Composition for improving intestinal metabolite composition containing a Lactobacillus plantarum strain

Abstract

The present invention relates to a composition for improving intestinal metabolite composition, which comprises a Lactobacillus plantarum strain. In one embodiment, the Lactobacillus plantarum strain can reduce intestinal polyamines (e.g., spermidine), and can be usefully used as a pharmaceutical composition, food (health functional food), feed composition, or anti-cancer aid for preventing, ameliorating, treating, or improving the prognosis of cancer, or as an anti-cancer aid.

Description

[Technical Field]

[0001] The present invention relates to a composition for improving intestinal metabolite composition, which comprises a Lactobacillus plantarum strain. [Background technology]

[0002] The microbiome refers to the entire set of microorganisms and their genetic information that exist in a specific environment. Microbiome information is being utilized in a variety of fields, including animals, agriculture, the ocean, and the environment, in addition to the human body. In particular, advances in human microbiome research based on advances in genetic information analysis and data analysis technology are expected to lead to growth in the diagnostic and healthcare industries.

[0003] Human intestinal microbes not only break down various substances that human enzymes cannot decompose and convert them into nutrients that human cells can absorb, but also suppress the growth of harmful bacteria from outside, preventing pathogenic infections. These intestinal microbes themselves or the various metabolic substances they secrete stimulate the numerous immune cells present in intestinal cells, activating or regulating the body's immune response. Furthermore, with the number and diversity of symbiotic microbial genes more than 100 times that of humans, the human microbiome is considered to be the human second genome, and its importance is recognized.

[0004] In particular, with the advancement of human microbiome research, there has been a rapid increase in research results showing that intestinal microbes are directly or indirectly related to numerous human diseases, and there is also an increasing number of studies showing that intestinal microbes are also related to various human cancers.

[0005] Polyamines are known to play a role in the growth, differentiation, and development of eukaryotic organisms. Polyamines, including spermine, spermidine, and the diamine precursor putrescine, are low-molecular-weight organic polycations containing at least two amino groups. Intracellular concentrations of polyamines can be maintained within specific physiological ranges through several regulatory mechanisms in normal cells. Furthermore, polyamine metabolism is known to be dysregulated in many neoplastic conditions, including cancer. Polyamine levels are elevated in various types of cancer, and a link between polyamine metabolism and oncogenic pathways, such as the mTOR and RAS pathways, is known. Therefore, polyamines may have potential therapeutic targets in the prevention and treatment of cancer.

[0006] Therefore, there is a need to develop substances for preventing or ameliorating diseases by improving the intestinal metabolite composition using the human microbiome. Summary of the Invention [Problem to be solved by the invention]

[0007] One aspect is to provide a composition for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

[0008] Another aspect is to provide a health functional food for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

[0009] Another aspect is to provide a probiotic composition for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted form of said strain, or a mixture thereof.

[0010] Another aspect is to provide a food composition for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted form of said strain, or a mixture thereof.

[0011] Another aspect is to provide a feed composition for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

[0012] Another aspect is to provide a pharmaceutical composition for the prevention or treatment of proliferative diseases, particularly cancer, comprising a Lactobacillus plantarum strain, a culture of said strain, a homogenate of said strain, or a mixture thereof as an active ingredient.

[0013] Another aspect is to provide an anti-cancer adjuvant comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

[0014] Another aspect is to provide a method for improving the intestinal metabolite composition of an individual, comprising administering to the individual an effective amount of a Lactobacillus plantarum strain, a culture of said strain, a disrupted form of said strain, or a mixture thereof.

[0015] Another aspect provides a method for preventing or treating cancer, comprising administering to an individual an effective amount of a Lactobacillus plantarum strain, a culture of said strain, a homogenate of said strain, or a mixture thereof.

[0016] Another aspect provides use of an effective amount of a Lactobacillus plantarum strain, a culture of said strain, a disruptant of said strain, or a mixture thereof for the manufacture of a formulation for improving the intestinal metabolite composition of an individual.

[0017] Another aspect provides use of an effective amount of a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof for the manufacture of a pharmaceutical preparation or a functional health food for preventing or treating cancer. [Means for solving the problem]

[0018] One aspect provides a composition (e.g., a pharmaceutical composition) for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of the strain, a disruptant of the strain, or a mixture thereof as an active ingredient.

[0019] Another aspect provides a method for improving the intestinal metabolite composition of an individual, comprising administering to the individual an effective amount of a Lactobacillus plantarum strain, a culture of the strain, a disrupted form of the strain, or a mixture thereof.

[0020] Another aspect provides use of an effective amount of a Lactobacillus plantarum strain, a culture of said strain, a disruptant of said strain, or a mixture thereof for the manufacture of a formulation for improving the intestinal metabolite composition of an individual.

[0021] Lactobacillus is a gram-positive, aerobic or facultatively anaerobic bacillus that is widely distributed in nature. Examples of Lactobacillus include Lactobacillus plantarum and Lactobacillus sakei. The present inventors conducted research to develop new strains with excellent anti-cancer effects and selected Lactobacillus plantarum GB104 as a candidate anti-cancer strain. This strain was deposited at the Korea Institute of Bioscience and Biotechnology (KIBIT) Biological Resources Center on January 14, 2020, under accession number KCTC14107BP. This strain corresponds to a probiotic strain and is harmless to the human body, allowing it to be used without side effects.

[0022] The Lactobacillus species has been renamed Limosilactobacillus or Lactiplantibacillus, and the modified strain names can be used interchangeably herein. For example, Lactobacillus plantarum has been renamed Lactiplantibacillus plantarum.

[0023] As used herein, the term "Lactobacillus plantarum GB104" may be used in conjunction with the L. Plantarum GB104 strain or the Lactobacillus plantarum GB104 strain (accession number: KCTC14107BP).

[0024] In one embodiment, the strain may be the strain deposited under accession number KCTC14107BP.

[0025] In one embodiment, the strain may be a strain comprising a 16S rRNA gene consisting of the nucleotide sequence of SEQ ID NO:1.

[0026] In one embodiment, the strain may have a 16S rRNA comprising the nucleotide sequence of SEQ ID NO: 1 or a nucleotide sequence having 97% or more identity thereto, specifically at least 93%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% identity to the nucleotide sequence of SEQ ID NO: 1 herein.

[0027] In one embodiment, the bacterial strain may be a live bacterium, a killed bacterium, or a cytoplasmic fraction obtained by disrupting the bacterial strain, preferably a live bacterium.

[0028] The term "culture" as used herein can be used interchangeably with "culture supernatant," "culture supernatant," "conditioned culture," or "conditioned medium," and can refer to the entire medium containing the Lactobacillus strain, its metabolites, and additional nutrients, obtained by culturing the strain for a certain period of time in a medium that can provide nutrients so that the strain can grow and survive in vitro. The culture refers to the product obtained by culturing a probiotic strain in a known medium, and the product may or may not contain the strain itself. The medium can be selected from known liquid or solid media, such as, but not limited to, MRS liquid medium, GAM liquid medium, MRS agar medium, GAM agar medium, and BL agar medium.

[0029] As used herein, the term "lysate" can be used interchangeably with "lysate" to refer to a solution or suspension of broken down microbial cells, such as Lactobacillus plantarum, in an aqueous medium. Cell lysates contain macromolecules, such as DNA, RNA, proteins, peptides, carbohydrates, lipids, and / or micromolecules, such as amino acids, sugars, fatty acids, or fractions thereof. Additionally, the lysate contains cellular debris, which may be smooth or granular in structure.

[0030] The culture medium may include the culture medium itself obtained by culturing the strain, a concentrate thereof, or a freeze-dried product thereof, or a culture supernatant obtained by removing the strain from the culture medium, or a concentrate or freeze-dried product thereof.

[0031] The culture medium may be obtained by culturing Lactobacillus plantarum in an appropriate medium (e.g., MRS plate medium) at a temperature above 10°C or below 40°C for a certain period of time, for example, 4 to 50 hours.

[0032] In one embodiment, the strain, a culture of the strain, or a lysate of the strain may contain acetylated spermidine.

[0033] In one embodiment, the strain, a culture of the strain, or a lysate of the strain may contain a polyamine acetylase, more specifically an enzyme involved in the acetylation of polyamines, including spermidine.

[0034] In one embodiment, the acetylated spermidine is N 1 -acetylspermidine, N 8 -acetylspermidine, or N 1 ,N 8 -diacetylspermidine. In one embodiment, the strain, a culture of the strain, or a lysate of the strain may contain glutaric acid or glutaconic acid.

[0035] In one embodiment, the glutarate or glutaric acid can be 2-oxoglutarate or 2-hydroxyglutarate. Further, in one embodiment, the glutaconic acid can be trans-glutarconic acid.

