KR20230047640A - Novel bacteroides uniformis and uses thereof - Google Patents

Abstract

The present invention relates to a pharmaceutical composition comprising at least one selected from a group consisting of the reported Bacteroides uniformis strain and Bacteroides uniformis, a culture, a lysate, and an extract thereof. A Bacteroides unipomis strain and a pharmaceutical composition of the present invention have excellent effects in preventing, alleviating, and treating cancer or inflammatory diseases.

Description

Novel Bacteroides uniformis strains and uses thereof {NOVEL BACTEROIDES UNIFORMIS AND USES THEREOF}

The present invention relates to a reported Bacteroides uniformis strain and its use, and more particularly, to a reported Bacteroides uniformis strain having efficacy in preventing or treating cancer or inflammatory diseases and cancer or inflammatory diseases including the same It relates to a pharmaceutical composition for preventing or treating disease, a food composition, and a cosmetic composition.

Cancer has infinite ability to proliferate, and the rate of proliferation is also rapid, and it forms a tumor microenvironment using blood vessels, lymphatic vessels, and fibroblasts surrounding cancer cells, causing metastasis to other tissues and loss of function of normal cells. It is one of the most terrifying diseases that cause death.

Targeted anticancer drugs targeting genetic mutations occurring in various cancer types have been developed, and the side effects caused by existing chemotherapy drugs have been greatly improved. There is a limit that cannot be expected.

Recently, several immune checkpoint inhibitors that regulate the patient's immunity while maintaining anticancer effects have been developed and used as therapeutic agents. It is not clearly identified, and there are problems such as high cost of treatment.

In order to overcome side effects caused by conventional drugs or immunomodulators, studies using probiotics alone or in combination with anticancer agents have been conducted. As a cancer treatment technology using probiotics, Korean Patent Publication No. 2021-0108900 discloses a pharmaceutical composition for preventing or treating inflammatory diseases containing a Lactobacillus helveticus strain as an active ingredient, and published in Korea Patent No. 2019-0034796 discloses a Lactobacillus plantarum strain having an anti-inflammatory effect.

However, since the above probiotics do not have sufficient improvement effects on cancer or inflammatory diseases, there are limitations in using them as new preventive and therapeutic tools.

The present invention is to solve the above technical problem, an object of the present invention is to provide a new treatment for cancer prevention and treatment using pharmabiotics.

One object of the present invention is to provide a pharmaceutical composition for preventing or treating cancer.

Another object of the present invention is to provide a pharmaceutical composition for preventing or treating inflammatory diseases.

Another object of the present invention is to provide a food composition for preventing or improving cancer or inflammatory diseases.

Another object of the present invention is to provide a cosmetic composition that helps improve inflammatory diseases.

One aspect of the present invention for solving the above problems is,

A pharmaceutical comprising at least one selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract thereof, for use in treating the cancer in a subject in need of treatment of cancer. It is about the enemy composition.

Another aspect of the present invention is selected from the group consisting of Bacteroides uniformis , its culture, its lysates and extracts for use in treating an inflammatory disease in a subject in need of such treatment. It relates to a pharmaceutical composition comprising at least one that is.

In the present invention , the Bacteroides uniformis ( Bacteroides uniformis ) is Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uniformis EB - It may be one or more strains selected from the group consisting of BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB-BUYK8 strain (KCCM13017P).

The anti-cancer or anti-inflammatory pharmaceutical composition of the present invention contains Bacteroides in the range of 1×10 ³ to 1×10 ¹³ cfu, or 1×10 ⁶ to 1×10 ¹¹ cfu, or 1×10 ⁸ to 1×10 ¹⁰ cfu. uniformis may be included.

In the present invention , Bacteroides uniformis can be administered together with other probiotic strains or one or more prebiotics.

Another aspect of the present invention relates to a food composition comprising at least one selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract thereof for preventing or improving cancer or inflammatory disease. will be.

Another aspect of the present invention relates to the reported strains, Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uniformis EB- It relates to a Bacteroides uniformis strain selected from the group consisting of BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB-BUYK8 strain (KCCM13017P).

Since the Bacteroides uniformis species strain of the present invention has been confirmed to have excellent anticancer activity, it can be used as an effective anticancer component.

The anti-cancer pharmaceutical composition of the present invention can be used as a less toxic anti-cancer agent or anti-tumor agent that can minimize side effects due to cytotoxicity of existing anti-cancer agents. As such, it may be effective in improving the quality of life of cancer patients by preventing or reducing the side effects of anticancer drugs. In addition, the pharmaceutical composition of the present invention has excellent efficacy as an additional agent for improving or controlling the effect of other treatments, in addition to its direct anticancer activity.

A composition containing Bacteroides uniformis of the present invention as an active ingredient may be used as a pharmaceutical composition for the treatment or prevention of cancer or inflammatory diseases, as well as food compositions and cosmetic compositions.

Figure 1 is a microscopic observation of the Bacteroides uniformis ( B. uniform is) strain of the present invention.
Figure 2 is a result of electrophoresis after PCR of Bacteroides uniformis strain with BU-specific primers.
Figure 3 is a phylogenetic tree created using the 16s rRNA nucleotide sequence of the Bacteroides uniformis strains of the present invention.
4 is a result of RAPD (Random Amplified Polymorphic DNA) analysis of genomic DNA of Bacteroides uniformis strains.
Figure 5 is a schematic diagram for explaining the animal test procedure for confirming the anti-cancer or anti-inflammatory activity of the Bacteroides uniformis strains of the present invention.
6 is E. coli in BMDC. coli. And it is a graph confirming the concentration of the inflammatory cytokine IL-6 of Bacteroides uniformis strains.
7 is E. coli in BMDC. It is a graph confirming the anti-inflammatory cytokine IL-10 concentration of E. coli and Bacteroides uniformis strains.
8 shows E. coli in spleen cells. It is a graph confirming the anti-inflammatory cytokine IL-12p70 concentration of E. coli and Bacteroides uniformis strains.
Figure 9 is the result of confirming the effect on the proliferation of spleen cells of the Bacteroides uniformis strains of the present invention.
10 shows E. coli in spleen cells. It is a graph confirming the anti-inflammatory cytokine IL-10 concentration of E. coli and Bacteroides uniformis strains.
11 is a graph showing the evaluation of anticancer activity by a Bacteroides uniformis strain using a wound healing test method in HT29 cell line.
12 is a graph showing the evaluation of anticancer activity by a Bacteroides uniformis strain using a wound healing test method in B16-F10 cell line.
13a-b are graphs showing changes in tumor volume according to time and experimental group in an allogeneic anti-cancer animal model to measure the inhibitory effect of the Bacteroides uniformis strain of the present invention on cancer development.

Hereinafter, the present invention will be described in more detail with reference to the accompanying drawings.

The singular forms include plural referents unless the context clearly dictates otherwise. In the present specification, when a part 'includes' a certain component, it means that it may further include other components, not excluding other components, unless otherwise stated.

The term 'cancer' when used in the present invention is meant to include tumors, neoplasias, and malignant tissues or cells. The cancer is colorectal cancer, lung cancer, small cell lung cancer, stomach cancer, liver cancer, blood cancer, bone cancer, pancreatic cancer, skin cancer, head or neck cancer, skin or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, proximal anal cancer, colon cancer, breast cancer, Fallopian tube carcinoma, endometrial carcinoma, cervical cancer, vaginal cancer, vulvar carcinoma, Hodgkin's disease, esophageal cancer, small intestine cancer, endocrine cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute including cancers such as leukemia, lymphocytic lymphoma, bladder cancer, kidney cancer, ureteral cancer, renal cell carcinoma, renal pelvic carcinoma, CNS tumor, primary CNS lymphoma, spinal cord tumor, brainstem glioma, pituitary adenoma, or a combination of one or more of these cancers do.

