KR20220130882A - Composition for preventing or treating cancer comprising Eubacterium callanderi, its culture medium or its culture medium extract thereof as an active ingredient - Google Patents

Abstract

The present invention relates to a composition for preventing or treating cancer comprising Eubacterium callanderi, a culture medium thereof, or a culture medium extract as an active component. Eubacterium callanderi isolated from feces of Koreans of the present invention, the culture medium, or the culture medium extract can be usefully used for the prevention or treatment of cancer through an effect of inhibiting cancer cell proliferation in vitro and in vivo.

Description

Eubacterium calanderi, a composition for preventing or treating cancer comprising Eubacterium callanderi, its culture medium or its culture medium extract as an active ingredient {Composition for preventing or treating cancer comprising Eubacterium callanderi, its culture medium or its culture medium extract thereof as an active ingredient}

The present invention relates to a composition for preventing or treating cancer comprising Eubacterium callanderi , a culture solution thereof, or an extract thereof as an active ingredient.

In the gastrointestinal tract of mammals, numerous types of bacteria that make up "microbiota" live symbiotically. Internationally, many studies have shown that the gut microbiome affects obesity, diabetes, cancer, childbirth, intestinal disease, growth, growth, aging, lifespan, Alzheimer's, depression, autism and even the human brain, affecting human behavior and behavior. It has been proven that it also affects sociability. With the development of next-generation sequencing technology, it has become possible to obtain extensive information about the gut microbiome (the entire genetic information of gut microbes and microorganisms).

Most of the gut microbiota live in an anaerobic environment, and it is very difficult to separate and culture them in an in vitro environment. Due to the characteristics of such intestinal microbes, it is very limited to identify the community and characteristics of the gut microbes through a culturing approach. Since the gut microbes identified in previous studies were cultured and reported from the guts of Westerners with completely different genetic traits and dietary habits, a new approach is needed to suit the gut microbiome environment of Koreans. Most of the new drug development using gut microbiota is in the R&D stage or clinical stage. Some university hospitals in Korea are using fecal transplantation to treat colitis and irritable bowel disease against antibiotic-resistant bacteria such as Clostridium difficile.

Eubacterium ( Eubacterium ) is a genus of Gram-positive anaerobic bacilli, and was isolated as a normal flora from feces, rumen and periodontal tissue. The Eubacterium is homogeneous or polymorphic in cell morphology and does not form spores. The optimum growth temperature for Eubacterium is 37° C., and the optimum pH is 7.0. Eubacterium callanderi ( Eubacterium callanderi ) is a kind of Gram-positive bacteria and corresponds to the genus Eubacterium in the family Eubacteriaceae . It is an environmental anaerobic rod-shaped bacterium first isolated in 1998 from an industrial anaerobic digester.

Korean Patent Application Laid-Open No. 2019-0117687 relates to a pharmaceutical composition of a bile acid derivative and a microbiome and uses thereof, and discloses a method for treating FXR-mediated diseases or disorders of the intestinal microbiome including Eubacterium, Korean Patent Application Laid-Open No. 2020-0053531 relates to bacterial extracellular vesicles, and discloses a method for treating autoimmune diseases, inflammatory diseases, metabolic diseases or cancer diseases of bacterial extracellular vesicles including Eubacterium, Korea Patent Publication No. 2009-0075566 relates to an immune-enhancing kimchi composition having anticancer effect, obesity suppression and lipid lowering effect, and a method for manufacturing the same. Intestinal bacteria such as Bifidobacterium, Clostridium, Bacteroides and Eubacterium convert cholesterol into coprostanol to prevent absorption, or inhibit the activity of hydroxymethylglutarate (HMG) reductase or acetyl CoA synthetase to synthesize cholesterol in vivo. is disclosed, and Korean Patent Application Laid-Open No. 2020-0125637 relates to microbiome-related immunotherapy, oncology induced by anticancer therapy, including administration to a subject containing a microbial agent - treatment induction A method of treating or preventing a condition comprising the fecal microbiota of a human donor as a type of microorganism is disclosed.

