed in the "Food Additives Fair" include, for example, chemical compounds such as ketones, glycine, potassium citrate, nicotinic acid, and cinnamon acid, natural additives such as chromosomes, licorice extract, crystalline cellulose, and guar gum, L- And mixed preparations such as sodium glutamate, noodle-added alkalis, preservatives, and tar colorants.
본 발명의 유효성분이 포함된 식품으로는 빵, 떡류, 건과류, 캔디류, 초콜릿류, 츄잉껌, 쨈류와 같은 과자류 아이스크림류, 빙과류, 아이스크림 분말류와 같은 아이스크림 제품류 우유류, 저지방 우유류, 유당분해 우유, 가공유류, 산양유, 발효유류, 버터유류, 농축유류, 유크림류, 버터유, 자연치즈, 가공치즈, 분유류, 유청류와 같은 유가공품류 식육가공품, 알가공품, 햄버거와 같은 식육제품류 어묵, 햄, 소세지, 베이컨 등의 어육가공품과 같은 어육제품류 라면류, 건면류, 생면류, 유탕면류, 호화건먼류, 개량숙면류, 냉동면류, 파스타류와 같은 면류 과실음료, 채소류음료, 탄산음료, 두유류, 요구르트 등의 유산균음료, 혼합음료와 같은 음료 간장, 된장, 고추장, 춘장, 청국장, 혼합장, 식초, 소스류, 토마토케첩, 카레, 드레싱과 같은 조미식품 마가린, 쇼트닝 및 피자를 들 수 있으나, 이에 한정되는 것은 아니다.Foods containing the active ingredient of the present invention include breads, rice cakes, dried fruits, candy, chocolates, chewing gum, confectionary ice creams such as confections, ice creams, ice cream products such as ice cream powders, low fat milks, lactose milk, Processed milk, goat milk, fermented milk, butter oil, concentrated milk, milk cream, butter oil, natural cheese, processed cheese, milk powder, dairy products such as whey, meat processed products, egg processed products, meat products such as hamburger fish cake, ham, Fish products such as sausages, bacon, and other processed meat products such as ramen, dried noodles, raw noodles, fried noodles, luxurious dried noodles, improved noodles, frozen noodles, pasta, fruit fruits, vegetable drinks, carbonated drinks, soy milk, Beverages such as lactic acid bacteria drinks such as yogurt, soy sauce, miso, red pepper paste, chunjang, cheonggukjang, mixed vinegar, sauces, tomato ketchup, curry, seasoning foods such as curry, dressing, margarine, shortening and pizza, but not limited to this It does not work.
상기 외에 본 발명의 조성물은 여러 가지 영양제, 비타민, 전해질, 풍미제, 착색제, 펙트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알코올, 탄산음료에 사용되는 탄산화제 등을 포함할 수 있다. 그밖에 본 발명의 조성물은 천연 과일주스, 과일주스 음료 및 야채 음료의 제조를 위한 과육을 포함할 수 있다. 이러한 성분은 독립적으로 또는 조합하여 사용할 수 있다. In addition to the above, the composition of the present invention includes various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, And carbonated agents used in carbonated beverages. In addition, the composition of the present invention may include flesh for the preparation of natural fruit juice, fruit juice beverages and vegetable beverages. These ingredients can be used independently or in combination.
본 발명의 유효성분을 포함한 음료 조성물은 다른 성분에는 특별한 제한이 없으며 통상의 음료와 같이 여러 가지 향미제 또는 천연 탄수화물 등을 추가성분으로 함유할 수 있다. 상술한 천연 탄수화물의 예는 모노사카라이드, (예를 들어, 포도당, 과당 등); 디사카라이드, (예를 들어 말토스, 슈크로스 등); 및 폴리사카라이드, (예를 들어 덱스트린, 시클로덱스트린 등)과 같은 통상적인 당, 및 자일리톨, 소르비톨, 에리트리톨 등의 당알콜이다. 상술한 것 이외의 향미제로서 천연 향미제(타우마틴, 스테비아 추출물(예를 들어 레바우디오시드 A, 글리시르히진 등)) 및 합성 향미제(사카린, 아스파르탐 등)를 유리하게 사용할 수 있다.The beverage composition containing the active ingredient of the present invention is not particularly limited to other ingredients, and may contain various flavoring agents or natural carbohydrates, etc. as additional ingredients, such as ordinary drinks. Examples of natural carbohydrates described above include monosaccharides (eg, glucose, fructose, etc.); Disaccharides (eg maltose, sucrose, etc.); And polysaccharides, common sugars such as (eg, dextrin, cyclodextrin, etc.), and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those described above, natural flavoring agents (such as taumatin and stevia extract (for example, rebaudioside A, glycyrrhizine)) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. have.
또한, 본 발명의 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 조성물은 가축의 암 예방 또는 개선용 사료 조성물 또는 사료 첨가제 조성물로 이용될 수 있다.In addition, the composition comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) or a culture thereof as an active ingredient of the present invention may be used as a feed composition or feed additive composition for preventing or improving cancer in livestock. .
상기 조성물이 사료 첨가제로서 제조될 경우, 상기 조성물은 20 내지 90% 고농축액이거나 분말 또는 과립 형태로 제조될 수 있다. 상기 사료 첨가제는 구연산, 후말산, 아디픽산, 젖산, 사과산등의 유기산이나 인산 나트륨, 인산 칼륨, 산성 피로인산염, 폴리인산염(중합인산염) 등의 인산염이나, 폴리페놀, 카테킨, 알파-토코페롤, 로즈마리 추출물, 비타민 C, 녹차 추출물, 감초 추출물, 키토산, 탄닌산, 피틴산 등의 천연 항산화제 중 어느 하나 또는 하나 이상을 추가로 포함할 수 있다. 사료로서 제조될 경우, 상기 조성물은 통상의 사료 형태로 제제화될 수 있으며, 통상의 사료 성분을 함께 포함할 수 있다.When the composition is prepared as a feed additive, the composition may be 20 to 90% highly concentrated or may be prepared in powder or granular form. The feed additives include organic acids such as citric acid, humic acid, adipic acid, lactic acid, and malic acid, phosphates such as sodium phosphate, potassium phosphate, acid pyrophosphate, polyphosphate (polymeric phosphate), polyphenols, catechins, alpha-tocopherols, and rosemary Extract, vitamin C, green tea extract, licorice extract, chitosan, tannic acid, phytinic acid, etc. may further include any one or more of natural antioxidants. When prepared as a feed, the composition may be formulated in a conventional feed form, and may include common feed ingredients.