In this specification, the term 'inflammation' is a response to protect a living body against harmful factors, and immune cells, blood vessels, and inflammatory mediators are involved to inhibit cell damage, remove damaged tissues and necrotic cells, and regenerate tissues. It includes a series of processes. 'Inflammatory disease' is a general term for diseases in which inflammation is the main lesion.

The inflammatory disease is inflammatory bowel disease, systemic lupus erythematosus, scleroderma, atopic dermatitis, psoriasis, anaphylaxis, dermatitis, diabetic retinopathy, retinitis, macular degeneration, uveitis, conjunctivitis, arthritis, rheumatoid arthritis, ankylosing spondylitis, osteoarthritis, osteoporosis , allergy, diabetes, diabetic nephropathy, pyelitis, nephritis, Sjogren's syndrome, autoimmune pancreatitis, periodontal disease, asthma, graft-versus-host disease, chronic pelvic inflammatory disease, endometritis, rhinitis, transplant rejection, and chronic prostatitis. It may be any one selected from.

As used herein, the term 'subject' refers to an animal, including but not limited to a primate (eg, human), cow, pig, sheep, goat, horse, dog, cat, rabbit, rat, or mouse.

As used herein, the terms 'treat', 'treating', and 'treatment' refer to alleviating or eliminating one or more of the disorder, disease or condition, or symptoms associated with the disorder, disease or condition; or alleviating or eradicating the causes of the disorder, disease or condition itself.

As used herein, the term 'prevention' refers to any action that inhibits or delays the occurrence, spread, and recurrence of a target disease by administration of the composition.

The term 'therapeutically effective amount' is meant to contain an amount of the strain sufficient to prevent or alleviate to some extent the development of one or more of the symptoms of the disorder, disease or condition being treated, when administered.

As used herein, the term 'pharmaceutically acceptable' refers to a substance that is physiologically acceptable and does not usually cause severe gastrointestinal disorders, dizziness, allergic reactions or similar reactions when administered to humans.

The term 'about' means an acceptable error for a particular value, the value of which depends on the method of measurement. In certain embodiments, the term 'about' is within 1, 2, 3 or 4 standard deviations.

The term 'culture medium' used throughout the present specification means obtained by culturing the strain of the present application in a known liquid medium or solid medium, and may be used interchangeably with 'culture medium'.

One aspect of the present invention relates to a new generation of pharmabiotics strains, which are strains of the species Bacteroides uniformis . Bacteroides uniformis species strains of the present invention are Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uniformis EB-BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB-BUYK8 strain (KCCM13017P).

Pharmabiotics are defined as human-derived bacteria or their products that have a proven pharmacological role for health or disease (“Probiotics and pharmabiotics,” Bioeng Bugs. 2010 Mar-Apr; 1 (2): 79-84.). The pharmaceutical composition of the present invention contains pharmabiotics as a main component and can be safely used without side effects.

Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uniformis EB-BUYK7 strain (KCCM13016P) and Bacteroides uniformis of the present invention Miss EB-BUYK8 strain (KCCM13017P) may be a strain having 16S rRNA having nucleotide sequences of SEQ ID NO: 1 to SEQ ID NO: 4, respectively.

The Bacteroides uniformis strains of the present invention may include a 16S rRNA gene composed of nucleotide sequences having 97% or more sequence identity with the nucleotide sequences of SEQ ID NOs: 1 to 4. Specifically, it has at least 107%, 98%, 99%, 99.5%, 99.8%, 99.9% or 100% homology with the nucleotide sequence consisting of SEQ ID NOs: 1 to 4 of the present specification.

The anticancer activity of the Bacteroides uniformis strain provided by the present invention may be an activity that inhibits the development or progression of cancer. Specifically, it may delay the occurrence of tumors or inhibit the growth rate of tumors, and may additionally have an activity to prevent metastasis of cancer due to inhibition of tumor growth rate.

The pharmaceutical composition of the present invention may be administered to a subject diagnosed with cancer or confirmed to be at risk of developing cancer. Treatment of cancer with the compositions of the present invention may reduce tumor size or slow tumor growth. The composition of the present invention is intended for use in patients with solid tumors. The composition of the present invention can be used to prevent metastasis of gastrointestinal cancer by delaying the development or growth of tumors in the treatment of gastrointestinal cancer. In certain embodiments, the composition of the present invention is used to treat one or more gastrointestinal cancers selected from the group consisting of esophageal cancer, gallbladder cancer, liver cancer, biliary tract cancer, pancreatic cancer, stomach cancer, small intestine cancer, colorectal cancer, colon cancer, anal cancer, or rectal cancer. can be used In addition, the pharmaceutical composition of the present invention can also be used to treat skin cancer such as basal cell carcinoma, squamous cell carcinoma, melanoma, Kaposi's sarcoma, Paget's disease, and mycosis fungoides.

In one embodiment of the present invention, the Bacteroides uniformis strain of the present invention may promote the production, expression, activity, etc. of anti-inflammatory factors, and specifically, the anti-inflammatory factors are cytokines, IL- 10 or IL-12, but is not limited thereto. IL-10 and IL-12 are involved in the downregulation of the inflammatory response by reducing or alleviating the inflammatory response.

IL-6 has long been considered a pro-inflammatory cytokine induced by LPS along with TNF-a and IL-1. IL-6 has little effect on the synthesis of anti-inflammatory cytokines such as IL-10 and transforming growth factor-b (TGF-b), while pro-inflammatory cytokines such as GM-CSF, IFN-g and MIP-2. Inhibits the production of caine. On the other hand, IL-10 is the most important anti-inflammatory cytokine found within the human immune response and is a potent inactivator of monocyte/macrophage inflammatory cytokine synthesis. IL-10 inhibits monocyte/macrophage derived TNF-a, IL-1, IL-6, IL-8, IL-12, granulocyte colony stimulating factor, MIP-1a and MIP-2a.

Another aspect of the present invention is selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof and an extract thereof for use in treating cancer in a subject in need thereof. It relates to a pharmaceutical composition comprising one or more.

Another aspect of the present invention is a group consisting of Bacteroides uniformis , its culture, its lysate and extract for use in treating the inflammatory disease in a subject in need thereof. It relates to a pharmaceutical composition comprising one or more selected from.

The Bacteroides uniformis species strains include Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uniformis EB- It may be one or more strains selected from the group consisting of BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB-BUYK8 strain (KCCM13017P). Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uniformis EB-BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB- BUYK8 strain (KCCM13017P) may be a strain having 16S rRNA having nucleotide sequences of SEQ ID NO: 1 to SEQ ID NO: 4, respectively.

The Bacteroides uniformis strain of the present invention may include a 16S rRNA gene composed of nucleotide sequences having 97% or more sequence identity with the nucleotide sequences of SEQ ID NOs: 1 to 4. Specifically, it has at least 107%, 98%, 99%, 99.5%, 99.8%, 99.9% or 100% homology with the nucleotide sequence consisting of SEQ ID NOs: 1 to 4 of the present specification.

The strain of the Bacteroides uniformis species may be in a cell-free form including or not including the strain. In the present specification, unless otherwise specified, the strain of Bacteroides uniformis species having anticancer activity is from the group consisting of the cell body of the strain, the culture of the strain, the lysate of the strain, and the extract of the strain It is used to mean one or more selected species. The lysate refers to a lysate obtained by disrupting the strain of the Bacteroides uniformis species or a supernatant obtained by centrifuging the lysate.

formulation of the composition

The pharmaceutical composition of the present invention includes a strain of Bacteroides uniformis, and the strain may be formulated in a freeze-dried form. For example, the composition of the present invention may include granules or gelatin capsules, eg, hard gelatin capsules containing the strain of the present invention. Alternatively, pharmaceutical compositions of the present invention may include live, active bacterial cultures.