However, Eubacterium callanderi ( Eubacterium callanderi ), a culture solution thereof or a composition for preventing or treating cancer comprising an extract thereof as an active ingredient has not been disclosed.

1. Korean Patent Publication No. 2019-0117687 2. Korean Patent Publication No. 2020-0053531 3. Korean Patent Publication No. 2009-0075566 4. Korean Patent Publication No. 2020-0125637

The present invention has been derived from the above needs, and the present inventors have found that Eubacterium callanderi isolated from Korean feces, its culture medium, or its culture medium extract has an inhibitory effect on cancer cell proliferation in vitro and in vivo . Found and completed the present invention.

In order to solve the above problems, the present invention is Eubacterium callanderi ( Eubacterium callanderi ), It provides a pharmaceutical composition for preventing or treating cancer comprising a culture solution thereof or an extract thereof as an active ingredient.

In one example of the present invention, the culture medium is anaerobic in a reinforced clostridial medium (RCM) for 2 days and then filtered through a filter to remove cells, and the culture medium extract is an ethyl acetate extract. .

In another embodiment of the present invention, the cancer is a solid cancer, and the cancer is colorectal cancer, colorectal cancer, stomach cancer, breast cancer, brain cancer, melanoma, cervical cancer, head and neck cancer, kidney cancer, lung cancer, ovarian cancer, prostate cancer It is at least one selected from the group.

In addition, the present invention is Eubacterium callanderi ( Eubacterium callanderi ), It provides a health functional food composition for improving cancer comprising its culture solution or its culture solution extract as an active ingredient.

Eubacterium callanderi isolated from Korean feces of the present invention, its culture medium, or its culture medium extract can be usefully used for the prevention or treatment of cancer through the effect of inhibiting the proliferation of cancer cells in vitro and in vivo . have.

1 shows that Eubacterium callanderi culture medium (CFS) isolated from human feces according to an embodiment of the present invention exhibits a concentration-dependent growth inhibitory activity of human colorectal cancer cells (HCT116).
Figure 2 shows that Eubacterium callanderi CFS according to an embodiment of the present invention exhibits cell proliferation inhibitory activity not only in HCT116 but also in mouse colorectal cancer cells (CT26), and that this activity does not appear in human colonic normal cells (CCD 841 CoN). will be.
Figure 3 shows that the colorectal cancer proliferation inhibitory activity of Eubacterium callanderi CFS according to an embodiment of the present invention is shown through apoptosis and cell cycle arrest.
Figure 4 shows that the active material of Eubacterium callanderi CFS according to an embodiment of the present invention is not affected by heat-inactivation and enzyme treatment and is present in the aqueous layer during organic solvent extraction.
5 shows the effect of inhibiting colon cancer growth of Eubacterium callanderi live bacteria and culture medium in BALB/C mice according to an embodiment of the present invention.

Hereinafter, preferred embodiments of the present invention will be described in detail. In addition, many specific details such as specific components are shown in the following description, which are provided to help a more general understanding of the present invention, and it is common in the art that the present invention can be practiced without these specific details. It will be self-evident to those who have the knowledge of And, in describing the present invention, if it is determined that a detailed description of a related known function or configuration may unnecessarily obscure the gist of the present invention, the detailed description thereof will be omitted.

In order to achieve the object of the present invention, the present invention is Eubacterium callanderi ( Eubacterium callanderi ), It provides a pharmaceutical composition for preventing or treating cancer comprising a culture solution thereof or an extract thereof as an active ingredient.

The strain may be derived from the human body, in particular, may be derived from a human feces sample, but is not limited thereto.

The strain may be a live strain or an attenuated strain (dead strain). By "attenuation" is meant modifying a strain to reduce its pathogenicity. Attenuation is for the purpose of reducing toxicity and other side effects when the strain is administered to a subject. The attenuated strain can be prepared through a variety of methods known in the art. For example, attenuation can be achieved by deleting or disrupting a virulence factor that allows the strain to survive in the host cell. Such deletions and disruptions are well known in the art and are carried out by methods such as, for example, homologous recombination, chemical mutagenesis, irradiation mutagenesis or transposon mutagenesis. In particular, since the strain of the present invention is derived from the human body and is safe, it may be used in the form of a live strain, but is not limited thereto.

