WO2022265447A1 - Composition comprenant une souche de lacticaseibacillus sp. ou lactobacillus sp. en tant que principe actif pour le diagnostic, la prévention ou le traitement du cancer - Google Patents

Composition comprenant une souche de lacticaseibacillus sp. ou lactobacillus sp. en tant que principe actif pour le diagnostic, la prévention ou le traitement du cancer Download PDF

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WO2022265447A1
WO2022265447A1 PCT/KR2022/008602 KR2022008602W WO2022265447A1 WO 2022265447 A1 WO2022265447 A1 WO 2022265447A1 KR 2022008602 W KR2022008602 W KR 2022008602W WO 2022265447 A1 WO2022265447 A1 WO 2022265447A1
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cancer
lactobacillus
composition
strain
curvatus
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PCT/KR2022/008602
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English (en)
Korean (ko)
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박승환
최승현
이정숙
이주혁
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한국생명공학연구원
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C12N1/205—Bacterial isolates
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K10/00—Animal feeding-stuffs
    • A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
    • A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A61K35/741—Probiotics
    • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
    • A61K35/747—Lactobacilli, e.g. L. acidophilus or L. brevis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/225—Lactobacillus
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • a composition for cancer diagnosis a kit comprising the composition; cancer prevention or treatment method using the pharmaceutical composition; And it relates to a method for diagnosing cancer using the composition for diagnosis and a method for providing information for diagnosing cancer.
  • Viral vectors A variety of agents for treating cancer have been developed, and non-viral vectors using viral vectors and nano-particles as gene carriers to deliver drugs into cancer tissues.
  • Viral vectors bacterial vectors using aerobic, anaerobic, and anaerobic strains are used.
  • Viral vectors showing high drug delivery efficiency are being used not only for anticancer treatment but also for gene therapy for other diseases.
  • the recently reported side effects of viral vectors clearly show the limitations of viral vectors (wide range of host cells, causing side effects on immunity upon repeated administration), and thus the development of safer gene delivery systems is required.
  • the present inventors have made diligent efforts to develop strains that are safe and have excellent anticancer effects without pathogenicity . ), and Lactobacillus brevis ( Lactobacillus brevis ) By confirming that the strain has a cancer targeting effect and cancer growth inhibitory effect, the strain can be used as an active ingredient in a composition for diagnosing, preventing or treating cancer By revealing that completed the present invention.
  • One object of the present invention is to provide a Lactobacillus paracasei strain deposited under accession number KCTC 14228BP.
  • Another object of the present invention is the Lactobacillus paracasei strain deposited with accession number KCTC 14228BP; Or to provide a pharmaceutical composition for the prevention or treatment of cancer, comprising a culture thereof as an active ingredient.
  • Another object of the present invention is the Lactobacillus paracasei strain deposited with accession number KCTC 14228BP; Or it relates to a food composition for preventing or improving cancer, comprising a culture thereof as an active ingredient.
  • Another object of the present invention is the Lactobacillus paracasei strain deposited with accession number KCTC 14228BP; Or it relates to a feed composition for preventing or improving cancer, comprising a culture thereof as an active ingredient.
  • Another object of the present invention is any one or more selected from the group consisting of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, or Lactobacillus brevis; Or to provide a composition for diagnosing cancer comprising a culture thereof as an active ingredient.
  • Another object of the present invention is to provide a kit for diagnosing cancer including the composition for diagnosing cancer.
  • Another object of the present invention is to provide a method for diagnosing cancer or providing information for diagnosing cancer, including administering the composition for diagnosing cancer to a subject.
  • Another object of the present invention is to provide a method for preventing or treating cancer, comprising administering the pharmaceutical composition to a subject.
  • Another object of the present invention is the Lactobacillus paracasei strain deposited with accession number KCTC 14228BP; Or to provide a cancer prevention, treatment, or improvement use of a composition containing a culture thereof.
  • Another object of the present invention is any one or more selected from the group consisting of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, or Lactobacillus brevis; Or to provide a cancer diagnosis use of the culture thereof.
  • the strain of the present invention itself can be used for diagnosis, prevention, improvement, or treatment of cancer without toxicity, and can be used as a carrier for a cancer therapeutic agent according to loading of an anticancer agent.
  • Lactobacillus paracasei Lactobacillus reuteri , Lactobacillus curvatus , and Lactobacillus brevis confirming the cancer targeting effect of strains in a mouse model of colorectal cancer. This is the diagram showing the result.