[0036] In another embodiment, the improvement in gut metabolite composition can include: - Reduction of the metabolite spermidine in the intestine or feces, - Increase in the intestinal or fecal metabolite acetylated spermidine, - Reduction of polyamines, their metabolites, in the intestine or feces, - Decreased activity of polyamine synthase in the intestine or feces, and -Increased activity of polyamine-degrading enzymes in the intestine or feces.

[0037] In one embodiment, the polyamine synthase may be ornithine decarboxylase (ODC), an enzyme that synthesizes spermidine from ornithine in cells, and the polyamine degrading enzyme may be spermidine / spermine N1-acetyltransferase (SSAT), an enzyme that acetylates spermidine, but these examples are illustrative and not limiting. Therefore, with the improvement in the intestinal metabolite composition of an individual administered the GB104 strain of the present invention, the activity of polyamine synthases (e.g., ornithine decarboxylase) in the individual's intestine or feces may decrease, the activity of polyamine degrading enzymes (e.g., spermidine / spermine N1-acetyltransferase) may increase, the levels of ornithine and polyamines (e.g., spermidine) in the intestine or feces may decrease, and the level of acetylated polyamines (e.g., acetylated spermidine) may increase.

[0038] The gut microbial composition, also known as the gut microbiome, can refer to the complex ecosystem within the gastrointestinal tract. Specifically, it comprises the entire gut microbial community, including bacteria, yeast, fungi, archaea, and viruses. The gut microbiome maintains several functions, including colonic fermentation of dietary fiber, nutrient extraction, synthesis of certain vitamins, prevention of pathogen colonization, maturation of the intestinal epithelium and immune system, release of metabolic products to systemic tissues, and regulation of gastrointestinal hormone secretion and neurological function. It is generally recognized that disruption of the normal balance in gut microbial composition can compromise the integrity of the intestinal barrier, which is often observed in many other diseases. The Lactobacillus plantarum GB104 strain of the present invention can restore and / or maintain a health-beneficial gut microbial composition in individuals through modulation of gut metabolite profiles (specifically, polyamine metabolism). Thus, the Lactobacillus plantarum strains herein may also provide compositions for restoring and / or maintaining a health-beneficial gut microbial composition (or for the prevention or treatment of disorders associated with impaired gut integrity).

[0039] Polyamine metabolism can include polyamine biosynthesis, catabolism, and transport. Natural polyamines can be synthesized in the cytoplasm of all cells. This biosynthesis begins with the urea cycle amino acids L-methionine and L-ornithine, followed by the decarboxylation of ornithine by ornithine decarboxylase (ODC) to form putrescine. Putrescine is a polyamine precursor in mammalian cells that generates spermine and spermidine when an aminopropyl group is added by decarboxylated S-adenosylmethionine (dcSAM). dcSAM is known to be generated by S-adenosylmethionine decarboxylase 1 (SAMDC, or adenosylmethionine decarboxylase 1, AMD1).

[0040] The role of polyamine transport in metabolism has led to the concept that polyamines are transported into cells via specific polyamine transport systems (PTSs). Polyamine uptake by PTSs is upregulated in proliferating cells, including tumor cells, suggesting that PTSs play a role in regulating intracellular polyamine concentrations. Because polyamine levels are upregulated in various cancer cell types, polyamines may be a common therapeutic target for cancer treatment. Rapidly proliferating cells, including tumor cells, exhibit higher activity of several enzymes involved in polyamine biosynthesis. Polyamine levels are elevated in cancer patients and may therefore be correlated with cancer development. High polyamine levels are known to be associated with the progression of neuroblastoma, hepatocellular carcinoma (HCC), prostate cancer, lung cancer, breast cancer, gastric cancer, and colorectal cancer (CRC). Decreased polyamine levels are known to induce cell death in postmitotic and senescent cells. Polyamines may also play a role in establishing tumor immunity, which promotes cancer cell growth through the excretion of sperm and their metabolites. Polyamines are also known to induce acquired chemoresistance to 5-fluorouracil and paclitaxel in colorectal and breast cancers. Reduction of polyamines can improve spontaneous IL-2 production, NK cell activity, and recovery of T lymphocyte populations without affecting tumor polyamines, preventing tumor-induced immunosuppression.

[0041] In one embodiment, the strain, a culture of the strain, or a lysate of the strain is capable of promoting the activity of intestinal immune cells or the expression of tight junction proteins between intestinal cells.

[0042] In one embodiment, the activity of intestinal immune cells can include an increase in the number of activated CD8+ T cells within immune cells, an increase in the proportion of activated CD8+ T cells within immune cells, or an increase in INF-γ secretion, more specifically, an increase in cytokine or interferon secretion by activated immune cells.

[0043] In other embodiments, the intestinal immune cells may include immune cells within the small intestine or large intestine (e.g., small intestinal intraepithelial lymphocytes (IELs), small intestinal lamina propria (siLP), colonic lamina propria (cLP)).

[0044] In one embodiment, the tight junction protein may include any one or more selected from the group consisting of claudin-1, claudin-2, claudin-3, claudin-4, claudin-5, ZO (Zonula Occludens)-1, ZO-2, ZO-3, and occludin.

[0045] Therefore, the bacterial strain, a culture thereof, or a lysate thereof may induce antitumor activity through the activation of intestinal immune cells, specifically through an increase in the proportion of cytotoxic T cells (CD8+ T cells) that are directly involved in suppressing cancer cell growth among immune cells in the small intestine or large intestine (e.g., small intestinal intraepithelial lymphocytes (IELs), small intestinal lamina propria (siLP), and colonic lamina propria (cLP)). Furthermore, the bacterial strain, a culture thereof, or a lysate thereof may promote or upregulate the expression of enterocyte tight junctions, thereby strengthening gut barrier function.

[0046] Therefore, in one embodiment, a Lactobacillus plantarum strain, a culture of the strain, a homogenate of the strain, or a mixture thereof is provided as a pharmaceutical composition, a food (health functional food), a feed composition, or an anti-cancer aid for preventing, ameliorating, treating, or improving the prognosis of cancer by reducing intestinal polyamines, or for anti-cancer aid.

[0047] In one embodiment, improving the intestinal metabolite composition of said individual may be for the prevention or treatment of cancer.

[0048] The cancer may be any one selected from the group consisting of gastric cancer, liver cancer, lung cancer, colon cancer, breast cancer, prostate cancer, ovarian cancer, pancreatic cancer, gallbladder cancer, biliary tract cancer, cervical cancer, thyroid cancer, laryngeal cancer, acute myeloid leukemia, brain tumor, neuroblastoma, retinoblastoma, salivary gland cancer, melanoma, bladder cancer, esophageal cancer, head and neck cancer, skin cancer, small intestine cancer, anal cancer, colon cancer, rectal cancer, kidney cancer, blood cancer, and lymphoma. Furthermore, the colon cancer may be a malignant tumor occurring in any site selected from the group consisting of the ascending colon, transverse colon, descending colon, sigmoid colon, and rectal mucosa.

[0049] The term "cancer," as used herein, refers to a physiological condition in an animal that is typically characterized by abnormal or uncontrolled cell growth. Cancer and cancer pathology can be associated with, for example, metastasis, interference with normally functioning surrounding cells, release of abnormal levels of cytokines or other secretory products, suppression or augmentation of inflammatory or immunological responses, neoplasia, premalignancy, malignancy, invasion of surrounding or distant tissues or organs, e.g., lymph node invasion, and the like.

[0050] The cancer can be a gastrointestinal cancer or a non-gastrointestinal cancer.

[0051] The gastrointestinal cancer is a malignant tumor that occurs in the gastrointestinal tract, such as the esophagus, stomach, small intestine, or large intestine. The gastrointestinal cancer may be, for example, one or more cancers selected from the group consisting of esophageal cancer, gallbladder cancer, liver cancer, biliary tract cancer, pancreatic cancer, gastric cancer, small intestine cancer, large intestine cancer, colon cancer, anal cancer, and rectal cancer, but is not limited to these, and one example may be large intestine cancer.

[0052] The non-gastrointestinal cancer includes, but is not limited to, malignant tumors occurring in organs other than the gastrointestinal tract or digestive system, such as, but not limited to, blood cancer, leukemia, acute myeloid leukemia, neuroblastoma, retinoblastoma, lung cancer, head and neck cancer, salivary gland cancer, melanoma, laryngeal cancer, prostate cancer, breast cancer, bladder cancer, kidney cancer, multiple myeloma, cervical cancer, thyroid cancer, ovarian cancer, urethral cancer, skin cancer, osteosarcoma, glioblastoma, brain tumor, or lymphoma.