The pharmaceutical composition of the present invention can be encapsulated so that the strain can be delivered to the intestine. Encapsulation protects the composition from degradation until delivery to the target location through rupture by chemical or physical stimuli, such as, for example, physical degradation that can be caused by pressure, enzyme activity, or pH change.

Typically, a probiotic, such as a pharmaceutical composition of the present invention, may optionally be combined with at least one suitable prebiotic compound.

Prebiotic compounds are generally indigestible carbohydrates such as oligosaccharides or polysaccharides, or sugar-alcohols that are not broken down or absorbed in the upper digestive tract. Known prebiotics include commercial products such as inulin and transgalacto-oligosaccharides.

The pharmaceutical composition of the present invention may further include pharmaceutically acceptable carriers and/or excipients in addition to the above active ingredients, and in addition, various additives commonly used pharmaceutically, such as binders, disintegrants, coating agents, lubricants, and the like. It can be formulated and prepared together with.

A pharmaceutically acceptable carrier may further include, for example, a carrier for oral administration or a carrier for parenteral administration. Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid and the like. In addition, various drug delivery materials used for oral administration may be included. In addition, carriers for parenteral administration may include water, suitable oil, saline, aqueous glucose and glycol, and the like. Stabilizers and preservatives may further be included. Suitable stabilizers include antioxidants such as sodium bisulfite, sodium sulfite or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben and chlorobutanol. The pharmaceutical composition of the present invention may further include a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifier, a suspending agent, and the like in addition to the above components. Other pharmaceutically acceptable carriers and formulations may be referred to those described in the following literature (Remington's Pharmaceutical Sciences, ^19th ed., Mack Publishing Company, Easton, PA, 1995).

Excipients usable in the present invention include sugars such as sucrose, lactose, mannitol, glucose and the like, and starches such as corn starch, potato starch, rice starch, and partially pregelantinated starch. Binders include polysaccharides such as dextrin, sodium alginate, carrageenan, guar gum, acacia, agar, tragacanth, gelatin, naturally-occurring macromolecular substances such as gluten, hydroxypropylcellulose, methylcellulose, hydroxypropylmethyl Cellulose derivatives such as cellulose, ethyl cellulose, hydroxypropylethyl cellulose, and carboxymethyl cellulose sodium, and polyvinylpyrrolidone, polyvinyl alcohol, polyvinyl acetate, polyethylene glycol, polyacrylic acid, polymethacrylic acid, and vinyl acetate resins. contains polymers;

Decomposers usable in the present invention include cellulose derivatives such as carboxymethyl cellulose, calcium carboxymethyl cellulose, and low-substituted hydroxypropyl cellulose, and sodium carboxymethyl starch, hydroxypropyl starch, corn starch, potato starch, rice starch, and partially starch. Starches such as gelatinized starch may be used.

Examples of the lubricant usable in the present invention include talc, stearic acid, calcium stearate, magnesium stearate, colloidal silica, hydrosilicone dioxide, various types of waxes and hydrogenated oils.

As the coating agent, dimethylaminoethyl methacrylate-methacrylic acid copolymer, polyvinyl acetal diethylaminoacetate, ethyl acrylate-methacrylic acid copolymer, ethyl acrylate-methyl methacrylate-chlorotrimethylammonium ethyl methacrylate Copolymers, water insoluble polymers such as ethyl cellulose, methacrylic acid-ethyl acrylate copolymers, enteric polymers such as hydroxypropylmethylcellulose phthalate, hydroxypropylmethylcellulose acetate succinate, and methylcellulose, hydroxypropylmethylcellulose , polyvinylpyrrolidone, and water-soluble polymers such as polyethylene glycol, but are not necessarily limited thereto.

administration of the composition

The pharmaceutical composition of the present invention is administered orally, but may be administered through the rectal, intranasal, intratumoral, or buccal or sublingual routes.

Compositions of the present invention may be administered once, or may be administered sequentially as part of a therapy regimen. In certain embodiments, the pharmaceutical composition of the present invention can be administered daily.

A composition of the present invention comprises a therapeutically effective amount of a Bacteroides eumiformis strain of the present invention. A therapeutically effective amount of the strain may be sufficient to deliver to the patient's intestine and/or cause partial or total colonization.

The dosage of the strains, which are active ingredients in the pharmaceutical composition for preventing or treating cancer or inflammatory diseases of the present invention, depends on the type of various diseases, the age, weight, sex of the patient, the medical condition of the patient, the severity of the condition, and the sensitivity to the drug. , time of administration, route of administration and rate of excretion, duration of treatment, factors including drugs used concurrently, and other factors well known in the medical field. Therefore, the dosage regimen can vary widely, but it is important to administer the amount that can obtain the maximum effect with the minimum amount without side effects, taking into account all the above factors, which can be easily determined by those skilled in the art using standard methods. .

A suitable daily dose of this strain for an adult human is, for example, about 1×10 ³ to about 1×10 ¹³ CFU for an adult human; eg about 1x10 ⁶ to about 1x10 ¹² CFU; in another embodiment from about 1x10 ⁶ to about 1x10 ¹⁰ CFU; In another embodiment, from about 1x10 ⁸ to about 1x10 ¹⁰ CFU.

combination therapy

Compositions of the present invention comprising a strain of Bacteroides uniformis may be particularly effective when used in combination with an additional therapeutic agent, such as a direct anti-cancer agent.

The pharmaceutical composition of the present invention may optionally further include a chemical anti-cancer agent or immuno-anticancer agent in addition to the Bacteroides uniformis strain. The combined administration of the Bacteroides unformis strain together with a chemo-anticancer agent or immuno-anticancer agent is not only the administration of the Bacteroides uniformis strain and the chemo-anticancer agent or immuno-anticancer agent together in one composition, but each of them separately. It includes administering the composition included in to a patient in need thereof. At this time , Bacteroides uniformis strain; and chemo-anticancer agents or immuno-anticancer agents may be administered sequentially or simultaneously to a patient in need thereof.

Non-limiting examples of the chemotherapy are oxaliplatin, pemetrexed, cisplatin, gemcitabine, carboplatin, fluorouracil (5-FU), cyclophosph Cyclophosphamide, Paclitaxel, Vincristine, Etoposide, Doxorubicin. In addition, the immuno-anticancer agent may be anti-PD1, anti-PDL1, anti-CTLA, anti-Tim3, or anti-LAG3 having an immune checkpoint inhibitory function, but is not necessarily limited thereto.

Another aspect of the present invention is a food containing at least one selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract thereof for use in preventing or improving cancer or inflammatory disease. It's about the composition. Food composition containing the strain of the present invention can be used for health functional foods, dairy products, fermented products, food additives or animal feed.

Bacteroides uniformis species strain-containing food of the present invention can be consumed as a food or nutritional product such as milk or milk product or as a food supplement or health functional food. Examples of the product include, but are not necessarily limited to, dairy products, beverages, juices, soups, or foods such as children's foods.

Another aspect of the present invention relates to a cosmetic composition comprising at least one selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract thereof. The cosmetic composition of the present invention may exhibit an anti-inflammatory effect or an effect of improving skin troubles, and may provide antioxidant and whitening effects.

The cosmetic composition of the present invention contains the bacterium-derived extracellular vesicles as an active ingredient, and may include components commonly used in cosmetic compositions. For example, it may include, but is not limited to, antioxidants, stabilizers, solubilizers, common adjuvants such as vitamins, pigments and flavors, and carriers.