  • Figure 2 is a diagram showing the results of confirming the cancer-targeting effect of the Lactobacillus paracasei strain in a colorectal cancer animal model.
  • Figure 3 is a diagram showing the results of confirming the cancer-targeting effect of the Lactobacillus paracasei strain in breast cancer animal models.
  • Figure 4 shows the anticancer effects of Lactobacillus paracasei KCTC 14228BP, Lactobacillus reuteri , Lactobacillus curvatus , and Lactobacillus brevis strains in a mouse model of colorectal cancer. It is a diagram showing the result of checking.
  • Lactobacillus paracasei KCTC 14228BP alone and in combination with Lactobacillus reuteri , Lactobacillus curvatus , and Lactobacillus brevis in a mouse model of colorectal cancer. It is a diagram showing the results of quantifying the anticancer effect.
  • Figure 6 is a diagram showing the results of confirming the anticancer effect of Lactobacillus paracasei KCTC 14228BP strain and Salmonella strain ( Salmonella - ⁇ ppGpp) in a breast cancer mouse model.
  • FIG. 7 is a diagram showing the results of quantifying the anticancer effect of Lactobacillus paracasei KCTC 14228BP strain and Salmonella strain ( Salmonella - ⁇ ppGpp) in a breast cancer mouse model.
  • FIG. 8 is a diagram showing the anticancer effect of the Lactobacillus paracasei KCTC 14228BP strain against various carcinoma types.
  • FIG. 9 is a diagram showing that the anticancer effect of the Lactobacillus paracasei KCTC 14228BP strain is excellent compared to other Lactobacillus paracasei.
  • One aspect of the present invention provides a Lactobacillus paracasei strain deposited with accession number KCTC 14228BP.
  • Lactobacillus paracasei is one of microorganisms belonging to the Lactobaciilus casei group, and is one of the microorganisms inhabiting the human intestine. According to the recent reclassification of the genus Lactobacillus by genetic analysis, Lactobacillus paracasei can be used interchangeably with Lactobacillus paracasei belonging to the genus Lacticaseibacillus.
  • the Lactobacillus paracasei may be a strain deposited as KCTC 14228BP, but is not limited thereto.
  • the strain may be a live strain or an attenuated strain (killed strain).
  • "Attenuation” means modifying a strain to reduce its pathogenicity. Attenuation is done for the purpose of reducing toxicity and other side effects when the strain is administered to a subject.
  • Attenuated strains can be prepared through various methods known in the art. For example, attenuation can be achieved by deleting or destroying virulence factors that allow the strain to survive in host cells. Such deletions and disruptions are well known in the art and are performed by methods such as homologous recombination, chemical mutagenesis, irradiation mutagenesis, or transposon mutagenesis.
  • the culture of the present invention refers to a culture solution obtained by culturing a specific microorganism in a culture medium, a concentrated culture solution, a dried product of a culture solution, a culture filtrate, a concentrated culture filtrate, a dried product of a culture filtrate, a diluted solution, a concentrated solution, and collecting cells of the strain It means a lysate, a fermented product, etc., which have been disrupted, and the culture medium means that it contains a specific bacterium, and the culture filtrate is substantially excluding a specific bacterium separated by filtration or the like. It does not mean that bacteria are completely excluded from the filtrate.) It means not included.
  • the fermentation product of a specific strain is not only the fermented material itself, but also the culture medium of the strain in which the strain and the culture coexist, the fermentation product obtained by filtering the strain from the culture medium, and the strain being sterilized from the culture medium and filtered.
  • fermented product an extract obtained by extracting the fermentation product or a culture medium containing the same, a diluent obtained by diluting the fermentation product or its extract, a concentrated solution, a dried product obtained by drying the fermentation product or its extract, and collecting and crushing the cells of the strain Lysates, etc., include all kinds of materials including fermented products generated from the above strains.
  • the Lactobacillus paracasei strain deposited under accession number KCTC 14228BP of the present invention may have cancer targeting.
  • Lactobacillus paracasei deposited under accession number KCTC 14228BP has a cancer target, enabling cancer diagnosis, and having significantly superior anticancer effect compared to other Lactobacillus paracasei strains (Example 3 and 6).
  • Another aspect of the present invention provides a pharmaceutical composition for preventing or treating cancer, including a Lactobacillus paracasei strain or a culture thereof deposited under accession number KCTC 14228BP.