In one embodiment of the present invention, the culture solution can be prepared to cover all of the culture conditions for a predetermined time or longer in which an effective active ingredient can be released. Preferably, it may be cultured for 24 hours or more, 48 hours or more, or 72 hours or more, and more preferably, cultured for 48 hours or more, but is not limited thereto.

In addition, in order to further prevent cell incorporation, it is used after filtration with a filter after liquid culture.

In the present invention, the culture medium includes all media in the field in which Eubacterium callanderi can be cultured, and preferably Reinforced clostridial medium (RCM) is used, but is not limited thereto.

The culture solution extract may be extracted using water or an organic solvent, and the extracted solution may be used in liquid form or concentrated and/or dried. The organic solvent is not necessarily limited thereto, but methanol, ethanol, isopropanol, butanol, ethylene, acetone, hexane, ether, chloroform, ethyl acetate, butyl acetate, dichloromethane, N, N-dimethylformamide (DMF), dimethyl Sulfoxide (DMSO), 1,3-butylene glycol, propylene glycol, or a mixed solvent thereof, preferably methanol, ethanol, butanol, hexane, dichloromethane, ethyl acetate, or a mixed solvent thereof, more preferably Ethyl acetate (ethyl acetate) extract, but is not limited thereto.

In addition, the extract can be extracted by heating or at room temperature under conditions in which the active ingredient of the extract is not destroyed or minimized. Depending on the organic solvent to be extracted, the degree of extraction and loss of the active ingredient of the extract may be different, so an appropriate organic solvent should be selected and used. The extraction method of the organic solvent is not particularly limited, and examples thereof include cold extraction, ultrasonic extraction, and reflux cooling extraction.

As used herein, the term 'cancer' broadly refers to the uncontrolled abnormal growth of the host's own cells, resulting in infiltration of tissues and surrounding tissues distal to the initial site of abnormal cell growth in the host. The main types include carcinomas, which are cancers of epithelial tissue (e.g., skin, squamous cells), sarcomas, which are cancers of connective tissue (e.g., bone, cartilage, fat, muscle, blood vessels, etc.), and cancers from brain and spinal tissues. cancers of the central nervous system. It refers to cancer, neoplasia or malignancy, encompassing all types of cancer, whether new or recurrent.

In another example of the present invention, the cancer is a solid cancer, preferably colon cancer, colorectal cancer, stomach cancer, breast cancer, brain cancer, melanoma, cervical cancer, head and neck cancer, kidney cancer, lung cancer, ovarian cancer, consisting of prostate cancer At least one selected from the group, but is not limited thereto.

As used herein, the term “prevention” refers to inhibiting or delaying the occurrence of a disease from the cause.

As used herein, the term "treatment" means to stop the progression of damage by inhibiting the progress and/or aggravation of symptoms even without complete cure, or to ameliorate some or all of the symptoms to guide them in the direction of healing.

The pharmaceutical composition of the present invention may further include suitable carriers, excipients and diluents commonly used in the preparation of the composition.

The pharmaceutical composition of the present invention may be used alone or in combination with methods using surgery, drug treatment, and biological response modifiers for the prevention and treatment of diseases.