[0053] In one embodiment of the present invention, the cancer may be colorectal cancer, which includes malignant tumors occurring in one or more sites selected from the group consisting of the ascending colon, transverse colon, descending colon, sigmoid colon, and rectal mucosa, and may be one or more types selected from the group consisting of adenocarcinoma, lymphoma, malignant carcinoid tumor, leiomyosarcoma, Kaposi's sarcoma, and squamous cell carcinoma, but is not limited to these.

[0054] According to one embodiment, the composition may contain 0.001 wt% to 80 wt% of the Lactobacillus plantarum strain based on the total weight of the composition. The dosage of the Lactobacillus plantarum strain may be 0.01 mg to 10,000 mg, 0.1 mg to 1,000 mg, 1 mg to 100 mg, 0.01 mg to 1,000 mg, 0.01 mg to 100 mg, 0.01 mg to 10 mg, or 0.01 mg to 1 mg. The strain is contained in the composition at a therapeutically or nutritionally effective amount, for example, when the strain is 10 3 ~10 16 CFU / g, 10 3 ~10 15 CFU / g, 10 3 ~10 14 CFU / g, 10 3 ~10 13 CFU / g, 10 3 ~10 12 CFU / g, 10 4 ~10 16 CFU / g, 10 4 ~10 15 CFU / g, 10 4 ~10 14 CFU / g, 10 4 ~10 13 CFU / g, 104 ~10 12 CFU / g, 10 5 ~10 16 CFU / g, 10 5 ~10 15 CFU / g, 10 5 ~10 14 CFU / g, 10 5 ~10 13 CFU / g, 10 5 ~10 12 CFU / g, 10 6 ~10 13 CFU / g, 10 6 ~10 12 CFU / g, 10 7 ~10 13 CFU / g, 10 7 ~10 12 CFU / g, 10 8 ~10 13 CFU / g or 10 8 ~10 12 They may be included in the composition at a CFU / g content or as cultures of the same number of live or dead bacteria.

[0055] Specifically, for adult patients, 1 × 10 3 ~1×10 16 CFU / g of live or killed bacteria can be administered in a single dose or in divided doses. However, the dosage can be varied depending on factors such as the formulation method, administration method, the patient's age, weight, sex, pathological condition, food, administration time, administration route, excretion rate, and reaction sensitivity, and those skilled in the art can appropriately control the dosage taking these factors into consideration. The number of administrations can be one or more than two within the range of clinically acceptable side effects, and the administration site can be one or more than two. For non-human animals, the dosage can be the same as that for humans per kg (body weight), or an amount converted to the dosage based on, for example, the volume ratio (e.g., average value) of organs (e.g., heart) between the target animal and humans can be administered.

[0056] Possible routes of administration may include oral, sublingual, parenteral (e.g., subcutaneous, intramuscular, intraarterial, intraperitoneal, intradural, or intravenous), rectal, topical (including transdermal), inhalation, and injection, or insertion of an implantable device or substance. Target animals for treatment in one embodiment include humans and other mammalian species, specifically humans, monkeys, mice, rats, rabbits, sheep, cattle, dogs, horses, pigs, etc. According to one embodiment, the composition contains a killed dried strain of bacteria, and can be administered at a dose of 1 g to 10 g, 0.5 g to 1.5 g, 2.5 g to 3.5 g, or 4.5 g to 5.5 g per dose, once to three times daily.

[0057] The term "therapeutically effective amount" as used herein refers to an amount of an anti-cancer agent for the methods and uses of the present invention, or a pharmaceutical composition containing an anti-cancer agent for the methods and uses of the present invention, that elicits the biological or medical response or desired therapeutic effect in a patient that a researcher, physician, or other clinician desires to achieve. A therapeutically effective amount of an anti-cancer agent can vary depending on factors such as the disease state, age, sex, and weight of the individual, and the ability of the anti-cancer agent to elicit a desired response in the individual. A therapeutically effective amount is one in which any toxic or detrimental effects are outweighed by the therapeutically beneficial effects.

[0058] The pharmaceutical composition according to one embodiment may contain pharmaceutically acceptable carriers and / or additives, such as sterile water, physiological saline, conventional buffers (such as phosphate, citric acid, and other organic acids), stabilizers, salts, antioxidants (such as ascorbic acid), surfactants, suspending agents, isotonicity agents, or preservatives. For topical administration, the composition may be combined with organic materials such as biopolymers, inorganic materials such as hydroxyapatite, specifically collagen matrices, polylactic acid polymers or copolymers, polyethylene glycol polymers or copolymers, and chemical derivatives thereof.

[0059] In one embodiment, the pharmaceutical composition may be an oral formulation.

[0060] In one embodiment, the oral formulation may be in the form of tablets, pills, capsules, lozenges, granules, powders, suspensions, sachets, or syrups.

[0061] When the pharmaceutical composition according to one embodiment is prepared in a formulation suitable for injection, the Lactobacillus bacteria may be dissolved or dispersed in a pharmaceutically acceptable carrier, or may be frozen in a solution in which they are dissolved or dispersed.

[0062] The pharmaceutical composition according to one embodiment may appropriately contain, if necessary depending on the administration method or formulation, a suspending agent, a solubilizing agent, a stabilizer, an isotonicity adjusting agent, a preservative, an anti-adsorption agent, a surfactant, a diluent, an excipient, a pH adjuster, a soothing agent, a buffer, a reducing agent, an antioxidant, etc. Pharmaceutically acceptable carriers and formulations suitable for the present invention, including those exemplified above, are described in detail in the literature "Remington's Pharmaceutical Sciences, 19th ed., 1995."

[0063] The pharmaceutical composition according to one embodiment can be prepared in unit dose form or in bulk containers by formulating it with pharmaceutically acceptable carriers and / or excipients according to a method readily understood by those skilled in the art, in which case the formulation may be in the form of a solution, suspension, or emulsion in an oily or aqueous medium, or in the form of a powder, granules, tablet, or capsule.

[0064] The pharmaceutical composition is administered in a pharmaceutically effective amount. As used herein, "pharmaceutical effective amount" refers to an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to any medical treatment, and the effective dose level can be determined according to factors including the type and severity of the patient's disease, the activity of the drug, sensitivity to the drug, administration time, administration route and excretion rate, treatment duration, concurrently used drugs, and other factors well known in the medical field.

[0065] The compositions of the present invention can be administered as an individual therapeutic agent or in combination with other therapeutic agents, and can be administered sequentially or simultaneously with conventional therapeutic agents, and can be administered singly or in multiple doses. Taking all of the above factors into consideration, it is important to administer an amount that can achieve the maximum effect with the minimum amount without side effects, and this can be easily determined by one skilled in the art.

[0066] Another aspect is to provide a health functional food for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

[0067] In one embodiment, improving the intestinal metabolite composition of the individual can include increasing beneficial intestinal bacteria, suppressing harmful bacteria, improving intestinal health by regulating the immune system, or improving bowel activity.

[0068] In one embodiment, the strain, a culture of the strain, or a lysate of the strain may contain acetylated spermidine or a polyamine acetylase.

[0069] In one embodiment, the acetylated spermidine is N 1 -acetylspermidine, N 8 -acetylspermidine, or N 1 ,N 8 -diacetylspermidine.

[0070] In one embodiment, the strain, a culture of the strain, or a disruptant of the strain may contain glutaric acid or glutaconic acid.

[0071] In one embodiment, the glutarate or glutaric acid can be 2-oxoglutarate or 2-hydroxyglutarate. Further, in one embodiment, the glutaconic acid can be trans-glutarconic acid.

[0072] In one embodiment, improving gut metabolite composition can include: - Reduction of the metabolite spermidine in the intestine or feces, - Increase in the intestinal or fecal metabolite acetylated spermidine, - Reduction of polyamines, their metabolites, in the intestine or feces, - Decreased activity of polyamine synthase in the intestine or feces, and -Increased activity of polyamine-degrading enzymes in the intestine or feces.

[0073] In one embodiment, the strain, a culture of the strain, or a lysate of the strain is capable of promoting or upregulating the activity of intestinal immune cells or the expression of tight junction proteins between intestinal cells.

[0074] In one embodiment, the activity of intestinal immune cells can include an increase in the number of activated CD8+ T cells within immune cells, an increase in the proportion of activated CD8+ T cells within immune cells, or an increase in INF-γ secretion, more specifically, an increase in cytokine or interferon secretion by activated immune cells.

[0075] In other embodiments, the intestinal immune cells may include immune cells within the small intestine or large intestine (e.g., small intestinal intraepithelial lymphocytes (IELs), small intestinal lamina propria (siLP), colonic lamina propria (cLP)).