The cosmetic composition of the present invention can be prepared in any formulation commonly prepared in the art, for example, solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing , It may be formulated as an oil, powder foundation, emulsion foundation, wax foundation and spray, but is not limited thereto. More specifically, it may be prepared in formulations of nourishing cream, astringent lotion, softening lotion, lotion, essence, nutritional gel, and massage cream.

Hereinafter, the present invention will be described in more detail by the following examples. However, the following examples are only for illustrating the present invention, and the scope of the present invention is not limited to these examples.

Example 1: Isolation and identification of Bacteroides uniformis strains

1.1. microscope observation

In order to isolate Bacteroides uniformis sp. strains from feces of healthy Koreans, the strains were anaerobic using the anaerobic M2GSC medium described by Barcenilla (Appl. Environ Microbiol., 2000, 66:1654-1661). isolated and cultured.

In order to confirm whether the isolated strain is the Bacteroides uniformis strain, the isolated strain was observed under a microscope and the results are shown in FIG. 1. In FIG. 1, A, B, C, and D are photomicrographs of Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 strains magnified at 400 times magnification, respectively.

Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 strains shown in FIG. 1 were enlarged at 400 times magnification, and all A, B, C, and D were short rod-shaped cells. It was confirmed that they exhibited a similar shape.

1.2. PCR analysis

In order to confirm whether the isolated strain is a Bacteroides uniformis strain, the isolated strain was subjected to PCR analysis using the Bacteroides uniformis -specific primers (SEQ ID NO: 5, SEQ ID NO: 6) in Table 1 below. Through this, a target DNA band of 350 bp was identified, and the results are shown in FIG. 2 . In FIG. 2, lane M is a DNA size marker, lanes 1, 2, 3, and 4 are Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 strains, respectively, and lane 5 is a negative control (distilled water) is the result. Referring to Figure 2, it can be seen that the strains of the present invention have similar band results.

Designation sequence number Direction Sequence (5’→3’) Amplicon size Bunf-F SEQ ID NO: 5 Forward TCC GTT TTC CAC TTA TAA GA 350 bp Bunf-R SEQ ID NO: 6 Reverse GGG TTB CCC CAT TCG G

1.3. Random Amplified Polymorphic DNA (RAPD) analysis

In order to confirm the genetic diversity between the strains isolated as described above, RAPD (Random Amplified Polymorphic DNA) analysis, which is a kind of molecular typing, was performed. To this end, DNA was amplified using universal primers (SEQ ID NOs: 7 to 10) in Table 2, targeting genomic DNA extracted from cells, followed by electrophoresis on a 1% agarose gel for 90 minutes, and UV perforator DNA fragmentation patterns were compared using a transilluminator, and the results are shown in FIG. 3 .

Universal primers Sequence (5′→3′) Tm sequence number OPA9 GGG TTA CGC C 35℃ SEQ ID NO: 7 OPA16 AGC CAG CGA A 36℃ SEQ ID NO: 8 OPH9 TGT AGC TGG G 35℃ SEQ ID NO: 9 910-30 GCG GCG GGG 32℃ SEQ ID NO: 10

As a result of DNA fragmentation pattern comparison, as confirmed in FIG. 3 , Bacteroides uniformis EB-BUYK1, Bacteroides uniformis EB- BUYK4, Bacteroides uniformis EB- BUYK7, and Bacteroides of the present invention The uniformis EB- BUYK8 strains showed different band patterns, and it was confirmed that the strains were different from each other, as the isolation sources were different and the 16S rDNA sequences were also different.

1.4. Phylogenetic tree analysis using 16s rRNA sequencing

As a result of the identification of the strains, strains similar to currently known strains exist, but exact matching results were not obtained, so a phylogenetic tree analysis was performed. For full-length 16S rRNA gene sequencing of the isolated Bacteroides uniformis strains, the 16S rRNA gene was amplified using the 27F (SEQ ID NO: 11) and 1492R (SEQ ID NO: 12) primers in Table 3, and then 3730xl DNA The nucleotide sequence was determined using Analyzer (Thermo Fisher Scientific, USA). Using the 16S rRNA gene sequences of the thus obtained EB-BUYK1, EB-BUYK4, EB-BUYK7, EB-BUYK8 strains and Bacteroides uniformis standard strain (ATCC 8492 ^T ), according to the Maximum Likelihood method, FIG. 4 and The same phylogenetic tree was created.

sequence number Designation Direction Sequence (5′→3′) SEQ ID NO: 11 27F Forward AGA GTT TGA TCM TGG CTC AG SEQ ID NO: 12 1492R Reverse GGT TAC CTT GTT ACG ACT T

As shown in Figure 4, as a result of analyzing the evolutionary relationship through a phylogenetic tree through 16s rDNA gene sequencing, the isolates are all genetically related to the Bacteroides uniformis species. It was confirmed that the strains belonged to it, and the four strains of Bacteroides uniformis showed a similarity of 100%, 99.798%, 99.798%, and 99.933%, respectively, confirming that there were some differences in nucleotide sequence between the isolated strains. did

Based on these results, the isolated strains were named Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 strains, respectively, and deposited with the Korea Center for Microorganisms (KCCM), accession number KCCM13014P (EB -BUYK1), KCCM13015P (EB-BUYK4), KCCM13016P (EB-BUYK7), and KCCM13017P (EB-BUYK8).

Example 2: Safety analysis of Bacteroides uniformis strains

2.1. Confirmation of hemolytic activity

To verify the safety of the four strains of Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 of the present invention, it was confirmed whether they had hemolytic activity.

For this, tryptic soy agar (17.0 g/L pancreatic digest of casein, 3.0 g/L pancreatic digest of soybean, 2.5 g/L dextrose, 5.0 g/L sodium chloride, 2.5 g/L potassium phosphate, 15 g/L After culturing the isolates using a blood agar medium prepared by adding 5% w/v defibrinated sheep blood to L agar), pathogenic β-hemolysis (completely transparent area around the colony) was confirmed. As a result, the table below As shown in Fig. 4, it was confirmed that the Bacteroides uniformis strains of the present invention did not cause β-hemolysis related to pathogenicity because there was no transparent area around the colony.

2.2. Check for presence of virulence gene

To verify the safety of the four strains of Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 of the present invention, the presence or absence of a virulence gene of Bacteroides species was confirmed. At this time, as toxic genes, the presence or absence of CfiA (class B zinc metallo-β-Lactamase encoding gene), bft (enterotoxin encoding gene), fpn (C11 protease encoding gene), and bfp (C10 protease encoding gene) genes were confirmed.

As a result of virulence gene PCR analysis using the gDNA of the four isolated strains of Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8, as shown in Table 4 below, Corresponding toxic genes were not detected in all strains. The Bacteroides uniformis strains of the present invention showed no cytotoxicity at concentrations inducing anticancer or anti-inflammatory effects.

Pathogenecity Bateroides uniformis strain number EB-BUYK1 EB-BUYK4 EB-BUYK7 EB-BUYK8 Hemolytic activity α-hemolysis doesn't exist doesn't exist doesn't exist doesn't exist β-hemolysis doesn't exist doesn't exist doesn't exist doesn't exist γ-hemolysis doesn't exist doesn't exist doesn't exist doesn't exist reading gene detection CfiA not detected not detected not detected not detected bft not detected not detected not detected not detected fpn not detected not detected not detected not detected bfp not detected not detected not detected not detected

Example 3: Confirmation of anti-inflammatory or anti-cancer efficacy of Bacteroides uniformis strains

3.1. Evaluation of anti-inflammatory efficacy in mouse bone marrow-derived dendritic cells

In order to evaluate the anti-inflammatory efficacy of the Bacteroides uniformis strains of the present invention, the secretion of cytokines from mouse bone marrow derived dendritic cells (BMDC) was confirmed.