  • the culture and the like are as described above.
  • the pharmaceutical composition is any one or more selected from the group consisting of Lactobacillus reuteri , Lactobacillus curvatus , and Lactobacillus brevis , or a culture thereof It may further include.
  • Lactobacillus reuteri is known as one of the most widely inhabited intestinal microorganisms in mammals.
  • Lactobacillus curvatus is one of non-starter (NSLAB) microorganisms, and has two subspecies of curvatus and melibiosus.
  • Lactobacillus brevis in the present invention can be found in fermented food, human intestines, feces, and vagina, and is the main Lactobacillus found in tibicos cereals that make kefir. one of the species
  • Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis are all Gram-positive microorganisms belonging to the Lactobaciilaceae family, and have the advantage of being non-toxic to humans.
  • Lactobacillus paracasei and Lactobacillus reuteri are microorganisms included in the Ministry of Food and Drug Safety Notice 19, and have excellent stability.
  • the Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis strains of the present invention may all be derived from the human body, specifically, from the microbiome of the human intestine, but are limited thereto. It doesn't work.
  • all of the Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis strains can be isolated from feces, etc., but the method of origin and separation is not limited as long as each strain has its own characteristics. don't
  • Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis strains may be live strains or attenuated strains (dead strains). Attenuation is as described above.
  • Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis strains of the present invention are derived from the human body and are safe, so they can be used in the form of live strains.
  • cancer refers to cells having characteristics of cancer-causing cells such as uncontrolled proliferation, immortality, metastatic potential, rapid growth and proliferation rate, and characteristic morphological characteristics known in the art. speaks of existence.
  • the cancer may be used in the same sense as "tumor".
  • the cancer is not limited as long as it can be targeted by Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and/or Lactobacillus brevis, but specifically colorectal cancer, breast cancer, and prostate cancer.
  • lung cancer non-small cell lung cancer, colon cancer, bone cancer, pancreatic cancer, gallbladder cancer, skin cancer, head or neck cancer, skin or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, stomach cancer, perianal cancer, colon cancer, fallopian tube carcinoma, endometrium Carcinoma, cervical carcinoma, vaginal carcinoma, 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, chronic or acute leukemia, It may be lymphocytic lymphoma, bladder cancer, renal or ureteric cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system (CNS) tumor, primary central nervous system lymphoma, spinal cord tumor, brainstem glioma or pituitary adenoma, etc.
  • CNS central nervous system
  • pancreatic cancer gallbladder cancer, kidney cancer, colon cancer, lung cancer, liver cancer, skin cancer, breast cancer, bladder cancer, and stomach cancer, but is not limited thereto.
  • Cancer can also include pre-malignant cancer as well as malignant cancer.
  • Lactobacillus paracasei Lactobacillus reuteri, Lactobacillus curvatus, or Lactobacillus brevis may be a genetically engineered strain to additionally express a gene having an anticancer effect.
  • the gene is not limited in type as long as it has anticancer activity, but specifically p53, rb1 (retinoblastoma susceptibility gene), wt1 (Wilms' tumors), nf1 (neurofibromatosis type-1), fap (familial adenomatosis polyposis) coli), vhl (von Hippel-Lindau syndrome), Cytolysin A (toxin gene), and antibody mimics.
  • p53 retinoblastoma susceptibility gene
  • wt1 Wide tumors
  • nf1 neurofibromatosis type-1
  • fap familial adenomatosis polyposis
  • vhl von Hippel-Lindau syndrome
  • Cytolysin A toxin gene
  • Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and/or Lactobacillus brevis strains or cultures thereof contain components known to have anticancer effects (eg, anticancer drugs) or metabolites of the strains. It may be further included, and for the purpose of the present invention, the component or metabolite is not limited in kind.
  • the component having the anticancer effect examples include doxorubicin, cyclophosphamide, mecholrethamine, uramustine, melphalan, chlorambucil, Ifosfamide, bendamustine, carmustine, lomustine, streptozocin, busulfan, dacarbazine, temozolomide ), thiotepa, altretamine, duocarmycin, cisplatin, carboplatin, nedaplatin, oxaliplatin, satrapl Satraplatin, triplatin tetranitrate, 5-fluorouracil, 6-mercaptopurine, capecitabine, cladribine , clofarabine, cystarbine, floxuridine, fludarabine, gemcitabine, hydroxyurea, methotrexate, pemetrexed (pemetrexed), pentostatin, thioguanine, camptothecin, topotecan, irinote
  • prevention refers to any one or more selected from the group consisting of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis or a culture thereof as an active ingredient. It means any action that inhibits or delays the onset or progression of cancer by administration of the composition.