상기 사료 및 사료 첨가제는 곡물, 예를 들면 분쇄 또는 파쇄된 밀, 귀리, 보리, 옥수수 및 쌀; 식물성 단백질 사료, 예를 들면 평지, 콩, 및 해바라기를 주성분으로 하는 사료; 동물성 단백질 사료, 예를 들면 혈분, 육분, 골분 및 생선분; 당분 및 유제품, 예를 들면 각종 분유 및 유장 분말로 이루어지는 건조성분 등을 더 포함할 수 있으며, 이외에도 영양보충제, 소화 및 흡수향상제, 성장촉진제 등을 더 포함할 수 있다.The feed and feed additives include grains, such as crushed or crushed wheat, oats, barley, corn and rice; Vegetable protein feeds, such as feeds based on rape, soybeans, and sunflower; Animal protein feeds, such as blood meal, meat meal, bone meal and fish meal; Sugars and dairy products, for example, may further include dry ingredients made of various milk powders and whey powders, and may further include nutritional supplements, digestion and absorption enhancers, growth promoters, and the like.
상기 사료 첨가제는 동물에게 단독으로 투여하거나 식용 담체 중에서 다른 사료 첨가제와 조합하여 투여할 수도 있다. 또한, 상기 사료 첨가제는 탑드레싱으로서 또는 이들을 동물사료에 직접 혼합하거나 또는 사료와 별도의 경구제형으로 용이하게 동물에게 투여할 수 있다. 상기 사료 첨가제를 동물사료와 별도로 투여할 경우, 당해 기술분야에 잘 알려진 바와 같이 약제학적으로 허용 가능한 식용 담체와 조합하여, 즉시 방출 또는 서방성 제형으로 제조할 수 있다. 이러한 식용 담체는 고체 또는 액체, 예를 들어 옥수수전분, 락토오스, 수크로오스, 콩플레이크, 땅콩유, 올리브유, 참깨유 및 프로필렌글리콜일 수 있다. 고체 담체가 사용될 경우, 사료 첨가제는 정제, 캡슐제, 산제, 트로키제 또는 함당정제 또는 미분산성 형태의 탑 드레싱일 수 있다. 액체 담체가 사용될 경우, 사료 첨가제는 젤라틴 연질 캡슐제, 또는 시럽제나 현탁액, 에멀젼제, 또는 용액제의 제형일 수 있다.The feed additive may be administered to animals alone or in combination with other feed additives in an edible carrier. In addition, the feed additives can be easily administered to animals as topdressing or by mixing them directly into animal feed or in a separate oral formulation from feed. When the feed additive is administered separately from the animal feed, it can be prepared in an immediate release or sustained release formulation in combination with a pharmaceutically acceptable edible carrier, as is well known in the art. Such edible carriers can be solid or liquid, for example corn starch, lactose, sucrose, soy flakes, peanut oil, olive oil, sesame oil and propylene glycol. When a solid carrier is used, the feed additive can be a tablet, capsule, powder, troche or sugar-coated tablet or a top dressing in undispersed form. When a liquid carrier is used, the feed additive may be a gelatin soft capsule, or a syrup or suspension, emulsion, or solution formulation.
또한, 상기 사료 및 사료 첨가제는 보조제, 예를 들어 보존제, 안정화제, 습윤제 또는 유화제, 용액 촉진제 등을 함유할 수 있다. 상기 사료 첨가제는 침주, 분무 또는 혼합하여 동물의 사료에 첨가하여 이용될 수 있다.In addition, the feed and feed additives may contain adjuvants, such as preservatives, stabilizers, wetting or emulsifying agents, solution accelerators, and the like. The feed additive may be used by adding to animal feed by acupuncture, spraying, or mixing.
본 발명의 사료 또는 사료 첨가제는 포유류, 가금 및 어류를 포함하는 다수의 동물 식이에 적용할 수 있다.The feed or feed additive of the present invention is applicable to a number of animal diets including mammals, poultry and fish.
상기 포유류로서 돼지, 소, 말, 양, 토끼, 염소, 설치동물 및 실험용 설치동물인 쥐, 햄스터, 기니피그뿐만 아니라, 애완동물(예: 개, 고양이) 등에게 사용할 수 있으며, 상기 가금류로서 닭, 칠면조, 오리, 거위, 꿩, 및 메추라기 등에도 사용할 수 있고, 상기 어류로서 잉어, 붕어 및 송어 등에 이용될 수 있으나, 이에 한정되는 것은 아니다.As the mammal, it can be used for pigs, cows, horses, sheep, rabbits, goats, rodents and experimental animals such as rats, hamsters, guinea pigs, pets (eg dogs, cats), and chickens as the poultry. It can also be used for turkey, duck, goose, pheasant, quail, and the like, and may be used as carp, crucian carp and trout, but is not limited thereto.
또한, 본 발명의 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 조성물은 인간을 제외한 개체에 투여하는 암의 예방 또는 치료 방법을 제공할 수 있다.In addition, the composition comprising Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, Accession No. KCCM11878P) or a culture thereof as an active ingredient of the present invention may provide a method for preventing or treating cancer administered to an individual except humans. .