[0076] In one embodiment, the tight junction protein may include any one or more selected from the group consisting of claudin-1, claudin-2, claudin-3, claudin-4, claudin-5, ZO (Zonula Occludens)-1, ZO-2, ZO-3, and occludin.

[0077] In one embodiment, the health functional food may be an oral formulation.

[0078] The strain, a culture of the strain, a disrupted product of the strain, or a mixture thereof, the administration route, the administration method, the administration dose, etc. are as described above.

[0079] In one embodiment, the health functional food may further comprise a nutrient-acceptable carrier.

[0080] The term "food-based acceptable" as used herein means that the compound exhibits the property of being non-toxic to cells or humans exposed to the compound.

[0081] The term "improvement" as used herein can refer to any action that at least reduces a parameter related to the condition being treated, such as the severity of symptoms. In this case, the health functional food can be used for the prevention or improvement of cancer, either before or after the onset of the disease, simultaneously or separately with a therapeutic drug.

[0082] In the health functional foods, the active ingredient may be added directly to the food or may be used in combination with other foods or food ingredients, or may be used appropriately according to conventional methods. The amount of the active ingredient to be mixed can be determined appropriately depending on the intended use (prevention or improvement). Generally, in the production of foods or beverages, the health functional foods can be added in an amount of about 15% by weight or less, more specifically about 10% by weight or less, based on the raw materials. However, in the case of long-term intake for health and hygiene purposes or health regulation purposes, the amount may be less than the above range.

[0083] The health functional food may be formulated into one selected from the group consisting of tablets, pills, powders, granules, powders, capsules, and liquid preparations, further comprising one or more of a carrier, a diluent, an excipient, and an additive. Foods to which the compound according to one embodiment can be added include various foods, powders, granules, tablets, capsules, syrups, beverages, gum, tea, vitamin complexes, health functional foods, etc.

[0084] Specific examples of the carrier, excipient, diluent, and additive may be at least one selected from the group consisting of lactose, dextrose, sucrose, sorbitol, mannitol, erythritol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium phosphate, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, polyvinylpyrrolidone, methylcellulose, water, gum syrup, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.

[0085] In addition to the active ingredient, the health functional food may contain other essential ingredients without particular limitation. For example, like regular beverages, it may contain various flavorings or natural carbohydrates as additional ingredients. Examples of the natural carbohydrates include conventional sugars such as monosaccharides (e.g., glucose, fructose, etc.), disaccharides (e.g., maltose, sucrose, etc.), polysaccharides (e.g., dextrin, cyclodextrin, etc.), and sugar alcohols (e.g., xylitol, sorbitol, erythritol, etc.). Other flavorings that can be advantageously used include natural flavorings (thaumatin, stevia extract (e.g., rebaudioside A, glycylrhizin, etc.)) and synthetic flavorings (saccharin, aspartame, etc.). The proportion of the natural carbohydrates can be appropriately determined by the selection of a person skilled in the art.

[0086] In addition to the above, the health functional food according to one embodiment may contain various nutrients, vitamins, minerals (electrolytes), flavorings such as synthetic flavorings and natural flavorings, coloring agents and enhancers (cheese, chocolate, etc.), pectinic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonation agents used in carbonated drinks, etc. Such ingredients may be used independently or in combination, and the proportions of such additives may also be appropriately selected by those skilled in the art.

[0087] The health functional food may be provided in a mixture with a conventionally known health functional food for preventing or improving cancer or other existing health functional foods, and the health functional food for preventing or improving cancer may be a conventionally known health functional food for preventing or improving metabolic diseases, an existing health functional food, or a newly developed health functional food.

[0088] When the health functional food contains other health functional foods that have cancer prevention or improvement effects, it is important to mix them in an amount that can achieve the maximum effect with the minimum amount without side effects, which can be easily determined by a person skilled in the art.

[0089] The food compositions for preventing or ameliorating cancer include all forms such as functional foods, nutritional supplements, health foods, and food additives, and the food compositions of this type can be prepared in various forms according to conventional methods known in the art.

[0090] The compositions herein can be considered food supplements. Food supplements, also known as dietary or nutritional supplements, can be considered another specific pharmaceutical product. They are prepared to supplement the diet and provide nutrients or beneficial ingredients that are not available or not available in sufficient amounts in a normal diet. Most food supplements are considered foods, but they are sometimes considered drugs, natural health products, or nutraceutical products. Within the meaning of the present invention, food supplements include nutraceuticals. Food supplements are usually sold over the counter without a prescription. When food supplements take the form of pills or capsules, they contain the same excipients as pharmaceuticals. However, food supplements can also take the form of food fortified with certain nutrients (e.g., infant formula). Therefore, in certain embodiments, the compositions of the present invention are food supplements.

[0091] The compositions according to the present invention may be administered as is, or mixed with an appropriate edible liquid or solid, or may be in the form of tablets, pills, capsules, lozenges, granules, powders, suspensions, sachets, syrups, or freeze-dried in unit dose form, or may be in the form of monodoses of freeze-dried compositions to be mixed in a separate liquid container provided together before administration.

[0092] The composition of the present invention may be included in various edible foods and foods, such as milk products, for infants. The term "edible product" as used herein broadly includes any form of product that can be ingested by an animal in any way (e.g., a product that is received by the organs of the body). The term "food product" is understood to mean an edible product that provides nutritional support to the body. Particularly interesting foods are food supplements and infant formulas. The food preferably comprises a carrier material, such as oatmeal gruel, lactic acid fermented foods, resistant starch, dietary fibers, carbohydrates, proteins, and glycosylated proteins. In certain embodiments, the bacterial cells of the present invention are homogenized with other ingredients, such as cereals or powdered milk, to form manufactured infant formula.

[0093] Another aspect is to provide a feed composition for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

[0094] The strain, a culture of the strain, a disrupted product of the strain, or a mixture thereof, the administration route, the administration method, the administration dose, etc. are as described above.

[0095] The feed composition can be prepared by adding the mixed strain composition in an appropriate effective concentration range according to various feed manufacturing methods known in the art, and can be used as a feed additive composition for the purpose of preventing or improving aging-related diseases.

[0096] The "feed" can mean any natural or artificial diet, meal, etc., or component of said meal, intended for or suitable for eating, ingesting, and digestion by an animal.

[0097] The type of feed is not particularly limited, and feed commonly used in the art can be used. Non-limiting examples of the feed include plant-based feeds such as grains, roots and fruits, food processing by-products, algae, fibers, pharmaceutical by-products, oils and fats, starches, gourds, and grain by-products; protein-based feeds, lipid-free feeds, oils and fats, minerals, oils and fats, single-cell proteins, zooplankton, and animal-based feeds such as food and drink.

[0098] Another embodiment provides an anti-cancer adjuvant comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

[0099] The strain, a culture of the strain, a disrupted product of the strain, or a mixture thereof, the administration route, the administration method, the administration dose, etc. are as described above.

[0100] As used herein, the term "adjuvant" refers to an agent that enhances and / or improves the therapeutic effect of an anticancer drug by supporting its efficacy, or that prevents or alleviates the adverse effects of the anticancer drug. The Lactobacillus plantarum strain of the present invention can improve the anticancer effect of other anticancer drugs by improving the composition of intestinal metabolites without itself placing a burden on the human body.

[0101] The other anticancer agent may be selected from the group consisting of conventional treatments that can be used in combination with other anticancer agents, such as chemotherapeutic agents, targeted anticancer agents, antibody therapeutic agents, immunological anticancer agents, and combinations thereof.

[0102] The term "chemo-cancer agent" as used herein is also referred to as an antitumor agent or a cytotoxic agent. It is a collective term for drugs that exhibit anticancer activity primarily by acting directly on DNA to block DNA replication, transcription, and translation processes, or by interfering with the synthesis of nucleic acid precursors in metabolic pathways, thereby inhibiting cell division. Specifically, a chemo-cancer agent may be any one selected from the group consisting of alkylating agents, microtubule inhibitors, antimetabolites, and topoisomerase inhibitors. The antitumor agent acts not only on tumor cells but also on normal cells, exhibiting cytotoxicity. Chemo-cancer agents can be used for maintenance therapy. Furthermore, the term "maintenance therapy" as used herein refers to a drug-based cancer treatment following initial anticancer therapy, intended to prevent or delay cancer recurrence.