  • treatment refers to clinically intervening to alter the natural process of a subject or cell to be treated, which can be performed during the course of a clinical pathological condition or to prevent it. Desired therapeutic effects include preventing occurrence or recurrence of the disease, alleviating symptoms, reducing any direct or indirect pathological consequences of the disease, preventing metastasis, reducing the rate of disease progression, and alleviating the disease state. or providing temporary relief, remission or improving prognosis.
  • all actions that improve the course of cancer by administering a composition containing Lactobacillus paracasei or its culture as an active ingredient, deposited under accession number KCTC 14228BP, are included.
  • Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis of the present invention have excellent cancer targeting, the efficiency of anticancer drug delivery is high, and the cancer prevention or treatment effect is excellent.
  • Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis of the present invention may be an active ingredient of a pharmaceutical composition, and a component having an anticancer effect by targeting cancer (eg, an anticancer agent) or It can be used as a carrier for the metabolite of the strain and used as a pharmaceutical composition.
  • a component having an anticancer effect by targeting cancer eg, an anticancer agent
  • a carrier for the metabolite of the strain can be used as a pharmaceutical composition.
  • composition of the present invention can be formulated in the form of tablets, pills, powders, granules, capsules, suspensions, internal solutions, emulsions, syrups, etc. according to conventional methods for the administration.
  • Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations are prepared by mixing at least one excipient, for example, starch, calcium carbonate, sucrose, lactose, gelatin, etc. It can be.
  • excipient for example, starch, calcium carbonate, sucrose, lactose, gelatin, etc. It can be.
  • Liquid preparations for oral administration include suspensions, internal solutions, emulsions, syrups, etc., and various excipients, such as wetting agents, in addition to water and liquid paraffin, which are commonly used simple diluents, may be used.
  • Preparations for parenteral administration may include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories, and the like.
  • Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents.
  • composition of the present invention may further include a carrier, excipient or diluent.
  • Carriers, excipients or diluents include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, hydrogel Mineral oils such as hydroxypropyl methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and silicon dioxide may be used.
  • Another aspect of the present invention provides a method for preventing or treating cancer, comprising administering the pharmaceutical composition for preventing or treating cancer to a subject.
  • the pharmaceutical composition for preventing or treating cancer, prevention, treatment, and cancer is as described above.
  • Another aspect of the present invention provides a food composition for preventing or improving cancer, comprising Lactobacillus paracasei or a culture thereof deposited under accession number KCTC 14228BP as an active ingredient.
  • the food composition of the present invention may further include any one or more selected from the group consisting of Lactobacillus reuteri, Lactobacillus, Curvatus, and Lactobacillus brevis, or a culture thereof.
  • Lactobacillus paracasei Lactobacillus reuteri, Lactobacillus curvatus, Lactobacillus brevis, cancer, prevention, etc. are as described above.
  • the term "improvement” refers to any one or more strains selected from the group consisting of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis, or a culture thereof, as an active ingredient. It refers to any activity in which cancer is improved or beneficially changed by administration of the composition.
  • the food composition for preventing or improving cancer of the present invention includes pills, powders, granules, precipitates, tablets, capsules or liquids, and the food to which the composition of the present invention can be added includes, for example, various foods. , For example, there are beverages, gum, tea, vitamin complexes, and health supplements.
  • supplementary food additives may be further added, and the supplementary food additives include conventional food supplement additives in the art, such as flavoring agents, flavoring agents, coloring agents, fillers, stabilizers, and the like. .
  • natural carbohydrates examples 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 dextrins, cyclodextrins, and the like, and sugar alcohols such as xylitol, sorbitol, and erythritol.
  • natural flavors for example, rebaudioside A, glycyrrhizin, etc.
  • synthetic flavors sacharin, aspartame, etc.
  • the food composition for preventing or improving cancer of the present invention contains various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants and enhancers (cheese, chocolate, etc.), pectic acid and salts thereof, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, and the like.
  • it may include fruit flesh for the manufacture of natural fruit juice, fruit juice beverages, and vegetable beverages. These components may be used independently or in combination.
  • the health supplements include health functional foods and health foods.