실시예 1. 균주의 분리 및 배양Example 1. Isolation and culture of strain
락토바실러스 사케아이 WIKIM30(기탁번호 KCCM11878P)은 세계김치연구소 미생물 유전자은행으로부터 분양받아 실험을 진행하였다.Lactobacillus Sakeai WIKIM30 (Accession No. KCCM11878P) was pre-sale from the World Kimchi Research Institute Microbial Gene Bank.
분양받은 락토바실러스 사케아이 WIKIM30의 단일 콜로니를 MRS agar 배지에 계대하여 30℃에서 48시간 동안 배양한 후 배양된 단일 콜로니를 MRS 액체배지 10㎖에 접종하여 37℃에서 24시간 동안 200rpm으로 진탕배양하였다. 배양 후, 균체를 8,000rpm에서 5분 동안 원심분리하여 배양액을 제거하고, PBS(Phosphate Buffered Saline)용액으로 3회 세척하여 남아 있는 배지 성분을 제거하였다.A single colony of pre-sold Lactobacillus Sakeai WIKIM30 was passaged in MRS agar medium for 48 hours at 30° C., and the cultured single colony was inoculated in 10 ml of MRS liquid medium and shake cultured at 200 rpm for 24 hours at 37° C. . After incubation, the cells were centrifuged at 8,000 rpm for 5 minutes to remove the culture medium, and washed with PBS (Phosphate Buffered Saline) solution three times to remove the remaining medium components.
실시예 2. 실험동물(마우스)의 조건Example 2. Experimental animal (mouse) conditions
실험에 사용된 실험동물은 수컷 5주령의 BALB/c 마우스, C57BL/6 마우스(오리엔트바이오, 한국)를 공급받았고, 실내 온도 20±2℃, 습도 55±15%가 유지되는 SPF 환경의 동물 사육실에서 1주일의 안정화 기간을 거쳐 실험기간 동안 사육하였다. 사료는 항생제가 첨가되지 않은 일반적인 팰렛 사료를 공급하였고, 물은 수시로 섭취할 수 있게 하였다. 실험 진행은 세계김치연구소 동물실험윤리위원회에서 승인된 프로토콜에 따라 모든 동물 사육, 실험 및 안락사를 실시하였다. 종양 크기 변화의 관찰은 3.14 x (길이 x 높이 x 넓이)/6의 식을 사용하여 종양의 부피(mm3)를 측정하여 진행하였다.The experimental animals used in the experiment were supplied with a male 5 week old BALB/c mouse and a C57BL/6 mouse (Orient Bio, Korea), and an animal breeding room in an SPF environment where the room temperature was maintained at 20±2℃ and humidity 55±15%. After a 1-week stabilization period, they were bred during the experimental period. The feed was fed with a general pellet feed without antibiotics, and water was allowed to be consumed from time to time. All animal breeding, experiments and euthanasia were conducted according to the protocol approved by the Animal Experimental Ethics Committee of the World Kimchi Research Institute. The observation of tumor size change was performed by measuring the volume of the tumor (mm 3 ) using the formula 3.14 x (length x height x width)/6.
실시예 3. CT26 마우스 대장암 세포 이식 동물모델의 제작Example 3. Construction of CT26 mouse colon cancer cell transplant animal model
3-1. 세포배양3-1. Cell culture
CT26 생쥐 대장암 세포(한국세포주은행, 한국)는 구입하여 사용하였다. CT26 생쥐 대장암 세포는 10% 우태아혈청과 1% 페니실린-스트렙토마이신(penicillin-streptomycin)을 포함한 DMEM 배지(Hyclone, 미국)를 사용하여 5% CO2, 37℃ 조건에서 배양하였다. CT26 mouse colon cancer cells (Korea Cell Line Bank, Korea) were purchased and used. CT26 mouse colon cancer cells were cultured in 5% CO 2 and 37° C. conditions using DMEM medium (Hyclone, USA) containing 10% fetal calf serum and 1% penicillin-streptomycin.
3-2. 세포 이식 암 동물모델의 제작3-2. Production of cell transplant cancer animal models
CT26 세포 이식 동물모델 제작을 위해서 6주령 BALB/c 마우스(18-21g)를 실험에 이용하였다. 배양한 생쥐 대장암 세포 CT26은 1x105 cell를 수확(harvest)한 후, 50㎕의 PBS에서 재부유한 후 마우스의 오른쪽 대퇴부(thigh) 피하에 주입하였다. For the production of the CT26 cell transplant animal model, 6-week-old BALB/c mice (18-21 g) were used in the experiment. Cultured mouse colon cancer cells CT26 were harvested 1x10 5 cells, resuspended in 50 µl PBS, and injected under the right thigh of the mouse.
실시예 4. CT26 세포 이식 동물모델에서의 항종양 효과 분석Example 4. Analysis of antitumor effect in CT26 cell transplant animal model
종양이 약 80~100mm3의 부피로 형성된 CT26 세포 이식 마우스의 꼬리에 락토바실러스 사케아이 WIKIM30을 정맥 내 주사하였다.The tail of a CT26 cell transplanted mouse with tumors formed in a volume of about 80-100 mm 3 was intravenously injected with Lactobacillus Sakeai WIKIM30.
투여용량 및 투여주기에 따른 항종양 효과를 분석하기 위해 상기 CT26 세포 이식 마우스에 락토바실러스 사케아이 WIKIM30이 함유된 조성물을 PBS를 이용하여 1Х109 CFU/0.1㎖와 5Х108 CFU/0.1㎖로 준비하여 각각 종양 이식 마우스에 1회 또는 3회 정맥주사하고, 음성대조군에는 PBS를 투여하였다. 시간 경과에 따라 CT26 세포 이식 마우스의 대장암의 종양 크기의 변화를 육안으로 확인한 결과를 도 1에 나타내었다. 또한, 시간 경과에 따라 CT26 세포 이식 마우스에서 종양 크기의 부피 변화를 측정한 결과를 도 2에 나타내었다. To analyze the anti-tumor effect according to the dosage and administration cycle, a composition containing Lactobacillus Sakeai WIKIM30 in the CT26 cell transplanted mouse was prepared using PBS to prepare 1Х10 9 CFU/0.1ml and 5Х10 8 CFU/0.1ml. The tumor-implanted mice were injected intravenously once or three times, and PBS was administered to the negative control group. The results of visually confirming the change in tumor size of colorectal cancer in CT26 cell transplanted mice over time are shown in FIG. 1. In addition, the results of measuring the volume change of tumor size in CT26 cell transplanted mice over time are shown in FIG. 2.