3.1.1. Isolation and cell culture of BMDCs

A hole was made in a 0.5 ml microtube using an 18G needle, and the femur and tibia of a 6-week-old C57BL/6 mouse were separated, placed in a 1.5 ml centrifuge tube, and centrifuged at 10,000 g for 15 seconds. After washing the pellet in the centrifuge tube three times with PBS, the pellet was cultured in RPMI-1640 (10% FBS, 1% P/S, media, 1X mercaptoethanol, 20 μg GM-CSF) medium in a 150 mm culture dish. cultured. The next day, the BMDCs were transferred to a 100 ml Petri dish and cultured, and on the 5th day, 10 ml of the culture medium was transferred to a 15 ml conical tube and centrifuged at 1,000 Хg for 15 minutes. The supernatant was removed, and 10 ml of BMDC medium was added and placed in a petri dish. Days 6 and 7 BMDCs were used in the experiment.

3.1.2. Confirmation of anti-inflammatory efficacy through measurement of IL-6 in BMDC cells

Representative anti-inflammatory cytokines IL-10 (Invitrogen, #88-7105-86) and IL-12 (Invitrogen, #88-7121-86) were used for the analysis of anti-inflammatory efficacy of Bacteroides uniformis strains of the present invention. The secretion of was analyzed by ELISA (Invitrogen, USA).

BMDC cells were treated with LPS (100 μg/ml), E. coli strain, and 4 EB-BUYK1 strains (10

7 cfu/ml, 10% v/v) for 1 hour in an antibiotic-free medium, then replaced with a medium containing penicillin/streptomycin antibiotics, incubated for 24 hours, and centrifuged the medium at 1,000 Хg to obtain the supernatant The secretion of IL-10 was measured by ELISA. Since there is no Bacteroides uniformis strain that can be used as a positive control with known anti-inflammatory or anti-cancer activity, it was compared with E. coli strain administration as a positive control.

As a result of measuring the concentration of IL-6, a pro-inflammatory cytokine, it was significantly higher in the positive control E. coli treated group than in the normal group ( P <0.001, One-way ANOVA). This means that the inflammatory response increased in the E. coli treated group. In all groups treated with the Bacteroides uniformis strain of the present invention, the concentration of IL-6 was significantly decreased compared to the positive control E. coli treated group ( P <0.001, One-way ANOVA). In addition, among the Bacteroides uniformis strain-treated group, the concentration of IL-10 in the Bacteroides uniformis EB-BUYK7 strain-treated group decreased by about 3 times compared to the positive control group, confirming the lowest IL-6 activity. (CON: 231.6±5.018 pg/ml, E.coli: 8269±74.33 pg/ml, EB-BUYK7: 2668±20.89 pg/m/).

3.1.3. Confirmation of anti-inflammatory efficacy of Bacteroides uniformis strains by measuring IL-10 and IL-12 concentrations in bone marrow-derived dendritic cells (BMDC)

IL-10 (Invitrogen, #88-7105-86) and IL-12 (Invitrogen, #88-7121), representative anti-inflammatory cytokines, were used to analyze the anti-inflammatory efficacy of EB-BUYK strains in BMDC cells. -86) was measured.

As a result of measuring the concentrations of anti-inflammatory cytokines, IL-10 and IL-12, all Bacteroides uniformis experimental groups compared to the experimental control group (untreated) or E.coli treated group. It was confirmed that there was a significant increase ( P <0.001, One-way ANOVA).

In particular, among the Bacteroides uniformis experimental group, the highest concentration of IL-10 was measured in the Bacteroides uniformis EB-BUYK4 strain-administered group (2679±72.46 pg/ml), which was higher than that of the E.coli experimental group (573.2± 6.386 pg/ml), which is about 5.7 times higher concentration. The concentrations of IL-10 in the EB-BUYK1, EB-BUYK7, and EB-BUYK8 experimental groups were 2006±19.43 pg/ml, 1887±28.98 pg/ml, and 2212±24.34 pg/ml, respectively.

In addition, as a result of measuring the concentration of IL-12, as shown in FIG. 8, the Bacteroides uniformis EB-BUYK8 experimental group showed the highest value at 589.6±9.327 pg/ml. Looking at the IL-12 concentration, respectively, 141.2 ± 1.031 pg / ml in the control group without treatment, 61.83 ± 1.699 pg / ml in the control group treated with E.coli , Bacteroides uniformis EB-BUYK1, EB-BUYK4, In the EB-BUYK7 administration group, it was measured as 328.2±32.3 pg/ml, 512.9±18.07 pg/ml, and 362.3±10.64 pg/ml, respectively. In all experimental groups treated with the Bacteroides uniformis strain, there was a significant increase compared to the normal control group or the control group treated with E.coli ( P <0.001, one-way ANOVA).

In conclusion, through in vivo experiments, it was confirmed that the Bacteroides uniformis strains of the present invention are useful for the treatment of inflammatory diseases, and in particular, when the Bacteroides uniformis EB-BUYK1 or EB-BUYK7 strains are administered, It was confirmed that the concentration of IL-6, an inflammatory cytokine, was the lowest, and the concentrations of IL-10 and IL-12, an anti-inflammatory cytokine, were significantly high, so that the treatment effect of inflammatory diseases was the most excellent.

3.2. Evaluation of anti-inflammatory efficacy in spleen cells

In order to evaluate the anti-inflammatory efficacy of the Bacteroides uniformis strains of the present invention, the secretion of cytokines from mouse splenocytes was confirmed.

3.2.1. Isolation and cell culture of spleen cells

C57BL/6 mice were sacrificed and spleens were removed. Prepare a cell strainer (mesh) (70 μm) on a Petri dish (60 mm), and put 3 mL of cold PBS (not containing Ca ⁺ , Mg ⁺ ). The removed spleen is raised here and the tissue is ground using a syringe. Collect the cells that passed through the cell strainer, add 7 mL of PBS, and centrifuge (1000 rpm, 4°C, 10 minutes). After discarding the supernatant, add ACK lysis buffer to the cell pellet, pipette carefully, and incubate at room temperature for 1 minute. Add 10 mL of cold PBS again and proceed with washing and centrifugation. The supernatant was discarded, and the pellet was released with RPMI medium containing 10% FBS, followed by cell counting and culture in an appropriate culture dish.

3.2.2. Confirmation of proliferative capacity of spleen cells

The spleen is a major peripheral immune organ responsible for the initial immune response, and is mainly composed of T cells, B cells, and macrophages, and these immune cells induce cellular and humoral immune responses against the invasion of foreign antigens. do. The spleen is a major lymphoid organ that collects antigens from blood and undergoes maturation of T cells and B cells and differentiation of lymphocytes after being stimulated by antigens. The proliferation of spleen cells has a very important meaning in the immune system. Therefore, the immune enhancing effect can be evaluated by measuring the proliferative ability of splenocytes.

Splenocytes cultured in a 96-well plate were treated with 2 μg/ml of Concanavalin A (ConA) and 1×10 ⁷ CFU/mL of four strains of Bacteroides uniformis for 24 hours. Thereafter, MTS solution (Promega, G3580) was added and incubated for 1 hour, and absorbance was measured at a wavelength of 540 nm using an ELISA reader.