Cancer therapeutic agents that can be used as additional therapeutic agents include chemotherapeutic agents, such as alkylating agents such as thiotepa and cyclophosphamide; alkyl sulfonates such as busulfan, improsulfan and piposulfan; aziridines such as benzodopa, carboquone, meturedopa and uredopa; ethylenimine and methyllamelamine, including altretamine, triethylenemelamine, triethylenephosphoramide, triethyylenethiophosphoramide and trimethylolomelamine; acetogenins (particularly bullatacin and bullatacinone); camptothecin (including the synthetic analogue topotecan); bryostatin; callistatin; CC-1065 (including its adozelesin, cazelesin and bizelesin synthetic analogs); cryptophycins (particularly cryptophycin 1 and cryptophycin 8); dolastatin; duocarmycin (including synthetic analogs, KW-2189 and CB1-TM1); eleuterobin; pancratistatin; sarcodictine; Spongistatin; nitrogen mustards such as chlorambucil, chlornaphazine, colophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembicin, phenesterine, prednimu Steen, trophosphamide, uracil mustard; nitrosureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine and ranimnustine; antibiotics such as enedyin antibiotics (eg, calicheamicins, particularly calicheamicin gamma 1I and calicheamicin omega 1I; dynemycins, including dynemycin A; bisphosphonates such as clodronate; esperamicin; as well as neocardinostatin chromophore and related chromoprotein enediin antibiotic chromophore, aclasinomycin, actinomycin, otrarnycin, azaserine, bleomycin, cactinomycin, carabicin, caminomycin, carzinophylline , chromomycinis, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin (morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin), epirubicin, esorubicin, idarubicin, marcelomycin, mitomycin such as mitomycin C, mycophenolic acid, nogalamicin, olibomycin, peflo mycin, poppyromycin, puromycin, quelamycin, rhodorubicin, streptonigrin, streptozocin, tubersidin, ubenimex, ginostatin, zorubicin; Lauracil (5-FU): folic acid analogues such as denopterin, methotrexate, pteropterin, trimetrexate; analogs such as ancitabine, azacitidine, 6-azauridine, camofur, cytarabine, dideoxyuridine, doxyfluridine, enocitabine, floxridine; androgens such as calusterone, dromostanolone pro Cypionate, epithiostanol, mephithiostan, testolactone; ; aminolevulinic acid; enyluracil; amsacrine; bestrabucil; bisantrene; edataxate; depopamine; demecholcin; diaziquone; elformitin; elliptinium acetate; epothilone; etoglucide; gallium nitrate ; hydroxyurea; lentinan; ronidinin; maytansinoids such as maytansine and ansamitosine; mitogua zone; mitoxantrone; fur short mole; nitraerin; pentostatin; phennamet; pyrarubicin; losoxantrone; podophyllic acid; 2-ethylhydrazide; procarbazine; PSK polysaccharide complex); Lazoxic acid; lyzoxine; sizofuran; spirogermanium; tenuazonic acid; triaziquone; 2,2',2''-trichlorotriethylamine; trichothecenes (especially T-2 toxins, veraculin A, loridine A and anguidin); urethane; vindesine; Dacarbazine; mannomustine; mitobronitol; mitolactol; Fivobroman; gastocin; arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids such as paclitaxel and docetaxel; chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum coordination complexes such as cisplatin, oxaliplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; vinorelbine; novantron; teniposide; edatrexate; daunomycin; aminopterin; Zeloda; ibandronate; irinotecan (eg, CPT-11); topoisomerase inhibitor RFS 2000; difluoromethylomitin (DMFO); retinoids such as retinoic acid; capecitabine; and pharmaceutically acceptable salts, acids or derivatives of any of the above.

Another form of cancer treatment is cancer immunotherapy. Immunotherapy refers to therapies that use a subject's immune system to treat cancer, such as checkpoint inhibitors, cancer vaccines, cytokines, cell therapy, CAR-T cell and dendritic cell therapy. Non-limiting examples of immunotherapy include nivolumab (BMS, anti-PD-1), pembrolizumab (Merck, anti-PD-1), ipilimumab (BMS, anti-CTLA-4) as checkpoint inhibitors, MEDI4736 (AstraZeneca, anti-PD-L1), and MPDL3280A (Roche, anti-PD-L1). Other immunotherapies include tumor vaccines such as Gardail, Cervarix, BCG, sipulencel-T, Gp100:209-217, AGS-003, DCVax-L, Algenpantucel-L, Tergenpantucel )-L, TG4010, ProstAtak, Prostvac-V/RTRICOM, Rindopepimul, E75 peptide acetate, IMA901, POL-103A, Belagenpumatucel-L, GSK1572932A, MDX-1279, GV1001, and It may be Tecemotide. Immunotherapy may be administered via injection (eg, intravenously, intratumorally, subcutaneously or into lymph nodes), but may also be administered orally, topically or via aerosol. Immunotherapy may include adjuvants such as cytokines.