도 1에 나타난 바와 같이, 음성대조군(PBS)에 비해 락토바실러스 사케아이 WIKIM30을 정맥 주입한 경우 13일이 경과한 후에 종양 세포가 육안으로 확인가능할 정도로 줄어든 것을 관찰하였다. 또한, 도 2에 나타난 바와 같이, CT26 세포 이식 마우스의 대장암 종양 크기가 락토바실러스 사케아이 WIKIM30을 1Х109 CFU/0.1㎖로 1회 또는 3회 주사하였을 경우 13일 후에 음성대조군 대비 약 4.6배, 약 3.5배의 항종양 효과가 있음을 확인할 수 있었고, 5Х108 CFU/0.1㎖로 1회 또는 3회 주사하였을 경우 13일 후에 음성대조군 대비 약 3.5배, 약 2.8배의 항종양 효과가 관찰되었다. 이에 따라 투여의 용량 및 주기는 1Х109 CFU/0.1㎖로 1회(single) 투여한 것이 항종양 효과가 가장 우수함을 확인할 수 있었다.As shown in FIG. 1, when intravenous injection of Lactobacillus Sakeai WIKIM30 compared to the negative control (PBS), it was observed that the tumor cells had shrunk to the naked eye after 13 days. In addition, as shown in FIG. 2, when CT26 cell transplantation mice were injected once or three times with Lactobacillus Sakeai WIKIM30 in 1Х10 9 CFU/0.1ml of colon cancer tumor size, about 4.6 times compared to the negative control after 13 days. It was confirmed that there was an anti-tumor effect of about 3.5 times, and when injected 1 or 3 times with 5Х10 8 CFU/0.1ml, an anti-tumor effect of about 3.5-fold and about 2.8-fold was observed after 13 days. Accordingly, it was confirmed that the dose and cycle of the administration were 1× 10 9 CFU/0.1ml once (single) and the anti-tumor effect was the best.
실시예 5. 마우스 피부 흑색종 B16F10 세포의 종양 크기 변화Example 5. Tumor size change of mouse skin melanoma B16F10 cells
5-1. 세포배양5-1. Cell culture
B16F10 마우스 흑색종 세포(한국세포주은행, 한국)는 구입하여 사용하였다. B16F10 마우스 흑색종 세포는 10% 우태아혈청과 1% 페니실린-스트렙토마이신(penicillin-streptomycin)을 포함한 DMEM 배지(Hyclone, 미국)를 사용하여 5% CO2, 37℃ 조건에서 배양하였다. B16F10 mouse melanoma cells (Korea Cell Line Bank, Korea) were purchased and used. B16F10 mouse melanoma cells were cultured at 5% CO 2 and 37° C. using DMEM medium (Hyclone, USA) containing 10% fetal calf serum and 1% penicillin-streptomycin.
5-2. 세포 이식 암 동물모델의 제작5-2. Production of cell transplant cancer animal models
B16F10 세포 이식 동물모델 제작을 위해서 6주령 C57BL/6 마우스 (18-21g)를 실험에 이용하였다. 배양한 마우스 흑색종 세포 B16F10은 각각 1x105 cell를 수확(harvest)한 후, 50㎕의 PBS에서 재부유한 후 마우스의 오른쪽 대퇴부(thigh) 피하에 주입하였다. For the production of the B16F10 cell transplant animal model, 6-week-old C57BL/6 mice (18-21 g) were used in the experiment. The cultured mouse melanoma cells B16F10 were harvested from each 1x10 5 cell, resuspended in 50 µl PBS, and injected under the right thigh of the mouse.
5-3. B16F10 세포 이식 동물모델에서의 항종양 효과 분석5-3. Analysis of anti-tumor effect in B16F10 cell transplant animal model
종양이 약 80~100mm3의 부피로 형성된 B16F10 세포 이식 마우스의 꼬리에 락토바실러스 사케아이 WIKIM30을 정맥 내 주사하였다. 락토바실러스 사케아이 WIKIM30은 PBS를 이용하여 1×1010 CFU/㎖ 균수로 정량하여 준비하였으며, 0.1㎖(1×109 CFU)를 실험동물에 정맥주사하고, 음성대조군에는 PBS를 투여하였다. 시간 경과에 따라 B16F10 세포 이식 마우스의 대장암 종양 크기의 변화를 육안으로 확인한 결과를 도 3에 나타내었다.The tail of a mouse transplanted with B16F10 cells formed with a tumor volume of about 80-100 mm 3 was intravenously injected with Lactobacillus Sakeai WIKIM30. Lactobacillus Sakeai WIKIM30 was prepared by quantifying 1×10 10 CFU/mL bacteria using PBS, 0.1 mL (1×10 9 CFU) was injected intravenously into the experimental animals, and PBS was administered to the negative control group. Fig. 3 shows the results of visually confirming the change in colorectal cancer tumor size of B16F10 cell transplanted mice over time.
도 3에 나타난 바와 같이, 음성대조군(PBS)에 비해 락토바실러스 사케아이 WIKIM30을 정맥 주입한 경우 14일이 경과한 후에 종양 세포가 육안으로 확인가능할 정도로 줄어든 것을 관찰하였다. As shown in FIG. 3, when intravenous injection of Lactobacillus Sakeai WIKIM30 compared to the negative control (PBS), it was observed that the tumor cells were reduced to a level that was visible to the naked eye after 14 days.