As a result of inducing proliferation of spleen cells by activating T cells by treating ConA alone, there was no significant increase compared to the control group (untreated group). However, in the group treated with the Bacteriodes uniformis strain of the present invention together with ConA, the proliferation of spleen cells was significantly increased compared to the ConA group. The proliferation rate of the control group was converted to 100 to confirm the proliferation rate of the experimental group. In the groups treated with EB-BUYK1, EB-BUYK4, and EB-BUYK7, the proliferation of spleen cells increased by 30.47%, 43.07%, and 45.31%, respectively, compared to the ConA-treated group. (ConA: 102.4 ± 1.431%, ConA + EB-BUYK1: 133.6 ± 4.882%, ConA + EB-BUYK4: 146.5 ± 5.099%, ConA + EB-BUYK7: 148.8 ± 7.112%, all P compared to ConA <0.001, one-way ANOVA).

When the proliferation of splenocytes increases, when exposed to an antigen from the outside, it can be expected to exhibit an immune enhancing effect that induces an effective defense against the antigen by increasing the number of immune cells that induce an immune response to the antigen.

3.2.3. Measurement of cytokines in spleen cells

To analyze the anti-inflammatory efficacy of the Bacteroides uniformis strains of the present invention in spleen cells, the secretion of IL-10 (Invitrogen, #88-7105-86), a representative anti-inflammatory cytokine, was measured, and the results are shown in FIG. 10 shown in

As a result of measuring the concentration of an anti-inflammatory cytokine, IL-10, as shown in FIG. 10, when ConA was treated, it slightly increased compared to the control group, but there was no significant difference. In addition, when the Bacteroides uniformis strain was treated with ConA, it was confirmed that the concentration of the anti-inflammatory cytokine IL-10 significantly increased about 4 to 5 times compared to the treatment with ConA alone (ConA: 308 ±19.38 pg/mL, ConA+EB-BUYK1: 1369±34.48 pg/mL, ConA+EB-BUYK4: 1564±66.63 pg/mL, ConA+EB-BUYK7: 1152±29.27 pg/mL, ConA+EB-BUYK8 : 1649±49.77 pg/mL, all P <0.001 versus ConA, one-way ANOVA). In inducing the differentiation of immune cells in splenocytes by ConA, when the Bacteroides uniformis strain was co-treated, the secretion of IL-10 was further promoted, which supports the immune enhancing effect of the strains of the present invention. do.

3.3. Evaluation of anticancer activity of Bacteroides uniformis strains through wound healing assay using human-derived colorectal cancer cell line (HT29) and mouse-derived melanoma cell line (B16-F10)

In order to investigate the anticancer activity of Bacteroides uniformis of the present invention, a wound healing activity test was conducted. Wounds were formed using the HT29 human-derived colon cancer cell line and the B16-F10 mouse melanoma cell line, and then each Bacteroides uniformis strain was treated to confirm the degree of metastasis.

HT29 human colorectal cancer cells and B16-F10 mouse melanoma cells were cultured in McCoy medium containing 10% FBS and 1% gentamycin and DMEM medium containing 10% FBS and 1% P/S in 5% CO ₂ , Incubated at 37 ℃ conditions. HT29 colorectal cancer cells or B16-F10 melanoma cells are dispensed into a 6-well plate for cell culture and cultured to confluent. Thereafter, the 6-well plate is regularly scratched using a pipette tip. Thereafter , the Bacteroides uniformis strains were treated with 10 ⁷ CFU/mL for 24 hours and observed under a microscope. The cell area was calculated using the Image J program, and the results are shown in tables in FIGS. 11 and 11 .

Referring to FIG. 11, as a result of wound healing experiments using HT29 cells, each of the groups treated with Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 strains was about 38% based on the control group. , 84%, 73%, and 19% reductions in metastatic properties were observed.

Referring also to FIG. 12, in the B16-F10 cell line administration group, the group treated with Bacteroides uniformis EB-BUYK4 strain and EB-BUYK7 strain reduced metastasis by about 38%, respectively, compared to the control group that was not treated with anything. has been confirmed

Example 4: Confirmation of anticancer efficacy of Bacteroides uniformis through animal experiments

In order to verify the anticancer treatment effect of the strain and composition of the present invention, a mouse syngeneic tumor model was constructed using a mouse melanoma cell line, B16-F10, and the antitumor effect was confirmed.

4.1. strain sample

Bacteroides uniformis EB-BUYK1 (KCCM 13014P), EB-BUYK4 (KCCM 13015P), EB-BUYK7 (KCCM 13016P), EB-BUYK8 (KCCM 13017P) live cells used in this experiment were 1x10 ⁸ CFU/150 μl PBS (25% glycerol, 0.05% cysteine/PBS).

4.2 . animal testing

Animal experiments were conducted in compliance with the Animal use and Care Protocol of the Institutional Animal Care and Use Committee (IACUC). To induce an anti-cancer animal model, 8-week-old female C57BL/6 mice were purchased, and after a week-long adaptation period, they were bred for 5 weeks. The breeding environment maintained a constant temperature (22 ℃) and relative humidity (40 ~ 60%), and was bred while adjusting the light and shade in a 12-hour cycle.

4.3 . Sample administration and experimental group setup

The antibiotics in Table 5 below were orally administered to the mice after one week of acclimation for one week.

Antibiotic density Ampicillin (Sigma-A0166) 1g/L Vancomycin (Sigma SBR00001) 0.5 g/L Metronidazole (Sigma M1547) 1 g/L Neomycin (Sigma N6386) 1g/L Amphotericin B (Sigma PHR1662) 0.1 g/L

B16-F10 cells, 2×10 ⁴ cells, were injected subcutaneously into the thigh with 100 μl Matrigel (SC, subcutaneous injection). From the 6th day (when cancer cells rise), the Bacteroides uniformis strain of the present invention was orally administered daily at 1×10 ⁸ CFU for 2 weeks. The size of the tumor was measured using a ruler once every two days from the time the tumor was measured, and the size of the tumor was calculated as the square of the long axis X the short axis X0.5 (mm ³ ), and shown in FIG. 13 .

group experimental group administered drug I Control group (untreated group) PBS administration II Bu EB-BUYK1 strain administration group EB-BUYK1 Live Cell, 1x10 ⁸ CFU III Bu EB-BUYK4 strain administration group EB-BUYK4 Live Cell, 1x10 ⁸ CFU IV Bu EB-BUYK7 strain administration group EB-BUYK7 Live Cell, 1x10 ⁸ CFU V Bu EB-BUYK8 strain administration group EB-BUYK8 Live Cell, 1x10 ⁸ CFU

In order to measure the anticancer effect of the Bacteroides uniformis strain of the present invention, changes in tumor size according to administration time and experimental group in an allogeneic anticancer animal model are shown in FIGS. 13a and 13b, respectively.

Referring to FIG. 13a, in the mouse tumor model, in the case of the PBS-administered control group, the tumor grew rapidly, but it was confirmed that the tumor size decreased in the group to which the Bacteroides uniformis strain of the present invention was orally administered. In particular, on the 20th day after administration, as shown in FIG. 13B, in the Bacteroides uniformis EB-BUYK1, EB-BUYK4, EB-BUYK7, and EB-BUYK8 administration groups, compared to the control group, respectively, 26.9%, 23.3%, Tumor size was reduced by 53.3% and 26.3%, respectively. In particular, compared to the control group's tumor size of 575.2±101.4 mm ³ , the tumor size of the EB-BUYK7-administered group was significantly reduced to 268.7±68.37 mm ³ (p<0.05; t-test).

The specific examples described in this specification are only for describing preferred embodiments of the present invention, and should not be construed as limiting the present invention. The present invention can be practiced with various modifications and changes without departing from the spirit and scope of the present invention, and these facts will be apparent to those skilled in the art. The protection scope of the present invention should be defined by the appended claims, and various modifications and changes of the above are intended to be included in the protection scope of the present invention.