  • the health functional food is the same term as food for special health use (FoSHU), and is a medicine processed to efficiently display bioregulatory functions in addition to nutrient supply, and has high medical effect.
  • “function (sex)” means to obtain useful effects for health purposes such as regulating nutrients for the structure and function of the human body or physiological functions.
  • the food of the present invention can be prepared by a method commonly used in the art, and can be prepared by adding raw materials and ingredients commonly added in the art during the preparation.
  • the formulation of the food may also be prepared without limitation as long as the formulation is recognized as a food.
  • the food composition of the present invention can be prepared in various types of formulations, and unlike general drugs, it has the advantage of not having side effects that may occur when taking drugs for a long time using food as a raw material, and has excellent portability, so that the composition of the present invention
  • the food composition can be ingested as an adjuvant to enhance the effect of preventing or improving cancer.
  • Another aspect of the present invention provides a feed composition for preventing or improving cancer comprising a Lactobacillus paracasei strain or a culture thereof deposited under accession number KCTC 14228BP.
  • Lactobacillus paracasei prevention, improvement, and cancer are as described above.
  • the feed composition may include known carriers, stabilizers, or additives acceptable for pharmaceutical, food, or feed use.
  • binders for example, there are binders, emulsifiers, and preservatives added to prevent quality deterioration, and amino acids, vitamins, enzymes, flavors, non-protein nitrogen compounds, silicates, and buffers added to feed to increase efficacy.
  • extractants, oligosaccharides, etc. may further include a feed mixture and the like, but is not limited thereto.
  • the feed composition may contain various nutrients such as vitamins, amino acids, and minerals, antioxidants, and other additives as needed, and may be in a suitable state such as powder, granule, pellet, or suspension.
  • the feed composition of the present invention can be supplied alone or mixed with feed for unit animals.
  • the feed of the present invention is not particularly limited, and is not particularly limited as long as it is for animals such as dogs, cats, horses, and cattle. Powder feed, solid feed, dry feed, wet feed, moist pellet feed, dry pellet feed, EP (Extruder Pellet ) Feed, raw food, etc., any feed is fine.
  • Another aspect of the present invention is selected from the group consisting of Lactobacillus paracasei , Lactobacillus reuteri , Lactobacillus curvatus , and Lactobacillus brevis It provides a composition for diagnosing cancer, comprising any one or more or a culture thereof as an active ingredient.
  • Lactobacillus paracasei Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis are as described above.
  • Lactobacillus paracasei may be deposited under accession number KCTC 14228BP, but is not limited thereto.
  • the Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curbatus, and Lactobacillus brevis of the present invention can target cancer, the Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curbatus, or Lactobacillus brevis strains It may include a means capable of detecting, and for example, the strain may be a genetically engineered strain to express a luminescent protein.
  • the type of the luminescent protein is not limited as long as it emits light of a wavelength that can be detected by a device capable of detecting light.
  • the luminescent protein may be green fluorescent protein, red fluorescent protein, blue fluorescent protein, yellow fluorescent protein, near-infrared fluorescent protein, or luciferase derived from prokaryotic or eukaryotic organisms.
  • the luciferase may be bacterial-derived luciferase, firefly ( Photinus pyralis ) luciferase, sea pansy ( Renilla ) luciferase, or Metridia luciferase.
  • the strain is a strain-specific gene known in the art in addition to the expression of a luminescent protein. It may include mutations, markers, and the like.
  • diagnosis means a process of confirming the presence or characteristics of a pathological state.
  • diagnosis may be interpreted as confirming whether cancer progresses or develops.
  • the diagnostic composition of the present invention is administered to a subject to determine whether or not cancer has occurred, and the level of the strain contained in the composition is measured at a specific location (tissue) of the subject to determine whether or not cancer has occurred at the location (tissue).
  • tissue a specific location of the subject to determine whether or not cancer has occurred at the location (tissue).
  • the term “individual” of the present invention refers to all animals, such as rats, mice, livestock, and the like, including humans who have or are likely to develop cancer. Specifically, it may be mammals including humans.
  • the "administration” means introducing the composition of the present invention to a subject by any suitable method, and the route of administration of the composition may be administered through various routes, oral or parenteral.
  • an injection method such as external skin or intraperitoneal injection, rectal injection, subcutaneous injection, intravenous injection, intramuscular injection, or intrathoracic injection may be selected.