또한, 시간 경과에 따라 B16F10 세포 이식 마우스에서 종양 크기의 부피 변화를 측정한 결과를 도 4에 나타내었다. In addition, the results of measuring the volume change of the tumor size in a B16F10 cell transplanted mouse over time are shown in FIG. 4.
도 4에 나타난 바와 같이, B16F10 세포 이식 마우스의 대장암 종양 크기가 락토바실러스 사케아이 WIKIM30을 주사하였을 경우 14일 후 음성대조군 대비 약 3.5배의 항종양 효과가 있음을 확인하였다.As shown in Figure 4, when the colorectal cancer tumor size of the B16F10 cell transplanted mouse was injected with Lactobacillus Sakeai WIKIM30, it was confirmed that the antitumor effect was about 3.5 times that of the negative control after 14 days.
실시예 6. In vitro 항암 활성 효능 분석Example 6. In vitro anticancer activity efficacy analysis
6-1. 세포주의 준비6-1. Preparation of cell lines
락토바실러스 사케아이 WIKIM30의 항암 활성 효과를 관찰하기 위해 Cell counting Kit-8을 이용하여 세포 성장률(cell viability assay)을 측정하였다. 항암 활성 효능실험에 사용된 세포는 마우스 유래 대장암 CT26 세포주, 사람 유래 대장암 SW620 및 HCT116 세포주, 사람 유래 비소세포폐암 H1650 세포주, 사람 유래 췌장암 ASPC1 및 PANC1 세포주, 사람 유래 간암 HepG2 세포주, 사람 유래 방광암 T24 세포주, 그리고 정상 피부 세포주인 CCD-986-sk 세포주를 이용하였다.To observe the anticancer activity effect of Lactobacillus Sakeai WIKIM30, cell viability assay was measured using Cell counting Kit-8. Cells used for the anticancer activity efficacy test include mouse-derived colon cancer CT26 cell line, human-derived colon cancer SW620 and HCT116 cell lines, human-derived non-small cell lung cancer H1650 cell line, human-derived pancreatic cancer ASPC1 and PANC1 cell lines, human-derived liver cancer HepG2 cell line, and human-derived bladder cancer The T24 cell line and the normal skin cell line CCD-986-sk cell line were used.
마우스 유래 대장암 CT26 세포주는 10% FBS(fetal bovine serum)과 1% 페니실린/스트렙토마이신(penicillin/streptomycin)을 첨가한 DMEM 배지를 사용하여 5% CO2, 37℃ 조건에서 계대배양하여 성장기의 세포를 사용하여 실험하였다. 배양된 B16F10 세포는 0.25% trypsin-EDTA로 배양기에 37℃에서 3분간 처리하여 세포를 탈착한 다음 DPBS로 2회 세척 후 1 X 104 cells/well로 준비하였다.The mouse-derived colorectal cancer CT26 cell line was passaged at 5% CO 2 and 37° C. using DMEM medium containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin. It was tested using. The cultured B16F10 cells were treated with 0.25% trypsin-EDTA in the incubator at 37°C for 3 minutes to detach the cells, washed twice with DPBS, and prepared at 1 X 10 4 cells/well.
사람 유래 암세포 SW620, HCT116, H1650, ASPC1, PANC1, HepG2 및 T24 세포주는 10% FBS(fetal bovine serum)과 1% 페니실린/스트렙토마이신(penicillin/streptomycin)을 첨가한 RPMI 배지를 사용하여 5% CO2, 37℃ 조건에서 계대배양하여 성장기의 세포를 사용하여 실험하였다. 배양된 SW620, HCT116, H1650, ASPC1, PANC1, HepG2 및 T24 세포는 0.25% trypsin-EDTA로 배양기에 37℃에서 3분간 처리하여 세포를 탈착한 다음 DPBS로 2회 세척 후 1 X 104 cells/well로 준비하였다.Human-derived cancer cells SW620, HCT116, H1650, ASPC1, PANC1, HepG2 and T24 cell lines are 5% CO 2 using RPMI medium containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin. , it was tested using the growth phase cells subcultured at 37 ℃ condition. Cultured SW620, HCT116, H1650, ASPC1, PANC1, HepG2 and T24 cells were treated with 0.25% trypsin-EDTA for 3 minutes at 37°C in the incubator, detached and washed twice with DPBS, then 1 X 10 4 cells/well Prepared.
사람 유래 정상 피부 세포 CCD-986-sk 세포주는 10% FBS(fetal bovine serum)과 1% 페니실린/스트렙토마이신(penicillin/streptomycin)을 첨가한 RPMI 배지를 사용하여 5% CO2, 37℃ 조건에서 계대배양하여 성장기의 세포를 사용하여 실험하였다. 배양된 CCD-986-나 세포는 0.25% trypsin-EDTA로 배양기에 37℃에서 3분간 처리하여 세포를 탈착한 다음 DPBS로 2회 세척 후 1 X 104 cells/well로 준비하였다.Human-derived normal skin cell CCD-986-sk cell line was passaged at 5% CO 2 at 37°C using RPMI medium containing 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin. Cultured and tested using cells in the growth phase. The cultured CCD-986-na cells were treated with 0.25% trypsin-EDTA in the incubator at 37°C for 3 minutes to detach the cells, washed twice with DPBS, and prepared at 1 X 10 4 cells/well.
6-2. 세포 성장률(cell viability assay) 측정 결과6-2. Cell viability assay measurement results
본 발명은 대장암(3종), 폐암(1종), 췌장암(2종), 간암(1종) 및 방광암(1종) 세포를 배양한 후 락토바실러스 사케아이 WIKIM30을 처리한 세포의 세포 성장률(cell viability assay)을 측정하였으며 락토바실러스 사케아이 WIKIM30을 처리하지 않는 암세포들의 흡광도를 100%의 성장률 기준으로 항암활성 효능을 확인하였다. Cell growth rate of cells treated with Lactobacillus Sakeai WIKIM30 after culturing cells of colorectal cancer (3 types), lung cancer (1 type), pancreatic cancer (2 types), liver cancer (1 type) and bladder cancer (1 type) (cell viability assay) was measured and anticancer activity efficacy was confirmed based on the growth rate of 100% based on the absorbance of cancer cells not treated with Lactobacillus Sakeai WIKIM30.