Korea Microbial Conservation Center (Overseas) KCCM13014P 20210622 Korea Microbial Conservation Center (Overseas) KCCM13015P 20210622 Korea Microbial Conservation Center (Overseas) KCCM13016P 20210622 Korea Microbial Conservation Center (Overseas) KCCM13017P 20210622

<110> Enterobiome Co., Ltd. <120> Novel Bacteroides uniformis and uses its <130> P21-ETB-01 <160> 12 <170> KoPatentIn 3.0 <210> 1 <211> 1485 <212> DNA 213 <213> <400> 1 agagtttgat cctggctcag gatgaacgct agctacaggc ttaacacatg caagtcgagg 60 ggcagcatga acttagcttg ctaagtttga tggcgaccgg cgcacgggtg agtaacacgt 120 atccaacctg ccgatgactc ggggatagcc tttcgaaaga aagattaata cccgatggca 180 tagttcttcc gcatggtaga actattaaag aatttcggtc atcgatgggg atgcgttcca 240 ttaggttgtt ggcggggtaa cggccaccca agccttcgat ggataggggt tctgagagga 300 aggtccccca cattggaact gagacacggt ccaaactcct acgggaggca gcagtgagga 360 atattggtca atggacgaga gtctgaacca gccaagtagc gtgaaggatg actgccctat 420 gggttgtaaa cttcttttat acgggaataa agtgaggcac gtgtgccttt ttgtatgtac 480 cgtatgaata aggatcggct aactccgtgc cagcagccgc ggtaatacgg aggatccgag 540 cgttatccgg atttattggg tttaaaggga gcgtaggcgg acgcttaagt cagttgtgaa 600 agtttgcggc tcaaccgtaa aattgcagtt gatactgggt gtcttgagta cagtagaggc 660 aggcggaatt cgtggtgtag cggtgaaatg cttagatatc acgaagaact ccgattgcga 720 aggcagcttg ctggactgta actgacgctg atgctcgaaa gtgtgggtat caaacaggat 780 tagataccct ggtagtccac acagtaaacg atgaatactc gctgtttgcg atatacagta 840 agcggccaag cgaaagcgtt aagtattcca cctggggagt acgccggcaa cggtgaaact 900 caaaggaatt gacgggggcc cgcacaagcg gaggaacatg tggtttaatt cgatgatacg 960 cgaggaacct tacccgggct tgaattgcaa ctgaatgatg tggagacatg tcagccgcaa 1020 ggcagttgtg aaggtgctgc atggttgtcg tcagctcgtg ccgtgaggtg tcggcttaag 1080 tgccataacg agcgcaaccc ttatcgatag ttaccatcag gttatgctgg ggactctgtc 1140 gagactgccg tcgtaagatg tgaggaaggt ggggatgacg tcaaatcagc acggccctta 1200 cgtccggggc tacacacgtg ttacaatggg gggtacagaa ggcagctaca cggcgacgtg 1260 atgctaatcc cgaaagcctc tctcagttcg gattggagtc tgcaacccga ctccatgaag 1320 ctggattcgc tagtaatcgc gcatcagcca cggcgcggtg aatacgttcc cgggccttgt 1380 acacaccgcc cgtcaagcca tgaaagccgg gggtacctga agtgcgtaac cgcaaggagc 1440 gccctagggt aaaactggtg attggggcta agtcgtaaca aggta 1485 <210> 2 <211> 1485 <212> DNA 213 <213> <400> 2 agagtttgat cctggctcag gatgaacgct agctacaggc ttaacacatg caagtcgagg 60 ggcagcatga acttagcttg ctaagtttga tggcgaccgg cgcacgggtg agtaacacgt 120 atccaacctg ccgatgactc ggggatagcc tttcgaaaga aagattaata cccgatggca 180 tagttcttcc gcatggtgga attattaaag aatttcggtc atcgatgggg atgcgttcca 240 ttaggttgtt ggcggggtaa cggccaccca agccttcgat ggataggggt tctgagagga 300 aggtccccca cattggaact gagacacggt ccaaactcct acgggaggca gcagtgagga 360 atattggtca atggacgaga gtctgaacca gccaagtagc gtgaaggatg actgccctat 420 gggttgtaaa cttcttttat acgggaataa agtgaggcac gtgtgccttt ttgtatgtac 480 cgtatgaata aggatcggct aactccgtgc cagcagccgc ggtaatacgg aggatccgag 540 cgttatccgg atttattggg tttaaaggga gcgtaggcgg acgcttaagt cagttgtgaa 600 agtttgcggc tcaaccgtaa aattgcagtt gatactgggt gtcttgagta cagtagaggc 660 aggcggaatt cgtggtgtag cggtgaaatg cttagatatc acgaagaact ccgattgcga 720 aggcagcttg ctggactgta actgacgctg atgctcgaaa gtgtgggtat caaacaggat 780 tagataccct ggtagtccac acagtaaacg atgaatactc gctgtttgcg atatacagta 840 agcggccaag cgaaagcgtt aagtattcca cctggggagt acgccggcaa cggtgaaact 900 caaaggaatt gacgggggcc cgcacaagcg gaggaacatg tggtttaatt cgatgatacg 960 cgaggaacct tacccgggct tgaattgcaa ctgaatgatg tggagacatg tcagccgcaa 1020 ggcagttgtg aaggtgctgc atggttgtcg tcagctcgtg ccgtgaggtg tcggcttaag 1080 tgccataacg agcgcaaccc ttatcgatag ttaccatcag gttatgctgg ggactctgtc 1140 gagactgccg tcgtaagatg tgaggaaggt ggggatgacg tcaaatcagc acggccctta 1200 cgtccggggc tacacacgtg ttacaatggg gggtacagaa ggcagctaca cggcgacgtg 1260 atgccaatcc cgaaagcctc tctcagttcg gattggagtc tgcaacccga ctccatgaag 1320 ctggattcgc tagtaatcgc gcatcagcca cggcgcggtg aatacgttcc cgggccttgt 1380 acacaccgcc cgtcaagcca tgaaagccgg gggtacctga agtgcgtaac cgcaaggagc 1440 gccctagggt aaaactggtg attggggcta agtcgtaaca aggta 1485 <210> 3 <211> 1485 <212> DNA 213 <213> <400> 3 agagtttgat cctggctcag gatgaacgct agctacaggc ttaacacatg caagtcgagg 60 ggcagcatga acttagcttg ctaagtttga tggcgaccgg cgcacgggtg agtaacacgt 120 atccaacctg ccgatgactc ggggatagcc tttcgaaaga aagattaata cccgatggca 180 tagttcttcc gcatggtaga attattaaag aatttcggtc atcgatgggg atgcgttcca 240 ttaggttgtt ggcggggtaa cggccaccca agccttcgat ggataggggt tctgagagga 300 aggtccccca cattggaact gagacacggt ccaaactcct acgggaggca gcagtgagga 360 atattggtca atggacgaga gtctgaacca gccaagtagc gtgaaggatg actgccctat 420 gggttgtaaa cttcttttat acgggaataa agtgaggcac gtgtgccttt ttgtatgtac 480 cgtatgaata aggatcggct aactccgtgc cagcagccgc ggtaatacgg aggattcgag 540 cgttatccgg atttattggg tttaaaggga gcgtaggcgg acgcttaagt cagttgtgaa 600 agtttgcggc tcaaccgtaa aattgcagtt gatactgggt gtcttgagta cagtagaggc 660 aggcggaatt cgtggtgtag cggtgaaatg cttagatatc acgaagaact ccgattgcga 720 aggcagcttg ctggactgta actgacgctg atgctcgaaa gtgtgggtat caaacaggat 780 tagataccct ggtagtccac acagtaaacg atgaatactc gctgtttgcg atatacagta 840 agcggccaag cgaaagcgtt aagtattcca cctggggagt acgccggcaa cggtgaaact 900 caaaggaatt gacgggggcc cgcacaagcg gaggaacatg tggtttaatt cgatgatacg 960 cgaggaacct tacccgggct tgaattgcaa ctgaatgatg tggagacatg tcagccgcaa 1020 ggcagttgtg aaggtgctgc atggttgtcg tcagctcgtg ccgtgaggtg tcggcttaag 1080 tgccataacg agcgcaaccc ttatcgatag ttaccatcag gttatgctgg ggactctgtc 1140 gagactgccg tcgtaagatg tgaggaaggt ggggatgacg tcaaatcagc acggccctta 1200 cgtccggggc tacacacgtg ttacaatggg gggtacagaa ggcagctaca cggcgacgtg 1260 atgctaatcc ctaaagcctc tctcagttcg gattggagtc tgcaacccga ctccatgaag 1320 ctggattcgc tagtaatcgc gcatcagcca cggcgcggtg aatacgttcc cgggccttgt 1380 acacaccgcc cgtcaagcca tgaaagccgg gggtacctga agtgcgtaac cgcaaggagc 1440 gccctagggt aaaactggtg attggggcta agtcgtaaca aggta 1485 <210> 4 <211> 1485 <212> DNA 213 <213> <400> 4 agagtttgat cctggctcag gatgaacgct agctacaggc ttaacacatg caagtcgagg 60 ggcagcatga acttagcttg ctaagtttga tggcgaccgg cgcacgggtg agtaacacgt 120 atccaacctg ccgatgactc ggggatagcc tttcgaaaga aagattaata cccgatggca 180 tagttcttcc gcatggtaga actattaaag aatttcggtc gtcgatgggg atgcgttcca 240 ttaggttgtt ggcggggtaa cggccaccca agccttcgat ggataggggt tctgagagga 300 aggtccccca cattggaact gagacacggt ccaaactcct acgggaggca gcagtgagga 360 atattggtca atggacgaga gtctgaacca gccaagtagc gtgaaggatg actgccctat 420 gggttgtaaa cttcttttat acgggaataa agtgaggcac gtgtgccttt ttgtatgtac 480 cgtatgaata aggatcggct aactccgtgc cagcagccgc ggtaatacgg aggatccgag 540 cgttatccgg atttattggg tttaaaggga gcgtaggcgg acgcttaagt cagttgtgaa 600 agtttgcggc tcaaccgtaa aattgcagtt gatactgggt gtcttgagta cagtagaggc 660 aggcggaatt cgtggtgtag cggtgaaatg cttagatatc acgaagaact ccgattgcga 720 aggcagcttg ctggactgta actgacgctg atgctcgaaa gtgtgggtat caaacaggat 780 tagataccct ggtagtccac acagtaaacg atgaatactc gctgtttgcg atatacagta 840 agcggccaag cgaaagcgtt aagtattcca cctggggagt acgccggcaa cggtgaaact 900 caaaggaatt gacgggggcc cgcacaagcg gaggaacatg tggtttaatt cgatgatacg 960 cgaggaacct tacccgggct tgaattgcaa ctgaatgatg tggagacatg tcagccgcaa 1020 ggcagttgtg aaggtgctgc atggttgtcg tcagctcgtg ccgtgaggtg tcggcttaag 1080 tgccataacg agcgcaaccc ttatcgatag ttaccatcag gttatgctgg ggactctgtc 1140 gagactgccg tcgtaagatg tgaggaaggt ggggatgacg tcaaatcagc acggccctta 1200 cgtccggggc tacacacgtg ttacaatggg gggtacagaa ggcagctaca cggcgacgtg 1260 atgctaatcc cgaaagcctc tctcagttcg gattggagtc tgcaacccga ctccatgaag 1320 ctggattcgc tagtaatcgc gcatcagcca cggcgcggtg aatacgttcc cgggccttgt 1380 acacaccgcc cgtcaagcca tgaaagccgg gggtacctga agtgcgtaac cgcaaggagc 1440 gccctagggt aaaactggtg attggggcta agtcgtaaca aggta 1485 <210> 5 <211> 20 <212> DNA <213> artificial sequence <220> <223> primer Bunf-F <400> 5 tccgttttcc acttataaga 20 <210> 6 <211> 16 <212> DNA <213> artificial sequence <220> <223> Primer Bunf-R <400> 6 gggttbcccc attcgg 16 <210> 7 <211> 10 <212> DNA <213> artificial sequence <220> <223> Primer OPA9 <400> 7 ggggtacgcc 10 <210> 8 <211> 10 <212> DNA <213> artificial sequence <220> <223> Primer OPA16 <400> 8 agccagcgaa 10 <210> 9 <211> 10 <212> DNA <213> artificial sequence <220> <223> Primer OPH9 <400> 9 tgtagctggg 10 <210> 10 <211> 9 <212> DNA <213> artificial sequence <220> <223> Primer 910-30 <400> 10 9 <210> 11 <211> 20 <212> DNA <213> artificial sequence <220> <223> primer 27F <400> 11 agagtttgat cmtggctcag 20 <210> 12 <211> 19 <212> DNA <213> artificial sequence <220> <223> primer 1492R <400> 12 ggttaccttg ttacgactt 19