Another cancer treatment is immune checkpoint inhibitors. Immune checkpoint inhibition broadly refers to inhibiting the checkpoint that cancer cells can create to prevent or downregulate an immune response. Examples of immune checkpoint proteins include, but are not limited to, CTLA4, PD-1, PD-L1, PD-L2, A2AR, B7-H3, B7-H4, BTLA, KIR, LAG3, TIM-3 or VISTA. The immune checkpoint inhibitor may be an antibody or antigen-binding fragment thereof that binds to the immune checkpoint protein and inhibits the immune checkpoint protein. Examples of immune checkpoint inhibitors include nivolumab, pembrolizumab, pidilizumab, AMP-224, AMP-514, STI-A1110, TSR-042, RG-7446, BMS-936559, MEDI-4736, MSB-0020718C, AUR -012 and STI-A1010.

The pharmaceutical composition of the present invention may be formulated in the form of oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, and sterile injection solutions according to conventional methods, and powders , tablets, capsules, injections and liquids are more preferred. Such formulation may be performed by a method conventionally performed in the pharmaceutical field, and may be preferably formulated according to each disease or component using the method disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA.

Carriers, excipients and diluents that may be included in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose , methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.

In the case of formulation, it may be prepared by additionally using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants.

Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and these solid preparations include at least one excipient, for example, starch, calcium carbonate, sucrose or lactose ( lactose), gelatin, etc. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used.

Liquid formulations for oral administration include suspensions, solutions, emulsions, syrups, etc., and various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, in addition to water and liquid paraffin, which are commonly used simple diluents, may be included. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. As the non-aqueous agent and suspending agent, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used. As a base of the suppository, witepsol, macrogol, tween, cacao butter, laurin, glycerogelatin, and the like may be used.

A preferred dosage of the pharmaceutical composition of the present invention is the species, size, body surface area, age, sex, immune capacity and general health of the subject, the specific microorganism to be administered, the duration and route of administration, the type and stage of the disease, for example, a tumor It may depend on many factors, including size, and other compounds such as drugs administered concurrently. In addition to the above factors, these levels may be affected by the infectivity of the microorganism and the nature of the microorganism as determined by one of ordinary skill in the art. In the present invention, an appropriate minimum dosage level of the microorganism may be a level sufficient for the microorganism to survive, grow and replicate. The dosage of the pharmaceutical composition described in the present invention may be appropriately set or adjusted according to the dosage form, administration route, the degree or stage of the target disease, and the like. For example, a typical effective dose of the formulation is 0.01 mg/kg body weight/day to 1000 mg/kg body weight/day, 0.1 mg/kg body weight/day to 1000 mg/kg body weight/day, 0.5 mg/kg body weight/day to 500 mg/kg body weight/day, 1 mg/kg body weight/day to 100 mg/kg body weight/day, or 5 mg/kg body weight/day to 50 mg/kg body weight/day. Effective doses are 0.01, 0.05, 0.1, 0.5, 1, 2, 3, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 200, 500, or 1000 mg/kg body weight/ It may be more than one day, but is not limited thereto.

In addition, effective dosages and treatment protocols are generally initiated at less than optimal dosages. After that, depending on the circumstances, it is preferable to increase it in small increments until the optimum effect is reached. This can be done according to the general capacity verification process in the art.

Individual administrations may include any number of two or more administrations, including 2, 3, 4, 5 or 6 administrations. One of ordinary skill in the art can readily determine the number of doses to be performed or it is desirable to perform one or more additional doses according to methods known in the art for monitoring the methods of treatment and other monitoring methods provided herein. Accordingly, in the present invention, when one or more administrations of the pharmaceutical composition are provided to a subject, the number of administrations may be determined by monitoring the subject, and based on the results of the monitoring, it is determined whether to provide one or more additional administrations. The determination of whether to give one or more additional doses may be based on various monitoring results.