세포 성장률(cell viability assay)는 MOI1 농도로 처리한 다음 72시간 동안 배양하였고, 배양 후 Cell counting Kit-8 시약을 처리한 후 흡광도(OD 450nm)를 측정하였다. 락토바실러스 사케아이 WIKIM30을 처리하지 않은 세포(comtrol)의 흡광도를 100% 기준으로, 락토바실러스 사케아이 WIKIM30을 처리한 세포의 성장률을 계산하였으며, 실험 결과를 하기에 기재하였다.The cell growth rate (cell viability assay) was treated with MOI1 concentration, and then cultured for 72 hours. After incubation, the cell counting Kit-8 reagent was treated and absorbance (OD 450 nm) was measured. Based on the absorbance of cells not treated with Lactobacillus Sakeai WIKIM30 (comtrol), the growth rate of cells treated with Lactobacillus Sakeai WIKIM30 was calculated, and the experimental results are described below.
마우스 유래 대장암 CT26 세포주, 사람 유래 대장암 SW620 및 HCT116 세포주에 락토바실러스 사케아이 WIKIM30을 처리하여 세포 성장률을 측정한 결과를 도 5에 나타내었다.The results of measuring the cell growth rate by treating Lactobacillus Sakeai WIKIM30 on mouse-derived colorectal cancer CT26 cell line, human-derived colorectal cancer SW620 and HCT116 cell lines are shown in FIG. 5.
도 5에 나타난 바와 같이, 락토바실러스 사케아이 WIKIM30을 처리한 마우스 유래 CT26 세포주에서 61.0%, 사람 유래 대장암 SW620 세포주에서 83.3%, 사람 유래 대장암 HCT116 세포주에서 67.3%의 세포 성장률이 측정되었다. 상기 결과로부터 락토바실러스 사케아이 WIKIM30을 처리한 대장암 세포주의 경우 세포 성장률이 현저하게 저해되므로 대장암에 대해 우수한 항암 활성 효능을 가짐을 확인할 수 있다.As shown in FIG. 5, cell growth rates of 61.0% in the mouse-derived CT26 cell line treated with Lactobacillus Sakeai WIKIM30, 83.3% in the human-derived colorectal cancer SW620 cell line, and 67.3% in the human-derived colorectal cancer HCT116 cell line were measured. From the above results, it can be seen that in the case of the colorectal cancer cell line treated with Lactobacillus Sakeai WIKIM30, cell growth rate is significantly inhibited, so it has excellent anticancer activity efficacy against colorectal cancer.
또한, 사람 유래 비소세포폐암 H1650 세포주에 락토바실러스 사케아이 WIKIM30을 처리하여 세포 성장률을 측정한 결과를 도 6에 나타내었다.In addition, the result of measuring the cell growth rate by treating the human-derived non-small cell lung cancer H1650 cell line with Lactobacillus Sakeai WIKIM30 is shown in FIG. 6.
도 6에 나타난 바와 같이, 락토바실러스 사케아이 WIKIM30을 처리한 사람 유래 비소세포폐암 H1650 세포주에서 59.1%의 세포 성장률이 측정되었다. 상기 결과로부터 락토바실러스 사케아이 WIKIM30을 처리한 폐암 세포주의 경우 세포 성장률이 현저하게 저해되므로 폐암에 대해 우수한 항암 활성 효능을 가짐을 확인할 수 있다.As shown in FIG. 6, a cell growth rate of 59.1% was measured in a human-derived non-small cell lung cancer H1650 cell line treated with Lactobacillus Sakeai WIKIM30. From the above results, it can be confirmed that in the case of the lung cancer cell line treated with Lactobacillus Sakeai WIKIM30, since the cell growth rate is significantly inhibited, it has excellent anticancer activity efficacy against lung cancer.
또한, 사람 유래 췌장암 ASPC1 및 PANC1 세포주에 락토바실러스 사케아이 WIKIM30를 처리하여 세포 성장률을 측정한 결과를 도 7에 나타내었다.In addition, the results of measuring the cell growth rate by treating the human-derived pancreatic cancer ASPC1 and PANC1 cell lines with Lactobacillus Sakeai WIKIM30 are shown in FIG. 7.
도 7에 나타난 바와 같이, 락토바실러스 사케아이 WIKIM30을 처리한 사람 유래 췌장암 ASPC1 세포주에서 35.6%, 사람 유래 췌장암 PANC1 세포주에서 51.6%의 세포 성장률이 측정되었다. 상기 결과로부터 락토바실러스 사케아이 WIKIM30을 처리한 췌장암 세포주의 경우 세포 성장률이 현저하게 저해되므로 췌장암에 대해 우수한 항암 활성 효능을 가짐을 확인할 수 있다.As shown in FIG. 7, cell growth rates of 35.6% in the human-derived pancreatic cancer ASPC1 cell line treated with Lactobacillus Sakeai WIKIM30 and 51.6% in the human-derived pancreatic cancer PANC1 cell line were measured. From the above results, it can be confirmed that in the case of the pancreatic cancer cell line treated with Lactobacillus Sakeai WIKIM30, the cell growth rate is significantly inhibited, and thus it has excellent anticancer activity efficacy against pancreatic cancer.
또한, 사람 유래 간암 HepG2 세포주에 락토바실러스 사케아이 WIKIM30을 처리하여 세포 성장률을 측정한 결과를 도 8에 나타내었다.In addition, the result of measuring the cell growth rate by treating the human-derived liver cancer HepG2 cell line with Lactobacillus Sakeai WIKIM30 is shown in FIG. 8.