[0103] The term "targeted anticancer drug" as used herein refers to a therapeutic agent that specifically kills cancer cells by targeting specific proteins or specific genetic alterations that are abundant only in cancer cells and blocking signals involved in the growth and development of cancer. Targeted anticancer drugs are classified into monoclonal antibodies that react extracellularly and small molecule substances that act intracellularly. Monoclonal antibodies are anticancer drugs that block cancer cell-inducing signals transmitted extracellularly and act on initiating signals related to proliferation, death, etc., while small molecule substances act on complex signal transduction that occurs intracellularly.

[0104] Specifically, target proteins may be epidermal growth factor receptor (EGFR), vascular endothelial growth factor receptor (VEGFR), CD20, CD38, RNAK-L, BTK, Bcr-abl, PDGFR / FGFR family, MEK / RAF, HER2 / Neu, ubiquitin, JAK, MAP2K, ALK, PARP, transforming growth factor beta receptor (TGFβR), proteasome, Bcl-2, c-Met, VR1, VR2, VR3, c-kit, AXL, RET, Braf, DNA methyltransferase (DNMT), CDK4 / 6, STING, and the like.

[0105] The term "antibody therapeutic agent" as used herein refers to a therapeutic agent that exhibits anticancer effects using an antibody that recognizes a specific protein on cancer cells as an antigen. Examples of antibody therapeutic agents include Cetuximab, Trastuzumab, Emtansine, Rituximab, Ibritumomab, Tositumomab, Brentuximab, Ofatumumab, Obinutuzumab, Necitumumab, Bevacizumab, Ramucirumab, Nivolumab, Pembrolizumab, Atezolizumab, Durvalumab, and Ipilimumab.

[0106] The term "immune anticancer agent" as used herein refers to a substance that inhibits the activity of immune checkpoint proteins, which suppress the differentiation, proliferation, and activity of immune cells, and is known to eliminate cancer cells by preventing them from evading the immune system. The immunoanticancer agent may be an antibody against any one selected from the group consisting of 2B4, 4-1BB (CD137), AaR, B7-H3, B7-H4, BAFFR, BTLA, CD2, CD7, CD27, CD28, CD30, CD40, CD80, CD83 ligand, CD86, CD160, CD200, CDS, CEACAM, CTLA-4, GITR, HVEM, ICAM-1, KIR, LAG-3, LAIR1, LFA-1 (CD11a / CD18), LIGHT, NKG2C, NKp80, OX40, PD-1, PD-L1, PD-L2, SLAMF7, TGFRp, TIGIT, Tim3, and VISTA. More specifically, the antibody may be any antibody selected from the group consisting of an anti-CTLA-4 antibody, an anti-PD-1 antibody, an anti-PD-L1 antibody, an anti-PD-L2 antibody, an anti-B7-H4 antibody, an anti-HVEM antibody, an anti-TIM3 antibody, an anti-GAL9 antibody, an anti-LAG3 antibody, an anti-VISTA antibody, an anti-KIR antibody, an anti-BTLA antibody, and an anti-TIGIT antibody, but is not limited to these.

[0107] The bacterial strain, culture thereof, or lysate thereof according to one embodiment may be administered in combination with the other anti-cancer agent.

[0108] As used herein, the terms "combination therapy" or "co-administration" or "in combination" refer to any form of simultaneous or combined therapy using at least two separate therapeutic agents. The components of the combination therapy can be administered simultaneously, sequentially, or in any order. The components can be administered in different doses, at different administration frequencies, or via different routes as appropriate.

[0109] Specifically, the combined administration may involve administering the Lactobacillus plantarum strain and the anticancer drug or antibiotic simultaneously, or administering the Lactobacillus plantarum strain followed by the anticancer drug or antibiotic. A combination therapy according to the present invention can be defined as providing a synergistic effect if the efficacy, measured, for example, through the degree of response, response rate, time to disease progression, or survival time, is therapeutically superior to the efficacy obtained by administering one or the other of the components of the combination therapy at a conventional dose. For example, the efficacy of a combination therapy is synergistic if its efficacy is therapeutically superior to the efficacy obtained by using each component alone. In particular, a synergistic effect is considered to exist if it does not adversely affect one or more of the degree of response, response rate, time to disease progression, and survival data, particularly the duration of response, reduces or decreases problematic side effects compared to those occurring when each component is used at a conventional dose, and allows for the reduction of the conventional doses of the Lactobacillus plantarum strain and the anticancer drug or antibiotic.

[0110] The term "administered simultaneously" as used herein is not particularly limited and means that the components of the combination therapy are administered substantially simultaneously, for example, as an admixture or in immediately subsequent order.

[0111] The term "sequentially administered" as used herein is not particularly limited and means that the components of a combination therapy are not administered simultaneously, but rather are administered one after the other or in groups with a specific time interval between administrations. The time interval may be the same or different between each administration of the components of the combination therapy, and may be selected from the ranges of, for example, 2 minutes to 96 hours, 1 day to 7 days, or 1 week, 2 weeks, or 3 weeks. Generally, the time interval between administrations may range from a few minutes to several hours, for example, from 2 minutes to 72 hours, 30 minutes to 24 hours, or 1 to 12 hours. Further examples include time intervals ranging from 24 to 96 hours, 12 to 36 hours, 8 to 24 hours, and 6 to 12 hours. [Effects of the Invention]

[0112] According to one embodiment, the Lactobacillus plantarum strain can reduce intestinal polyamines (e.g., spermidine), and can be usefully used as a pharmaceutical composition, food (health functional food), feed composition, or anti-cancer aid for preventing, ameliorating, treating, or improving the prognosis of cancer, or as an anti-cancer aid. [Brief explanation of the drawings]

[0113] [Figure 1] FIG. 1 is a graph showing the relative differences in the content of metabolites in GB104 culture supernatant and MRS culture supernatant (control group) by metabolic pathways via CE-TOF-MS. [Figure 2] FIG. 2 is a table showing the changes in trans-glutaconic acid content in metabolites in GB104 culture supernatant, L. plantarum F0077 strain, and MRS culture supernatant (control group) expressed as overall average fold change. [Figure 3] FIG. 3 is a table showing the changes in spermidine and three acetylated spermidine contents in metabolites in GB104 culture supernatant and MRS culture supernatant (control group) expressed as fold change based on the overall mean. [Figure 4] FIG. 4 is a graph showing the relative differences in the contents of spermidine and three types of acetylated spermidine among metabolites in GB104 culture supernatant and MRS culture supernatant (control group). [Figure 5] FIG. 5 is a graph showing the changes in the relative contents of spermidine and acetylspermidine in the colon cancer cell line HCT116 after treatment with GB104 culture supernatant and MRS culture supernatant (control group). [Figure 6] Figure 6 is a graph showing changes in the expression of polyamine synthase ODC and polyamine degrading enzyme SSAT after treatment with GB104 culture supernatant and MRS culture supernatant (control group) in colon cancer cell lines HCT116 and HT-29; ODC: Ornithine decarboxylase, SSAT: Spermidine / spermine N1-acetyltransferase. [Figure 7]FIG. 7 is a graph showing the change in relative spermidine content in feces after mice transplanted with colon cancer cells MC38 were treated with GB104 or PBS (control group). [Figure 8] FIG. 8 is a graph showing changes in N1-acetylspermidine / spermidine and ornithine contents in tumors after treatment of mice transplanted with colon cancer cells MC38 with GB104 and PBS (control group). [Figure 9] FIG. 9 is a graph showing the change in spermidine content in feces after antibiotic-treated mice were treated with GB104 and PBS (control group). [Figure 10] FIG. 10 is a graph showing changes in the number of intestinal immune cells after treatment of antibiotic-treated mice with GB104 or PBS (control group). [Figure 11] FIG. 11 is a graph showing changes in tight junction-related gene expression in intestinal tissues after treatment of antibiotic-treated mice with GB104 and PBS (control group); ZO: Zonula Occludens. [Figure 12] FIG. 12 is a graph showing the cell viability after treating the colon cancer cell line HCT116 with culture supernatants of bacterial strains different from GB104. [Figure 13] FIG. 13 is a graph showing the cell cycle of colon cancer cell line HCT116 after treatment with the culture supernatant of GB104 and a control strain (WCFS1). [Figure 14] FIG. 14 is a graph showing cell death after treating the colon cancer cell line HCT116 with the culture supernatant of GB104 and the control strain (WCFS1). DETAILED DESCRIPTION OF THE INVENTION

[0114] Preferred embodiments are presented below to aid in understanding the present invention. However, the following embodiments are provided to facilitate understanding of the present invention, and the contents of the present invention are not limited to the following embodiments. Various modifications can be made to the embodiments, and the embodiments are not limited to the embodiments disclosed below, and can be implemented in various forms.