  • composition of the present invention can be formulated in the form of tablets, pills, powders, granules, capsules, suspensions, internal solutions, emulsions, syrups, etc. according to conventional methods for the administration.
  • Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations are prepared by mixing at least one excipient, for example, starch, calcium carbonate, sucrose, lactose, gelatin, etc. It can be.
  • excipient for example, starch, calcium carbonate, sucrose, lactose, gelatin, etc. It can be.
  • Liquid preparations for oral administration include suspensions, internal solutions, emulsions, syrups, etc., and various excipients, such as wetting agents, in addition to water and liquid paraffin, which are commonly used simple diluents, may be used.
  • Preparations for parenteral administration may include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories, and the like.
  • Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents.
  • composition of the present invention may further include a carrier, excipient or diluent.
  • Carriers, excipients or diluents include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, hydrogel Mineral oils such as hydroxypropyl methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and silicon dioxide may be used.
  • kits for diagnosing cancer including the composition for diagnosing cancer.
  • composition for diagnosing cancer and cancer are as described above.
  • the kit can measure the presence level of the strain in an individual or sample and the strain in normal tissue with the kit. After measuring and comparing the presence level, if the object or sample has a higher level of strain than normal tissue, it can be diagnosed as having cancer.
  • the kit treats a subject suspected of having cancer or a composition containing one or more strains selected from Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis, or a culture thereof, to a sample of the subject. And, it can be used to diagnose cancer by measuring the level of presence of one or more of the strains present in the subject or a sample of the subject, as long as it is suitable for a luminometer, primer or probe as well as an analysis method for measuring the level of existence
  • One or more other component compositions, solutions or devices may also be included.
  • a diagnostic kit for measuring the presence level of the strain of the present invention may be a kit including a luminometer.
  • the strain since the strain may be a strain genetically engineered to express a luminescent protein, the measuring instrument is not limited to that type as long as it can measure the luminescence of the expressed protein.
  • a diagnostic kit for specifically measuring the presence level of the strain may be a kit including essential elements required to perform RT-PCR.
  • the RT-PCR kit contains, in addition to each primer pair for the strain-specific gene, a test tube or other suitable container, reaction buffer (with varying pH and magnesium concentration), deoxynucleotides (dNTPs), Taq-polymerase and reverse transcriptase. enzymes such as enzymes, DNase, RNAse inhibitors, DEPC-water, sterile water, and the like.
  • a primer pair specific to a gene used as a quantitative control may be included.
  • the diagnostic kit for specifically measuring the levels of the Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis strains may be a kit including a probe.
  • the probe refers to a nucleic acid fragment such as RNA or DNA corresponding to as short as several bases to as long as several hundred bases, capable of specific binding with the strain-specific gene, and labeled with a fluorescent marker or a radioactive marker, etc. (labeled), it is possible to confirm the presence or absence of a specific gene region.
  • the probe may be in the form of an oligonucleotide probe, a single stranded DNA probe, a double stranded DNA probe, or an RNA probe.
  • Another aspect of the present invention is cancer diagnosis comprising the step of detecting any one or more strains selected from the group consisting of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis in a sample.
  • the sample may be a sample isolated from a subject treated with the composition for diagnosing cancer, or a sample isolated from a subject to which the composition for diagnosing cancer is administered.
  • Lactobacillus paracasei Lactobacillus reuteri, Lactobacillus curvatus, Lactobacillus brevis, the composition for diagnosing cancer, and the subject are as described above.
  • the step of detecting one or more strains selected from the group consisting of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis may include measuring the level of the strain in a tissue. However, it is not limited thereto. The detection or level measurement may be performed through biopsy biopsy, measurement of luminescent protein, etc., but is not limited thereto. A step of diagnosing cancer or providing information for diagnosing cancer when a strain of a certain level or higher (eg, the strain detection level of the normal control group) is detected may be further included, but is not limited thereto.
  • the sample may be a cell or tissue of an individual, but is not limited thereto.
  • administering the composition for diagnosing cancer to a subject; and/or measuring the level of detection in the tissue, but is not limited thereto.
  • composition for diagnosing cancer, diagnosing cancer, and the like are as described above.
  • composition for diagnosing cancer of the present invention has an effect of targeting cancer, it is possible to track the occurrence of cancer and the location of a tumor by administering the composition, and by providing such information, information for diagnosing cancer is provided. can do.