도 8에 나타낸 바와 같이, 락토바실러스 사케아이 WIKIM30을 처리한 사람 유래 간암 HepG2 세포주에서는 18.8%의 세포 성장률이 측정되었다. 상기 결과로부터 락토바실러스 사케아이 WIKIM30을 처리한 간암 세포주의 경우 세포 성장률이 현저하게 저해되므로 간암에 대해 우수한 항암 활성 효능을 가짐을 확인할 수 있다.As shown in FIG. 8, a cell growth rate of 18.8% was measured in a human-derived liver cancer HepG2 cell line treated with Lactobacillus Sakeai WIKIM30. From the above results, it can be confirmed that in the case of a liver cancer cell line treated with Lactobacillus Sakeai WIKIM30, cell growth rate is remarkably inhibited and thus has excellent anticancer activity efficacy against liver cancer.
또한, 사람 유래 방광암 T24 세포주에 락토바실러스 사케아이 WIKIM30을 처리하여 세포 성장률을 측정한 결과를 도 9에 나타내었다.In addition, the results of measuring the cell growth rate by treating the human-derived bladder cancer T24 cell line with Lactobacillus Sakeai WIKIM30 are shown in FIG. 9.
도 9에 나타낸 바와 같이, 락토바실러스 사케아이 WIKIM30을 처리한 사람 유래 방광암 T24 세포주에서 53.9%의 세포 성장률이 측정되었다. 상기 결과로부터 락토바실러스 사케아이 WIKIM30을 처리한 방광암 세포주의 경우 세포 성장률이 현저하게 저해되므로 방광암에 대해 우수한 항암 활성 효능을 가짐을 확인할 수 있다.As shown in FIG. 9, a cell growth rate of 53.9% was measured in a bladder cancer T24 cell line derived from humans treated with Lactobacillus Sakeai WIKIM30. From the above results, it can be seen that in the case of the bladder cancer cell line treated with Lactobacillus Sakeai WIKIM30, the cell growth rate is significantly inhibited, and thus it has excellent anticancer activity efficacy against bladder cancer.
상기 실험과 대비하여 사람 유래 정상 피부 CCD-986-sk 세포주에 락토바실러스 사케아이 WIKIM30을 처리하여 세포 성장률을 측정한 결과를 도 10에 나타내었다. In comparison to the above experiment, the result of measuring the cell growth rate by treating Lactobacillus Sakeai WIKIM30 on a human-derived normal skin CCD-986-sk cell line is shown in FIG. 10.
도 10에 나타낸 바와 같이, 락토바실러스 사케아이 WIKIM30을 처리한 정상 피부 세포주에서는 세포 성장율이 97.4%로 측정되었으며, 상기 결과로부터 락토바실러스 사케아이 WIKIM30이 정상 피부 세포주에 대해서는 독성을 갖지 않음을 확인할 수 있다.As shown in FIG. 10, in the normal skin cell line treated with Lactobacillus Sakeai WIKIM30, the cell growth rate was measured to be 97.4%, and it can be confirmed from the above results that Lactobacillus Sakeai WIKIM30 has no toxicity to normal skin cell lines. .
양방검정 t-test는 음성대조군과 락토바실러스 사케아이 WIKIM30 처리군 사이에 종양 성장에서 통계적으로 유의미한 차이를 결정하였다. P < 0.05는 모든 종양성장 그래프 분석에 대하여 유의미하게 고려되었다.The two-way t-test determined statistically significant differences in tumor growth between the negative control group and the Lactobacillus Sakeai WIKIM30 treatment group. P <0.05 was considered significantly for all tumor growth graph analysis.
Figure WO-DOC-FIGURE-ro134
Figure WO-DOC-FIGURE-ro134

Claims (18)

  1. 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 암의 예방 또는 치료용 약학 조성물.Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a pharmaceutical composition for the prevention or treatment of cancer comprising a culture thereof as an active ingredient.
  2. 제1항에 있어서, According to claim 1,
    상기 락토바실러스 사케아이 WIKIM30은 서열번호 1의 핵산서열을 갖는 것을 특징으로 하는 암의 예방 또는 치료용 약학 조성물.The Lactobacillus Sakeai WIKIM30 is a pharmaceutical composition for the prevention or treatment of cancer, characterized by having the nucleic acid sequence of SEQ ID NO: 1.
  3. 제1항에 있어서, According to claim 1,
    상기 암은 대장암, 피부암, 폐암, 췌장암, 간암 및 방광암으로 이루어지는 군으로부터 선택되는 어느 하나의 암인 것을 특징으로 하는 암의 예방 또는 치료용 약학 조성물.The cancer is a cancer cancer, skin cancer, lung cancer, pancreatic cancer, liver cancer and bladder cancer is a pharmaceutical composition for the prevention or treatment of cancer, characterized in that any one selected from the group consisting of cancer.
  4. 제3항에 있어서, According to claim 3,
    상기 폐암은 비소세포폐암인 것을 특징으로 하는 암의 예방 또는 치료용 약학 조성물.The lung cancer is a pharmaceutical composition for the prevention or treatment of cancer, characterized in that the non-small cell lung cancer.
  5. 제3항에 있어서, According to claim 3,
    상기 피부암은 흑색종인 것을 특징으로 하는 암의 예방 또는 치료용 약학 조성물.The skin cancer is a pharmaceutical composition for the prevention or treatment of cancer, characterized in that the melanoma.
  6. 제1항에 있어서,According to claim 1,
    상기 조성물은 경구 투여 또는 비경구 투여의 방법으로 투여 가능한 것을 특징으로 하는 암의 예방 또는 치료용 약학 조성물.The composition is a pharmaceutical composition for the prevention or treatment of cancer, characterized in that it can be administered by a method of oral administration or parenteral administration.