The period between administrations can be any of a variety of periods. The period between administrations can be a function of a variety of factors including the number of administrations, a monitoring step as described with respect to the period during which the subject develops an immune response. In one example, the duration may be a function of time in which the subject elicits an immune response; For example, the period of time may be longer than the period in which the subject develops an immune response, such as greater than about 1 week, greater than about 10 days, greater than about 2 weeks, or greater than about 1 month; In other examples, the period of time may be shorter than the period in which the subject elicits an immune response, such as less than about 1 week, less than about 10 days, less than about 2 weeks, or less than about 1 month.

In some embodiments, delivery of an additional therapeutic agent in combination with a pharmaceutical composition comprising the present invention may be for reducing side effects. Side effects or toxicity are those that include all kinds of symptoms experienced by a subject during and after treatment. Typical side effects or toxicity are abdominal pain, acid excess, acid reflux, allergic reaction, alopecia, anaphylaxis, anemia, anxiety, anorexia, arthralgia, asthenia, ataxia, azotemia, loss of balance, bone pain, bleeding, blood clots, hypotension, hypertension , dyspnea, bronchitis, bruising, low white blood cell count, red blood cell count, low platelet count, cardiotoxicity, cystitis, hemorrhagic cystitis, arrhythmia, heart valve disease, cardiomyopathy, coronary artery disease, cataract, central neurotoxicity, cognitive impairment, Confusion, conjunctivitis, constipation, cough, convulsions, cystitis, deep vein thrombosis, dehydration, depression, diarrhea, dizziness, dry mouth, dry skin, indigestion, dyspnea, edema, electrolyte imbalance, esophagitis, fatigue, loss of fertility, fever , flatulence, flushing, gastric reflux, gastroesophageal reflux disease, genital pain, granulocytopenia, gynecomastia, glaucoma, hair loss, hand-foot syndrome, headache, hearing loss, heart failure, palpitations, heartburn, hematoma, hemorrhagic cystitis, hepatotoxicity, hyperamylase Hyperemia, hypercalcemia, hyperchloremia, hyperglycemia, hyperkalemia, hyperlipaseemia, hypermagnesemia, hypernatremia, hyperphosphatemia, hyperpigmentation, hypertriglyceridemia, hyperuricemia, hypoalbuminemia, hypocalcemia, Hypochloremia, hypoglycemia, hypokalemia, hypomagnesemia, hyponatremia, hypophosphatemia, asthenia, infection, injection site reaction, insomnia, iron deficiency, pruritus, joint pain, nephropathy, leukopenia, hepatic insufficiency, memory loss, menopause, mouth ulcers, mucositis, myalgia, myalgias, myelosuppression, myocarditis, neutrophil fever, nausea, nephrotoxicity, neutropenia, nosebleeds, paralysis, ototoxicity, pain, paresthesia erythematosus, pancytopenia, Pericarditis, peripheral neuritis, pharyngitis, glare, photosensitivity, pneumonia, pneumonia, proteinuria, pulmonary embolism, pulmonary fibrosis, pulmonary toxicity, rash, rapid heartbeat, rectal bleeding, anxiety, rhinitis, seizures, dyspnea, Sinusitis, thrombocytopenia, tinnitus, urinary tract infection, vaginal bleeding, vaginal dryness, dizziness, water retention, weakness, weight loss, weight loss increase and dry mouth. In general, toxicity is acceptable if the benefit to the subject achieved with the therapy outweighs the side effects experienced by the subject with the therapy.

In addition, the present invention is Eubacterium callanderi ( Eubacterium callanderi ), It provides a health functional food composition for improving cancer comprising its culture solution or its culture solution extract as an active ingredient.