  • compositions comprising a Lactobacillus paracasei strain and/or a culture thereof deposited under Accession No. KCTC 14228BP for use in preventing, treating, or improving cancer.
  • the composition may be a pharmaceutical composition, food composition, and / or feed composition, but is not limited thereto.
  • the composition is any one or more selected from the group consisting of Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis; And / or may further include a culture thereof, but is not limited thereto.
  • Another aspect of the present invention is any one or more selected from the group consisting of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis; Or it provides a cancer diagnostic use of its culture.
  • Lactobacillus paracasei may be deposited under accession number KCTC 14228BP, but is not limited thereto.
  • Lactobacillus paracasei Lactobacillus reuteri, Lactobacillus curvatus, Lactobacillus brevis, cancer, prevention, treatment, improvement, diagnosis, and the like are as described above.
  • Example 1 Production of colon cancer and breast cancer animal models by injecting colon cancer or breast cancer cells into normal mice
  • a colorectal cancer animal model was prepared by subcutaneously administering CT26 cell line 1x10 6 cells/50 ⁇ L, a colorectal cancer cell line, to the thigh of a known BALB/c (male) mouse.
  • a breast cancer animal model was prepared by subcutaneously administering 4T1 cell line 1 ⁇ 10 6 cells/50 ⁇ L, a breast cancer cell line, to the thigh of a known BALB/c (male) mouse.
  • Example 2 Preparation of an anticancer composition comprising Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and/or Lactobacillus brevis strains
  • Example 2-1 Obtaining Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis
  • Lactobacillus paracasei In order to obtain Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis, known Lactobacillus reuteri from KGMB, a strain bank derived from Korean intestinal microbiome banking business, Lactobacillus curvatus , and Lactobacillus brevis strains were distributed.
  • the present inventors newly discovered a Lactobacillus paracasei strain, and the first Lactobacillus strain discovered by the present inventors is Lactobacillus paracasei subsp.
  • KGMB04157 or V1bio04157, BCT04157 is the same
  • KCTC Korea Center for Biological Resources
  • Example 2-2 Preparation of anti-cancer composition for administration to cancer animal models
  • a composition containing Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and/or Lactobacillus brevis obtained in Example 2-1 was prepared as follows.
  • the composition contained only microorganisms or PBS, and the composition is classified according to the type of microorganisms included as shown in [Table 2].
  • Example 3 Confirmation of cancer targeting and diagnostic effects of Lactobacillus paracasei (KCTC 14228BP), Lactobacillus reuteri, Lactobacillus curvatus, and/or Lactobacillus brevis strains
  • Example 3-1 Confirmation of cancer targeting and diagnostic effects of Lactobacillus paracasei (KCTC 14228BP), Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis
  • the microorganism has a cancer target, it suggests that cancer can be diagnosed based on this and suggests the possibility of an excellent cancer treatment effect, so the cancer target of the microorganism of the genus Lactobacillus of the present invention was analyzed.
  • composition No. 2 of [Table 2] was administered to the cancer animal model, and 5 days later, the tumor was extracted from the animal model. After each strain, the number of strains per tumor mass was measured. As a result, it was confirmed that all of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis have cancer targets (FIG. 1).
  • Example 3-2 Confirmation of cancer targeting and diagnostic effects of Lactobacillus paracasei (KCTC 14228BP)
  • Lactobacillus paracasei KGMB04157 or V1bio04157, BCT04157, KCTC 14228BP was further specifically confirmed.
  • the Lactobacillus paracasei (1x10 9 CFU) was injected into the colorectal cancer animal model, and 5 days later, the colorectal Tumors were extracted from cancer animal models, smeared on MRS agar, and the resulting colonies were finally confirmed. Colonies were identified by 16S rRNA sequencing.
  • Lactobacillus paracasei (KCTC 14228BP) has excellent colorectal cancer tumor targeting (FIG. 2).
  • Lactobacillus paracasei injection, tumor extraction, smearing, and colony confirmation were performed in the same manner as in the colorectal cancer animal model described above.
  • Lactobacillus paracasei The targeting properties of Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus, and Lactobacillus brevis suggest that they have cancer diagnostic effects.
  • Example 4 Confirmation of anticancer effects of Lactobacillus paracasei (KCTC 14228BP), Lactobacillus reuteri, Lactobacillus curvatus, and/or Lactobacillus brevis strains (in vivo)
  • Example 4-1 Confirmation of the effect of reducing the volume of colorectal cancer tumors by the Lactobacillus strain mixture (in vivo)
  • the mixed composition including Lactobacillus paracasei and Lactobacillus paracasei It was confirmed that significantly reduced the volume of cancer (FIG. 4).