  7. 제6항에 있어서,The method of claim 6,
    상기 비경구 투여의 방법은 정맥주사 투여인 것인, 암의 예방 또는 치료용 약학 조성물.The method of parenteral administration is intravenous administration, a pharmaceutical composition for the prevention or treatment of cancer.
  8. 제1항에 있어서,According to claim 1,
    상기 조성물은 생균 형태인 것을 특징으로 하는 암의 예방 또는 치료용 약학 조성물.The composition is a pharmaceutical composition for the prevention or treatment of cancer, characterized in that in the form of live bacteria.
  9. 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 암의 예방 또는 개선용 식품 조성물.Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a food composition for preventing or improving cancer comprising a culture thereof as an active ingredient.
  10. 제9항에 있어서,The method of claim 9,
    상기 식품은 건강기능식품인, 암의 예방 또는 개선용 식품 조성물.The food is a health functional food, food composition for preventing or improving cancer.
  11. 제9항에 있어서,The method of claim 9,
    상기 식품은 빵, 떡, 캔디, 초콜릿, 껌, 아이스크림, 우유, 치즈, 식육가공품, 어육가공품, 김치, 간장, 된장, 고추장, 춘장, 청국장, 식초, 케첩, 카레, 드레싱, 음료 및 발효유로 이루어지는 군으로부터 선택되는 어느 하나의 식품인 것을 특징으로 하는 암의 예방 또는 개선용 식품 조성물.The food is made of bread, rice cake, candy, chocolate, gum, ice cream, milk, cheese, processed meat products, processed fish products, kimchi, soy sauce, miso, red pepper paste, chunjang, cheonggukjang, vinegar, ketchup, curry, dressing, beverage and fermented milk Food composition for the prevention or improvement of cancer, characterized in that any one food selected from the group.
  12. 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 암의 예방 또는 개선용 식품 첨가제 조성물.Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a food additive composition for preventing or improving cancer comprising a culture thereof as an active ingredient.
  13. 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 가축의 암 예방 또는 개선용 사료 조성물.Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a feed composition for preventing or improving cancer in livestock comprising a culture thereof as an active ingredient.
  14. 제13항에 있어서,The method of claim 13,
    상기 가축은 돼지, 소, 말, 양, 토끼, 염소, 쥐, 햄스터, 기니피그, 개, 고양이, 닭, 칠면조, 오리, 거위, 꿩, 메추라기, 잉어, 붕어 및 송어로 이루어지는 군으로부터 선택되는 어느 하나인 것인, 가축의 암 예방 또는 개선용 사료 조성물.The livestock is any one selected from the group consisting of pig, cow, horse, sheep, rabbit, goat, rat, hamster, guinea pig, dog, cat, chicken, turkey, duck, goose, pheasant, quail, carp, crucian and trout. Phosphorus, a feed composition for preventing or improving cancer in livestock.
  15. 락토바실러스 사케아이 WIKIM30(Lactobacillus sakei, WIKIM30, 기탁번호 KCCM11878P) 또는 이의 배양물을 유효성분으로 포함하는 가축의 암 예방 또는 개선용 사료 첨가제 조성물.Lactobacillus sakeai WIKIM30 (Lactobacillus sakei, WIKIM30, accession number KCCM11878P) or a feed additive composition for the prevention or improvement of cancer in livestock comprising a culture thereof as an active ingredient.
  16. 제1항 내지 제8항 중 어느 하나의 조성물을 인간을 제외한 개체에 투여하는 암의 예방 또는 치료 방법.A method for preventing or treating cancer, wherein the composition of any one of claims 1 to 8 is administered to an individual other than a human.
  17. 제16항에 있어서, The method of claim 16,
    상기 투여의 방법 및 투여량은 정맥주사의 방법으로 락토바실러스 사케아이 WIKIM30을 1Х1010 CFU/㎖ 농도로 0.1㎖ 투여하는 것인, 암의 예방 또는 치료 방법.The method of administration and dosage is 0.1 ml of Lactobacillus Sakeai WIKIM30 at a concentration of 1Х10 10 CFU/ml by intravenous injection, a method for preventing or treating cancer.
  18. 제17항에 있어서,The method of claim 17,
    상기 투여의 횟수는 1회 투여하는 것인, 암의 예방 또는 치료 방법.The number of times of administration is to administer once, a method for preventing or treating cancer.
PCT/KR2019/016627 2018-12-10 2019-11-28 Pharmaceutical composition comprising lactobacillus sakei wikim30 as active ingredient for prevention or treatment of cancer WO2020122476A1 (en)

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EP19896691.3A EP3895719A4 (en) 2018-12-10 2019-11-28 Pharmaceutical composition comprising lactobacillus sakei wikim30 as active ingredient for prevention or treatment of cancer
CN201980081668.4A CN113164536A (en) 2018-12-10 2019-11-28 Pharmaceutical composition for preventing or treating cancer comprising lactobacillus sakei WIKIM30 as an active ingredient
AU2019396986A AU2019396986A1 (en) 2018-12-10 2019-11-28 Pharmaceutical composition comprising Lactobacillus sakei WIKIM30 as active ingredient for prevention or treatment of cancer
SG11202106163RA SG11202106163RA (en) 2018-12-10 2019-11-28 Pharmaceutical composition for prevention or treatment of cancer comprising lactobacillus sakei wikim30 as active ingredient
CA3122124A CA3122124A1 (en) 2018-12-10 2019-11-28 Pharmaceutical composition for prevention or treatment of cancer comprising lactobacillus sakei wikim30 as active ingredient
US17/312,698 US20220062355A1 (en) 2018-12-10 2019-11-28 Pharmaceutical composition comprising lactobacillus sakei wikim30 as active ingredient for prevention or treatment of cancer
JP2021532434A JP2022534150A (en) 2018-12-10 2019-11-28 Pharmaceutical composition for prevention or treatment of cancer containing Lactobacillus salmoni WIKIM30 as an active ingredient

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