Claims (10)

A pharmaceutical comprising at least one selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract thereof, for use in treating the cancer in a subject in need of treatment of cancer. enemy composition. For use in treating an inflammatory disease in a subject in need of treatment for an inflammatory disease, Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract comprising at least one selected from the group consisting of pharmaceutical composition. The method of claim 1 or 2, wherein the Bacteroides uniformis (Bacteroides uniformis ) is Bacteroides unformis EB-BUYK1 strain (KCCM13014P), Bacteroides unformis EB-BUYK4 strain (KCCM13015P), A pharmaceutical composition, characterized in that at least one strain selected from the group consisting of Bacteroides uniformis EB-BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB-BUYK8 strain (KCCM13017P). The pharmaceutical composition according to claim 1, wherein the pharmaceutical composition contains Bacteroides in the range of 1×10 ³ to 1×10 ¹³ cfu, or 1×10 ⁶ to 1×10 ¹¹ cfu, or 1×10 ⁸ to 1×10 ¹⁰ cfu. A pharmaceutical composition wherein an amount of Uniformis is administered to a subject. The pharmaceutical composition according to claim 1 or 2, wherein the composition further comprises other probiotic strains or one or more prebiotics in addition to the Bacteroides uniformis species strain . A food composition comprising at least one selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract thereof for use in preventing or improving cancer or inflammatory diseases. According to claim 6, wherein the Bacteroides uniformis ( Bacteroides uniformis ) Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uni A food composition, characterized in that at least one strain selected from the group consisting of Formis EB-BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB-BUYK8 strain (KCCM13017P). A cosmetic composition comprising at least one selected from the group consisting of Bacteroides uniformis , a culture thereof, a lysate thereof, and an extract thereof for use in preventing or improving inflammatory diseases. According to claim 8, wherein the Bacteroides uniformis ( Bacteroides uniformis ) Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uni A pharmaceutical composition, characterized in that at least one strain selected from the group consisting of Formis EB-BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB-BUYK8 strain (KCCM13017P). Bacteroides uniformis EB-BUYK1 strain (KCCM13014P), Bacteroides uniformis EB-BUYK4 strain (KCCM13015P), Bacteroides uniformis EB-BUYK7 strain (KCCM13016P) and Bacteroides uniformis EB- Bacteroides uniformis strain selected from the group consisting of BUYK8 strain (KCCM13017P).

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