[0115] Example 1. Isolation and identification of Lactobacillus plantarum GB104 strain

[0116] The isolation and identification of Lactobacillus plantarum strain GB104 was performed according to the methods described in Korean Patent Application Nos. 10-2020-0186738 and 10-2022-0080567, both of which are incorporated herein by reference in their entireties.

[0117] Briefly, Lactobacillus plantarum GB104 was isolated from vaginal samples of healthy women who visited a hospital for a health checkup. First, vaginal samples were collected with a cotton swab and pre-inoculated onto Rogosa SL (MRS) plates. They were then cultured in an anaerobic chamber at 37°C for 48 hours. Once bacterial colonies had grown, single colonies were subcultured onto new MRS plates for pure isolation. After isolation, the strains were cultured in MRS medium. Next, the Lactobacillus plantarum GB104 strain, which exhibited low levels of adipocyte accumulation inhibition and cytotoxicity, was selected from the cultured strains. To identify the selected Lactobacillus plantarum GB104 strain, the 16S rRNA gene sequence obtained by PCR using primers targeting the 16S rRNA gene was analyzed by Sanger sequencing. The 16S rRNA sequence of Lactobacillus plantarum GB104 is shown in SEQ ID NO: 1. The inventors named GB104 "Lactobacillus plantarum GB104" (deposit number: KCTC 14107BP) and deposited it with the Korean Collection for Type Cultures (KCTC) at the Korea Institute of Bioscience and Biotechnology on January 14, 2020. Furthermore, the strain name of Lactobacillus plantarum was changed to Lactiplantibacillus plantarum. In the following embodiments, the changed strain names of existing strains are used interchangeably.

[0118] Experimental Example 1. Analysis of polyamines and trans-glutaconic acid in GB104 culture supernatant and comparison of their contents

[0119] Metabolite analysis of the MRS culture supernatant (control) and GB104 culture supernatant was performed by Human Metabolome Technologies (HMT). The MRS and GB104 culture supernatant samples were added to Milli-Qwater containing internal standards and subjected to CE-TOF-MS analysis. Analysis of cation and anion metabolites was performed using an Agilent CE-TOF-MS system (Agilent Technologies Inc.), and the results are shown in Figure 1. Metabolite separation was performed using a fused silica capillary. Putative metabolites analyzed by HMT were identified through the HMT internal library.

[0120] FIG. 1 is a graph showing the relative differences in the content of metabolites in GB104 culture supernatant and MRS culture supernatant (control group) by metabolic pathways via CE-TOF-MS.

[0121] FIG. 2 is a table showing the changes in trans-glutaconic acid content in metabolites in GB104 culture supernatant, L. plantarum F0077 strain, and MRS culture supernatant (control group) in terms of fold change based on the total bacterial type.

[0122] FIG. 3 is a table showing the changes in spermidine and three acetylated spermidine contents in metabolites in GB104 culture supernatant and MRS culture supernatant (control group) expressed as fold change based on the overall mean.

[0123] FIG. 4 is a graph showing the relative differences in the contents of spermidine and three types of acetylated spermidine among metabolites in GB104 culture supernatant and MRS culture supernatant (control group).

[0124] As shown in Figures 1-4, a total of 296 metabolites were analyzed, including both substances identified through the HMT internal library and unknown substances. The OPLS-DA model, which was used to confirm differences between the two groups, was used to further analyze the metabolites produced and eliminated by GB104, resulting in the selection of 87 of the 296 metabolites. These metabolites included a variety of metabolites, including amino acids, peptides, nucleosides, nucleotides, organic acids, and sugars. Of particular note was the change in polyamines, which are known to be involved in cancer cell proliferation. GB104 reduced spermidine, a polyamine component, by acetylating it, while acetylated spermidine increased.

[0125] Experimental Example 2. Changes in metabolites and gene expression in cancer cells treated with GB104 culture supernatant

[0126] The effects of L. Plantarum GB104 culture supernatant on metabolites and gene expression in human colon cancer cell lines were investigated.

[0127] Human colon cancer cell line (HCT116) was placed in a 6-well plate at 2x10 5 The cells were dispensed into each well and cultured for 24 hours, after which the culture supernatant of the L. Plantarum GB104 strain was treated at a concentration of 10% and cultured for 24 hours under conditions of 37°C and 5% CO2. The culture supernatant was prepared by culturing the L. Plantarum strain in MRS medium for 8, 16, and 24 hours, then centrifuging to precipitate the strain, collecting only the supernatant, and filtering it through a 0.22 μm filter to obtain supernatants according to the culture time.

[0128] Changes in polyamine content in cancer cells were investigated by metabolite extraction and HPLC-DAD analysis of HCT116 cell pellets treated with GB104 culture supernatant at various times. The results are shown in Figure 5 as peak areas. Furthermore, changes in the expression of genes that inhibit polyamine synthesis and promote degradation by GB104 metabolites were investigated by RNA extraction and cDNA synthesis from HCT116 and HT-29 cell pellets treated with culture supernatant at various times. mRNA expression levels were then determined using the QuantStudio 3 Real-Time PCR Instrument. The results are shown in Figure 6. Polyamines, which are involved in cell growth and proliferation, are synthesized in particularly large amounts in cancer cells and play an important role in their proliferation. Polyamines are synthesized from ornithine by ornithine decarboxylase (ODC) in cells and then acetylated by spermidine / spermine N1-acetyltransferase (SSAT) for degradation. It has been reported that increased SSAT expression in cancer cells suppresses cancer cell proliferation, and high SSAT expression in cancer tissues leads to a favorable prognosis for anti-cancer treatment.

[0129] FIG. 5 is a graph showing the changes in the relative contents of spermidine and acetylspermidine in the colon cancer cell line HCT116 after treatment with GB104 culture supernatant and MRS culture supernatant (control group).

[0130] Figure 6 is a graph showing changes in the expression of polyamine synthase ODC and polyamine degrading enzyme SSAT after treatment with GB104 culture supernatant and MRS culture supernatant (control group) in colon cancer cell lines HCT116 and HT-29; ODC: Ornithine decarboxylase, SSAT: Spermidine / spermine N1-acetyltransferase.

[0131] As shown in Figure 5, in HCT116 colon cancer cells treated with GB104 culture supernatant, spermidine decreased and acetylspermidine increased in a culture time-dependent manner, confirming that GB104 acetylates polyamines.

[0132] As shown in Figure 6, the expression of the polyamine synthase ODC decreased and the expression of the polyamine degrading enzyme SSAT increased in both colon cancer cell lines HCT116 and HT-29 treated with GB104 culture supernatant. This confirmed that GB104 inhibits polyamine synthesis and increases degradation in cancer cells, thereby suppressing polyamine-induced cancer cell proliferation.

[0133] Experimental Example 3: Changes in spermidine content in feces of a mouse model 3.1. Comparison of changes in spermidine content in feces of MC-38 mouse colon cancer model

[0134] Changes in spermidine content in feces and tumors following administration of L. Plantarum GB104 were examined in a colon carcinoma MC-38 allograft model. Seven-week-old c57BL / 6 mice were inoculated with 2 × 10 colon cancer cell lines per mouse into the right flank. 5 Tumor models were established by subcutaneously injecting 100 μL of cells. Five days after tumor cell injection, tumors were 20–40 mm in size. 3 Only mice falling within the range were selected and randomly assigned to each group, and the L. Platarum GB104 strain was then added to the animal model at 1 × 10 per mouse. 9CFU were orally administered daily from day 6 until just before the end of the study. Feces were collected from all mice in each group on day 6 (just before oral GB104 administration) (day 0 sample) and again on day 12 (day 7 sample). Tumor samples were collected from mice on day 20 after tumor cell administration, the end of the study. Polyamines were extracted and derivatized from mouse feces and tumor samples, followed by HPLC-DAD analysis. Polyamines were detected in both feces and tumor samples, confirming that the changes in polyamine content were caused by GB104. The change in spermidine content in feces was expressed as a fold change, calculated by dividing the area of ​​the spermidine peak in fecal samples taken after 7 days of oral GB104 administration by the area of ​​the spermidine peak in fecal samples taken just before oral GB104 administration. This value was compared by group and is shown in Figure 7. The change in acetylspermidine content in the tumor was expressed as the area of ​​the N1-acetylspermidine peak divided by the area of ​​the spermidine peak, and the change in ornithine content was shown in FIG. 8 as the area value of the ornithine peak.

[0135] FIG. 7 is a graph showing the change in relative spermidine content in feces after mice transplanted with colon cancer cells MC38 were treated with GB104 or PBS (control group).

[0136] FIG. 8 is a graph showing changes in N1-acetylspermidine / spermidine and ornithine contents in tumors after treatment of mice transplanted with colon cancer cells MC38 with GB104 and PBS (control group).