  • Example 4-3 Confirmation of breast cancer tumor volume reduction effect by Lactobacillus paracasei (KCTC 14228BP) (in vivo)
  • Lactobacillus paracasei (KCTC 14228BP) is more cancerous than Salmonella- ⁇ ppGpp (Park SH et al., Theranostics 2016; 6(10):1672-1682.), which is known to have anticancer effects, as well as the PBS group. It was confirmed that the volume was greatly reduced (FIG. 6).
  • Example 4-4 Confirmation of breast cancer tumor volume reduction effect by Lactobacillus paracasei (KCTC 14228BP) (in vivo)
  • Lactobacillus paracasei (KCTC 14228BP) significantly reduced the volume of cancer than not only the PBS group but also Salmonella - ⁇ ppGpp known to have anticancer effects (FIG. 7).
  • Example 5 Confirmation of anticancer effect of Lactobacillus paracasei KCTC 14228BP strain against various cancer types
  • the anticancer effect was confirmed by treating the Lactobacillus paracasei KCTC 14228BP strain of the present invention with various types of carcinoma.
  • each cancer cell was cultured in a 37°C CO 2 incubator using DMEM or RPMI medium supplemented with 10% FBS and 1% Antibiotic solution, and then the number of cancer cells in a 96 well plate was prepared to be 1x10 4 cells/well. After overnight culture, Lactobacillus paracasei KCTC 14228BP was treated at an MOI of 10 with DMEM or RPMI medium supplemented with 2% FBS. After 48 hours, absorbance was measured using LDH assay reagent.
  • Lactobacillus paracasei KCTC 14228BP strain of the present invention has excellent anticancer effects against pancreatic cancer, gallbladder cancer, kidney cancer, colon cancer, lung cancer, liver cancer, skin cancer, breast cancer, and stomach cancer (FIG. 8).
  • Example 6 Comparison of anticancer effects of Lactobacillus paracasei KCTC 14228BP strain and other Lactobacillus paracasei strains
  • Lactobacillus paracasei KCTC 3189 and KCTC 3510 were distributed, and the cancer cell inhibition levels of Lactobacillus paracasei KCTC 14228BP (KGMB04157), KCTC 3189, and KCTC 3510 were measured.
  • a colorectal cancer cell line in a 37°C CO 2 incubator using DMEM or RPMI medium supplemented with 10% FBS and 1% Antibiotic solution
  • the number of cancer cells in a 96 well plate is 1x10 4 cells/well.
  • Lactobacillus paracasei was treated at an MOI of 10 with DMEM or RPMI medium supplemented with 2% FBS. After 48 hours, absorbance was measured using LDH assay reagent.
  • Lactobacillus paracasei KCTC 14228BP (KGMB04157) significantly reduced the survival rate of cancer cells, but Lactobacillus paracasei KCTC 3189 and KCTC 3510 had little effect of reducing cancer cell viability (FIG. 9).
  • Lactobacillus paracasei KCTC 14228BP has a significantly superior anticancer effect compared to other Lactobacillus paracasei.

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Abstract

La présente invention concerne : une souche de Lactobacillus paracasei déposée sous le numéro d'enregistrement KCTC14228BP ; une composition pharmaceutique pour la prévention ou le traitement du cancer et une composition alimentaire pour la prévention ou le soulagement du cancer, comprenant chacune la souche ou une culture de celle-ci en tant que principe actif ; une composition comprenant au moins un lactobacille choisi dans le groupe constitué par Lactobacillus paracasei, Lactobacillus reuteri, Lactobacillus curvatus et Lactobacillus brevis en tant que principe actif pour le diagnostic du cancer ; un kit comprenant la composition ; une méthode de prévention ou de traitement du cancer utilisant la composition pharmaceutique ; un procédé de diagnostic du cancer utilisant la composition de diagnostic ; et un procédé de fourniture d'informations pour le diagnostic du cancer.
PCT/KR2022/008602 2021-06-18 2022-06-17 Composition comprenant une souche de lacticaseibacillus sp. ou lactobacillus sp. en tant que principe actif pour le diagnostic, la prévention ou le traitement du cancer WO2022265447A1 (fr)

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