As used herein, "improvement" refers to any action that improves or beneficially changes the symptoms of cancer, such as alleviation, reduction, or disappearance of symptoms of cancer by ingestion of the health functional food composition according to the present invention

When the composition of the present invention is included in health functional food, the composition can be added as it is or used together with other health functional food or health functional food ingredients, and there is no particular limitation on the other ingredients, and various herbal medicines like ordinary food Extracts, food supplement additives or natural carbohydrates may be included as additional ingredients, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be suitably determined according to the purpose of use. In general, in the production of food or beverage, the composition of the present invention may be added in an amount of preferably 15 parts by weight or less, more preferably 10 parts by weight or less, based on the raw material. However, in the case of long-term intake for the purpose of health control and hygiene, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range.

There is no particular limitation on the type of health functional food that can contain the composition of the present invention, and specific examples include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, and ice cream. There are dairy products, various soups, beverages, teas, drinks, alcoholic beverages and vitamin complexes, and may include all health functional foods in a conventional sense, and may include foods used as feed for animals.

Examples of the natural carbohydrate include monosaccharides such as glucose, fructose and the like; disaccharides such as maltose, sucrose and the like; and polysaccharides such as conventional sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those described above, natural flavoring agents (eg rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used.

In addition to the above, the composition of the present invention contains various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and thickening agents (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts. salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated beverages, and the like. Other natural fruit juices and fruit juice beverages and fruit juices for the production of vegetable beverages may be included. These components may be used independently or in combination.

In the present invention, health functional (sex) food (functional food) is the same term as food for special health use (FoSHU), and in addition to nutritional supply, medical and medical processed to efficiently exhibit bioregulatory functions. Foods that are highly effective. Here, "function (sex)" means to obtain a useful effect for health purposes such as regulating nutrients or physiological action with respect to the structure and function of the human body. The food of the present invention can be prepared by a method commonly used in the art, and at the time of manufacture, it can be prepared by adding raw materials and components commonly added in the art. In addition, the formulation of the food may be prepared without limitation as long as it is a formulation recognized as a food. The food composition of the present invention can be prepared in various types of dosage forms, and unlike general drugs, it has the advantage of not having side effects that may occur during long-term administration of the drug using food as a raw material, and has excellent portability.

Advantages and features of the present invention, and methods for achieving them, will become apparent with reference to the embodiments described below in detail. However, the present invention is not limited to the embodiments disclosed below, but will be implemented in a variety of different forms, and only these embodiments allow the disclosure of the present invention to be complete, and common knowledge in the technical field to which the present invention belongs It is provided to fully inform the possessor of the scope of the invention, and the present invention is only defined by the scope of the claims.

<Example 1> Eubacterium calenderi ( Eubacterium callanderi ) Isolation of strains, preparation of culture medium, and inhibition of human colorectal cancer cells

Eubacterium callanderi isolated from human feces was cultured in anaerobic conditions in RCM medium for 2 days and then filtered through a filter to prepare a microbial culture medium (CFS; cell free supernatant). Human colorectal cancer cells (HCT116) were seeded in 96-well plates at 1x10 ⁴ cells/well, and the filtered culture solution was treated for each concentration after 24 hours (0 ~ 100 μl/ml). As a result of observing cell proliferation through MTT assay 3 days after treatment with the culture medium, it was confirmed that there was a concentration-dependent cell growth inhibitory activity (FIG. 1).

<Example 2> Eubacterium calenderi ( Eubacterium callanderi ) Colorectal cancer-specific anticancer effect of the culture medium

Similarly, it was confirmed by MTT assay whether the activity of CFS prepared in Example 1 was also active in mouse colorectal cancer cells (CT26) for in vivo mouse animal experiments. And in order to confirm that this activity is a specific activity for cancer cells, human colonic normal cells (CCD 841 CoN) were treated with a culture medium and confirmed through MTT assay, it was confirmed that cell proliferation inhibitory activity hardly appeared in normal cells (Fig. 2).