[0137] As shown in Figure 7, GB104 dose-dependently reduced spermidine content, particularly at a dose of 1 × 10 9 The change in spermidine content in the group administered 100mg of ...

[0138] As shown in Figure 8, the proportion of acetylspermidine was higher in tumors from GB104-treated mice, indicating that GB104 not only acetylates polyamines to weaken cancer cell proliferation but also facilitates polyamine degradation. Furthermore, ornithine, a substrate of the polyamine synthase ODC, increased in tumors from GB104-treated mice, indicating that GB104 inhibited the activity of polyamine synthase.

[0139] 3.2. Comparison of changes in spermidine content in feces of antibiotic-treated mouse colon cancer models

[0140] Changes in fecal spermidine content following administration of the L. plantarum GB104 strain were examined using an antibiotic-treated mouse model. Antibiotics were mixed into sterilized drinking water, which was then shielded from light and given to mice ad libitum for 7 days. The antibiotic-treated drinking water was replaced with fresh water every 2-3 days. From the 7th day onwards, mice were provided with sterilized regular drinking water instead of antibiotic-containing water until the end of the study. Lyophilized GB104 was suspended in D-PBS and diluted to 1 × 10 9 After adjusting the CFU / head dose, the test substance was orally administered to each mouse using an oral tube at 200 μL per day for 7 days, starting from the day the antibiotic treatment was discontinued. After antibiotic treatment was discontinued, feces were collected from all mice in each group immediately prior to GB104 administration (day 0 sample), and again on day 7 of GB104 oral administration (day 7 sample). Polyamines were extracted from the mouse fecal samples, derivatized, and analyzed by HPLC-DAD. Spermidine was detected in the fecal samples, confirming that the change in spermidine content was caused by GB104. The change in spermidine content was expressed as a fold change, calculated by dividing the area of ​​the spermidine peak in the fecal samples collected after 7 days of GB104 oral administration by the area of ​​the spermidine peak in the fecal sample collected immediately prior to GB104 oral administration. This value was compared by group and is shown in Figure 9.

[0141] FIG. 9 is a graph showing the change in spermidine content in feces after antibiotic-treated mice were treated with GB104 and PBS (control group).

[0142] As shown in Figure 9, fecal spermidine content decreased with antibiotic treatment but increased on day 7, when the antibiotic's efficacy decreased. However, this increase in spermidine was significantly reduced by GB104.

[0143] Experimental Example 4: Changes in the number of intestinal immune cells by GB104 administration

[0144] In an antibiotic-treated mouse model, we confirmed changes in intestinal immune cells caused by administration of L. plantarum GB104 strain.

[0145] Antibiotics were mixed into sterilized drinking water, which was then shielded from light and given to mice ad libitum for 7 days. The antibiotic-treated drinking water was replaced with fresh water every 2-3 days. From the 7th day onwards, mice were given sterilized regular drinking water instead of antibiotic-containing water until the end of the study. Freeze-dried GB104 was suspended in D-PBS and given at 1 × 10 daily for 14 days from the end of antibiotic treatment. 9After oral administration of 200 μL of CFU / head, small intestinal and large intestinal tissues were obtained from mice. The small intestine was then cleaned of fat and Peyer's patches. The intestinal tissue was opened longitudinally, washed with PBS, and cut into 1-2 cm pieces. To remove epithelial cells, the tissue was stirred in FACS buffer (PBS containing 3% FBS, 20 mM HEPES, 100 U / mL penicillin, 100 μg / mL streptomycin, 1 mM sodium pyruvate, and 10 mM EDTA) at 37°C for 20 minutes. The filtered supernatant was used to further isolate intestinal epithelial cells (intraepithelial lymphocytes, IELs). The intestinal tissue fragments, from which epithelial cells had been removed, were washed with PBS, minced, and incubated at 37°C for 45 minutes in enzyme medium (RPMI 400 medium containing 400 U / mL Collagenase D and 10 μg / mL DNase I for small intestine; 800 U / mL Collagenase D and 10 μg / mL DNase I for large intestine, containing 3% FBS, 20 mM HEPES, 100 U / mL penicillin, 100 μg / mL streptomycin, 1 mM sodium pyruvate, and 1 mM NEAA). The enzyme reaction was stopped by adding 10 mM EDTA. The cells were passed through a strainer, resuspended in 40% Percoll solution, and then centrifuged with 75% Percoll solution. After centrifugation, the middle layer was collected and the lamina propria (LP) cells were isolated. The isolated immune cells were stained with fluorescent antibodies corresponding to the markers of the cells to be identified and analyzed using a FACSymphony instrument. The results are shown in FIG.

[0146] FIG. 10 is a graph showing changes in the number of intestinal immune cells after treatment of antibiotic-treated mice with GB104 or PBS (control group).

[0147] As shown in Figure 10, changes in immune cells in the small and large intestines were examined. The group administered with L. Plantarum GB104 showed a significant increase in the proportion of cytotoxic T cells (CD8+ T cells), known as immune cells that directly suppress cancer growth, compared to the control group. The amount of IFN-γ secreted by these cells also increased significantly. This confirmed that L. Plantarum GB104 has an effect on the increase and activation of CD8+ anti-tumor immune T cells.

[0148] Experimental Example 5. Changes in intestinal tight junction gene expression by GB104 administration

[0149] In an antibiotic-treated mouse model, we confirmed the changes in tight junction-related gene expression in the intestinal tissue caused by the administration of L. plantarum GB104 strain.

[0150] Antibiotics were mixed into sterilized drinking water, which was then shielded from light and given to mice ad libitum for 7 days. The antibiotic-treated drinking water was replaced with fresh water every 2-3 days. From the 7th day onwards, mice were given sterilized regular drinking water instead of antibiotic-containing water until the end of the study. Freeze-dried GB104 was suspended in D-PBS at a concentration of 1 × 10 9 After adjusting the dose to CFU / head, the test substance was orally administered to each mouse at 200 μL per day using an oral zondelle for 14 days starting from the end of antibiotic treatment. On day 14 after oral administration of GB104, the ileum tissue from the small intestine was excised and disrupted to extract RNA, which was then synthesized into cDNA. The mRNA expression levels of tight junction-associated genes (ZO-1, Occludin, and Claudin-4) were then measured using a QuantStudio 3 Real-Time PCR Instrument. The results are shown in Figure 11.

[0151] FIG. 11 is a graph showing changes in tight junction-related gene expression in intestinal tissues after treatment of antibiotic-treated mice with GB104 and PBS (control group); ZO: Zonula Occludens.

[0152] As shown in Figure 11, GB104 increased the expression of tight junction genes (ZO-1, Occludin, and Claudin-4) in the small intestine, confirming that GB104 can increase the expression of tight junction genes and strengthen the intestinal barrier function.

[0153] Experimental Example 6: Confirmation of cancer cell viability and cancer cell death effects by treatment with GB104 strain culture supernatant 6.1. Confirmation of cancer cell viability by treatment with GB104 culture supernatant

[0154] Human colon cancer cell lines were treated with various L. plantarum culture supernatants, including the L. plantarum GB104 strain, and cell viability was screened by MTT assay.

[0155] 2x10 human colon cancer cell line HCT116 cells were placed in a 96-well plate. 3 After 24 hours of incubation, cells were dispensed into each well and treated with 10% culture supernatant of various L. plantarum strains in culture medium supplemented with DFMO and aminoguanidine. Cultures were then incubated at 37°C and 5% CO2 for 72 hours. Culture supernatants were prepared by culturing L. plantarum strains in MRS medium, centrifuging the cultures, and collecting the supernatant. The supernatant was then filtered through a 0.22 μm filter. Using the Cell Proliferation Kit I (MTT) (Roche), the MTT solution was diluted to 0.5 mg / mL and the cells were incubated for an additional 4 hours. Purple formazan crystals, produced by metabolic activity in live cells, were dissolved in solubilization solution, and the absorbance was measured at 570 nm. The results are shown in Figure 12.

[0156] FIG. 12 is a graph showing the cell viability after treating the colon cancer cell line HCT116 with culture supernatants of bacterial strains different from GB104.

[0157] As shown in Figure 12, various L. Plantarum culture supernatants did not show the same inhibitory effect on cancer cell proliferation. In particular, GB104 culture supernatant showed an effect of reducing cancer cell viability by approximately 95%, confirming that it was the most effective in inhibiting cancer cell proliferation.

[0158] 6.2. Confirmation of cancer cell cycle by treatment with GB104 culture supernatant

[0159] The culture supernatant of L. Plantarum GB104 was treated with a human colon cancer cell line, and changes in the cancer cell cycle were observed using a flow cytometer.