<Example 3> Eubacterium calander ( Eubacterium callanderi ) of the strain culture Effect on cell cycle and apoptosis

Flow cytometry assay was performed to confirm the mechanism of cell proliferation inhibitory activity in human colorectal cancer cells (HCT116) against Eubacterium callanderi CFS. When HCT116 cells were treated with Eubacterium callanderi culture medium, cultured for 3 days, and flow cytometry assay was performed with AnnexinV and Propidium Iodide (PI), HCT116 cells in CFS-treated group increased by 6.8% in G2 phase cells compared to control, and apoptosis was confirmed to increase by 12%. Therefore, it was confirmed that the culture medium of Eubacterium callanderi affects both cell cycle arrest and apoptosis (FIG. 3).

<Example 4> Eubacterium calenderi ( Eubacterium callanderi ) Analysis of the properties of the active substances in the culture medium of the strain

To characterize the active substance of Eubacterium callanderi CFS, heat-inactivation (A), protease and α-amylase treatment (B), and ethyl acetate extraction (C) were performed. RCM and CFS were heat-treated at 100° C. for 10 minutes, then the activity was checked, and the enzymes were also treated with RCM and CFS at 37° C. for one hour, and then the activity was checked. In the case of ethyl acetate extraction, ethyl acetate was mixed with RCM and CFS in the same volume, and the aqueous phase and organic phase were separated through layer separation. The organic phase was used by evaporating ethyl acetate with a rotary evaporator and then dissolving it in methanol. As a result, it was confirmed that the material was not affected by heat-inactivation and enzyme treatment, and it was confirmed that the active material was present in the aqueous phase when extracted with ethyl acetate (FIG. 4).

<Example 5> Eubacterium calander ( Eubacterium callanderi ) of the strain or culture medium in vivo effect

To confirm in vivo the ability of Eubacterium callanderi culture medium (A) and strain (B) to inhibit colorectal cancer cell proliferation, mouse colorectal cancer cells CT26 were subcutaneously injected into the flank of 6-week-old BALB/C mice, followed by inoculation of cancer cells with PBS and strains It was orally administered before/after, and RCM medium and CFS were injected subcutaneously around the tumor to measure the tumor size for a total of 30 days from 4 days after cancer cell inoculation at 2 day intervals. As a result, it was confirmed that the size of the culture medium was reduced by 25.6% and the weight by 32%, and the size of colorectal cancer was reduced by 30.5% and the weight by 29.5% when the strain was inoculated (FIG. 5).

Claims (10)

Eubacterium callanderi ( Eubacterium callanderi ), A pharmaceutical composition for preventing or treating cancer comprising its culture solution or its culture solution extract as an active ingredient The pharmaceutical composition for preventing or treating cancer according to claim 1, wherein the culture medium is filtered through a filter after liquid culture in anaerobic state. The pharmaceutical composition for preventing or treating cancer according to claim 1, wherein the culture solution extract is an ethyl acetate extract. The pharmaceutical composition for preventing or treating cancer according to claim 1, wherein the cancer is a solid cancer. According to claim 1, wherein the cancer is colorectal cancer, colorectal cancer, stomach cancer, breast cancer, brain cancer, melanoma, cervical cancer, head and neck cancer, kidney cancer, lung cancer, ovarian cancer, characterized in that at least one selected from the group consisting of prostate cancer A pharmaceutical composition for preventing or treating cancer Eubacterium callanderi ( Eubacterium callanderi ), Health functional food composition for improving cancer comprising its culture solution or its culture solution extract as an active ingredient [Claim 7] The health functional food composition for improving cancer according to claim 6, wherein the culture medium is filtered through a filter after liquid culture in an anaerobic state. The health functional food composition for improving cancer according to claim 6, wherein the culture solution extract is an ethyl acetate extract. The health functional food composition for improving cancer according to claim 6, wherein the cancer is a solid cancer. According to claim 6, wherein the cancer is colorectal cancer, colorectal cancer, stomach cancer, breast cancer, brain cancer, melanoma, cervical cancer, head and neck cancer, kidney cancer, lung cancer, ovarian cancer, characterized in that at least one selected from the group consisting of prostate cancer Health functional food composition for improving cancer

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