[0160] 5x10 human colon cancer cell line HCT116 cells were placed in a 6-well plate. 4 After 24 hours of incubation, the cells were treated with 10% L. plantarum GB104 culture supernatant in a culture medium supplemented with DFMO and aminoguanidine and incubated at 37°C and 5% CO2 for 48 hours. The culture supernatant was prepared by culturing L. plantarum in MRS medium, centrifuging the culture to precipitate the cells, and filtering the supernatant through a 0.22 μm filter. After 48 hours, the cells were detached using trypsin-EDTA, centrifuged, and fixed in 70% ethanol. The cells were then stored at 4°C for at least 1 hour. After RNase A treatment and staining with propidium iodine (PI), the extent of cancer cell death was assessed by cell cycle analysis using a flow cytometer (BD FACSymphony A3 Cell Analyzer), as shown in Figure 13.

[0161] FIG. 13 is a graph showing the cell cycle of colon cancer cell line HCT116 after treatment with the culture supernatant of GB104 and a control strain (WCFS1).

[0162] As shown in Figure 13, the sub-G1 region of cells treated with L. plantarum GB104 culture supernatant was increased by approximately 9.8-fold compared to control cells treated with MRS, confirming that GB104 induces cancer cell death.

[0163] 6.3. Confirmation of the cancer cell death effect by treatment with GB104 culture supernatant

[0164] The culture supernatant of L. Platarum GB104 was treated with a human colon cancer cell line, and the cancer cell killing effect was confirmed using a flow cytometer.

[0165] 5x10 human colon cancer cell line HCT116 cells were placed in a 6-well plate. 4 After 24 hours of incubation, cells were added to each well and treated with 10% L. plantarum GB104 culture supernatant in a culture medium supplemented with DFMO and aminoguanidine. The culture supernatant was then incubated at 37°C and 5% CO2 for 48 hours. L. plantarum cultures were cultured in MRS medium, centrifuged to precipitate the cells, and the supernatant was collected and filtered through a 0.22 μm filter. After 48 hours, cells were harvested by trypsin-EDTA treatment and centrifuged. The FITC Annexin V Apoptosis Detection Kit with 7-AAD (BioLegend) was used to mix the cells with 100 μL of binding solution. Then, 5 μL of FITC Annexin V and 5 μL of 7-AAD were added and incubated at room temperature, protected from light, for 15 minutes. Fluorescence measurements to confirm cancer cell death were performed using a flow cytometer (BD FACSymphony A3 Cell Analyzer), and the results are shown in Figure 14. Normal, living cells are not labeled with Annexin V or 7-AAD, while cells in the early stage of cell death are labeled with Annexin V but not with 7-AAD. Cells in the later stage of cell death can be distinguished by being labeled with both Annexin V and 7-AAD.

[0166] FIG. 14 is a graph showing cell death after treating the colon cancer cell line HCT116 with the culture supernatant of GB104 and the control strain (WCFS1).

[0167] As shown in Figure 14, early and late cell death of cancer cells treated with L. plantarum GB104 culture supernatant was approximately four-fold increased compared to control cells treated with MRS. These results confirmed that GB104 is involved in inducing early cell death of cancer cells.

[0168] The above description of the present invention is for illustrative purposes only, and those skilled in the art will understand that the present invention can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. Therefore, it should be understood that the above-described embodiments are illustrative in all respects and are not limiting. [Accession number]

[0169] Name of depository institution: Korea Institute of Bioscience and Biotechnology Accession number: KCTC14107BP Date of acceptance: 20200114

Claims

1. A composition for improving the intestinal metabolic product composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

2. The composition of claim 1 , wherein the strain is deposited under accession number KCTC14107BP.

3. The composition of claim 1 , wherein the strain comprises the 16S rRNA of SEQ ID NO:

1.

4. The composition of claim 1 , wherein the strain, a culture of the strain, or a lysate of the strain contains acetylated spermidine.

5. The acetylated spermidine is N １ -acetylspermidine, N ８ -acetylspermidine, or N １ , N ８ The composition according to claim 4, wherein the hydroxybenzoate is diacetylspermidine.

6. The composition of claim 1 , wherein the strain, a culture of the strain, or a lysate of the strain comprises a polyamine acetyltransferase.

7. The composition of claim 1 , wherein the strain, a culture of the strain, or a disruption of the strain comprises glutaric acid or glutaconic acid.

8. The composition of claim 7, wherein the glutaric acid is 2-oxoglutaric acid or 2-hydroxyglutaric acid.

9. 2. The composition of claim 1, wherein the improvement in intestinal metabolite composition comprises any one or more of the following properties: - Reduction of the metabolite spermidine in the intestines or feces, - Increase in the intestinal or fecal metabolite acetylated spermidine, - Reduction of polyamines, which are metabolic products in the intestine or feces, - reduced activity of polyamine synthases in the intestine or faeces, and - Increased activity of polyamine degrading enzymes in the intestine or feces.

10. The composition of claim 1, wherein the strain, a culture of the strain, or a lysate of the strain promotes the activity of intestinal immune cells or the expression of tight junction proteins between intestinal cells.

11. The composition according to claim 10, wherein the activity of intestinal immune cells comprises an increase in the number of activated CD8+ T cells within the immune cells or an increase in INF-γ secretion.

12. The composition according to claim 10, wherein the tight junction protein is any one or more selected from the group consisting of ZO (Zonula Occludens)-1, ZO-2, ZO-3, occludin, claudin-1, claudin-2, claudin-3, and claudin-4.

13. A pharmaceutical composition for preventing or treating cancer, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

14. The strain is 10 ３ ~10 １６ 14. The pharmaceutical composition of claim 13, wherein the amount of the virus is in an amount of 0.01 cfu / g.

15. 14. The pharmaceutical composition of claim 13, which is an oral formulation.

16. The pharmaceutical composition of claim 13, wherein the cancer is any one selected from the group consisting of gastric cancer, liver cancer, lung cancer, colon cancer, breast cancer, prostate cancer, ovarian cancer, pancreatic cancer, gallbladder cancer, biliary tract cancer, cervical cancer, thyroid cancer, laryngeal cancer, acute myeloid leukemia, brain cancer, neuroblastoma, retinoblastoma, salivary gland cancer, melanoma, bladder cancer, esophageal cancer, head and neck cancer, skin cancer, small intestine cancer, anal cancer, colon cancer, rectal cancer, kidney cancer, blood cancer, and lymphoma.

17. 17. The pharmaceutical composition of claim 16, wherein the colon cancer occurs at any site selected from the group consisting of the ascending colon, transverse colon, descending colon, sigmoid colon, and rectal mucosa.

18. A health functional food for improving the intestinal metabolic product composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

19. The health functional food according to claim 18, wherein the strain, a culture of the strain, or a disrupted product of the strain contains acetylated spermidine or polyamine acetyltransferase.

20. The health functional food according to claim 18, wherein the strain, a culture of the strain, or a disrupted product of the strain contains glutaric acid or glutaconic acid.

21. The health functional food according to claim 18, wherein the improvement of intestinal metabolite composition includes: - Reduction of the metabolite spermidine in the intestines or feces, - Increase in the intestinal or fecal metabolite acetylated spermidine, - Reduction of polyamines, which are metabolic products in the intestine or feces, - reduced activity of polyamine synthases in the intestine or faeces, and - Increased activity of polyamine degrading enzymes in the intestine or feces.

22. The health functional food according to claim 18, wherein the strain, a culture of the strain, or a homogenate of the strain promotes the activity of intestinal immune cells or the expression of tight junction proteins between intestinal cells.

23. The health functional food according to claim 18, which is an oral formulation.

24. An anti-cancer adjuvant comprising, as an active ingredient, a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof.

25. A feed composition for improving the intestinal metabolite composition of an individual, comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof as an active ingredient.

26. A method for improving the intestinal metabolite composition of an individual, comprising administering to the individual in need thereof an effective amount of a Lactobacillus plantarum strain, a culture of said strain, a disrupted form of said strain, or a mixture thereof.

27. A method for preventing or treating cancer, comprising the step of administering an effective amount of a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof to an individual in need thereof.

28. Use of a composition comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof for the manufacture of a preparation for improving the intestinal metabolite composition of an individual.

29. Use of a composition comprising a Lactobacillus plantarum strain, a culture of said strain, a lysate of said strain, or a mixture thereof for the manufacture of a pharmaceutical preparation for preventing or treating cancer.

30. Use of a composition comprising a Lactobacillus plantarum strain, a culture of said strain, a disrupted product of said strain, or a mixture thereof for producing a health functional food for preventing or treating cancer.

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