Novel pericodepside and composition for inhibiting cancer metastatis and invasion, or treating cancer comprising the novel pericodepside

Abstract

The present invention relates to novel pericodecside compounds that exhibit cytotoxicity to cancer cells, inhibit the expression of EMT markers in cancer cells, and inhibit metastasis and invasion of cancer cells, pharmaceutically or food-grade acceptable salts thereof, and compositions for inhibiting cancer metastasis and invasion or for treating cancer comprising these compounds.

Description

Technology Field

[0001] The present invention relates to a novel pericodeptide compound and a composition for inhibiting cancer metastasis and invasion or for improving or treating cancer containing the same. More specifically, the invention relates to a novel pericodeptide compound that has cytotoxicity against cancer cells and inhibits cancer cell motility by downregulating epithelial-mesenchymal transition (EMT) markers, which are the basic mechanisms of cancer cell motility and progression; a pharmaceutically or food-acceptable salt thereof; and a composition for inhibiting cancer metastasis and invasion or for improving or treating cancer containing said novel pericodeptide compound and / or a pharmaceutically or food-acceptable salt thereof as an active ingredient. Background Technology

[0002] Cancer is one of the leading causes of death worldwide. According to GLOBOCAN 2020 data, nearly 10 million people died from cancer in 2020. Breast cancer is the most frequently diagnosed cancer globally. Lung, prostate, stomach, and colorectal cancers are among the top cancers associated with death, with lung cancer showing the second-highest cancer-related mortality rate in 2020. Brain cancer is the 21st most common cause of death but has the lowest survival rate of all types of cancer. Tumor metastasis is a major cause of treatment failure, and the prognosis for patients with metastatic tumors remains poor. Therefore, new therapeutic approaches targeting tumor metastasis are needed.

[0003] Metastatic cancer occurs when cancer cells spread from their site of origin to other parts of the body. The motility of cancer cells contributes to tumor metastasis through various stages, such as migration, invasion, and movement to distant sites. Epithelial-mesenchymal transition (EMT) is a common mechanism of tumor metastasis and progression. During EMT, tumor epithelial cells lose intercellular adhesion and acquire migratory and invasive characteristics due to the upregulation of EMT transcription factors such as N-cadherin, Snail, Slug, Twist, and ZEB1 / 2. EMT is a fundamental early stage of oncogenesis. Therefore, inhibiting EMT is a key objective in the development of anti-metastatic therapeutics.

[0004] Natural products derived from lichens are chemically diverse and biologically active, and have the potential to be a valuable source of compounds capable of addressing various health problems. Depsides and their derivatives are promising natural products found in lichens, characterized by a structure containing two or more phenolic groups similar to orsellinic acid linked by ester bonds. These compounds exhibit anticancer, antibacterial, antioxidant, and anti-inflammatory activities, suggesting potential utility for therapeutic purposes [Non-patent Literature 1]. Atranorin, evernic acid, and olivetoric acid are representative examples of depsides with cytotoxic activity, suggesting that they could be potential anticancer agents [Non-patent Literatures 2-4].

[0005] Advancements in biotechnology and genetic engineering have facilitated the biosynthesis of various depths, making them more economical to produce [Non-patent Literature 5-7]. In addition, the production of specific depths and their derivatives through biosynthesis has provided an environmentally friendly and sustainable alternative compared to traditional chemical synthesis methods. Furthermore, improvements in the biosynthesis of these compounds will ultimately facilitate large-scale industrial production and use.

[0006] Recently, the inventors of the "clean host" Ascotina laviey ( Ascochyta rabiei Three lichen genes in the ) strain ( atr1, atr2 and atr3 Atlanorin was biosynthesized by heterologous expression of ) [Non-patent Literature 8]. In the process of atlanorin biosynthesis, lichen depths 4- O -Dimethylbaratic acid(4- O The formation of -demethylbaratic acid) was catalyzed by ATR1, a non-reducing polyketide synthase. 4- O -Dimethylbaratic acid is hydroxylated to proatranorin II and then further oxidized by the CYP450 enzyme ATR2 to produce proatranorin III (1). O - Methyltransferase ATR3 completed the biosynthesis of atranorin by introducing a methyl group to the carboxylic acid group of proatranorin III [Non-patent Literature 9]. Prior art literature

[0007]

[01] Ibrahim, S. R.; Sirwi, A.; Eid, B. G.; Mohamed, S. G.; Mohamed, G. A. Fungal depsides―Naturally inspiring molecules: Biosynthesis, structural characterization, and biological activities. Metabolites. 2021, 11, 683.

[02] Zhou, R.; Yang, Y.; Park, S.Y.; Nguyen, T. T.; Seo, Y.W.; Lee, K. H.; Lee, J. H.; Kim, K. K.; Hur, J.S.; Kim, H. G. The lichen secondary metabolite atranorin suppresses lung cancer cell motility and tumorigenesis. Sci Rep. 2017, 7, 8136.

[03] Kalin. S. N.; Altay, A.; Budak, H. Effect of evernic acid on human breast cancer MCF7 and MDA-MB-453 cell lines via thioredoxin reductase 1: A molecular approach. J Appl Toxicol. 2023, 43, 1148-1158.

[04] Emsen, B.; Aslan, A.; Togar, B.; Turkez, H. In vitro antitumor activities of the lichen compounds olivetoric, physodic and psoromic acid in rat neuron and glioblastoma cells. Pharm Biol. 2016, 54, 1748-1762.

[05] Chen, L.; Wei, X.; Matsuda, Y.Depside bond formation by the starter-unit acyltransferase domain of a fungal polyketide synthase. J Am Chem Soc. 2022, 144, 19225-19230.

[06] Kim, W.; Liu, R.; Woo, S.; Kang, K. B.; Park, H.; Yu, Y. H.; Ha, H.-H.; Oh, S.-Y.; Yang, J. H.; Kim, H. J. Linking a gene cluster to atranorin, a major cortical substance of lichens, through genetic dereplication and heterologous expression. mbio. 2021, 12, 10-1128.

[07] Liu, Q.; Zhang, D.; Gao, S.; Cai, X.; Yao, M.; Xu, Y.; Gong, Y.; Zheng, K.; Mao, Y.; Yang, L. Didepside Formation by the Nonreducing Polyketide Synthase Preu6 of Preussia isomera Requires Interaction of Starter Acyl Transferase and Thioesterase Domains. Angew Chem Int Ed Engl. 2023, 135, e202214379.

[08] Kim, W.; Liu, R.; Woo, S.; Kang, K. B.; Park, H.; Yu, Y. H.; Ha, H.-H.; Oh, S.-Y.; Yang, J. H.; Kim, H. J. Linking a gene cluster to atranorin, a major cortical substance of lichens, through genetic dereplication and heterologous expression. mbio. 2021, 12, 10-1128.

[09] Kim, W.; Liu, R.; Woo, S.; Kang, K.B.; Park, H.; Yu, YH; Ha, H.-H.; Oh, S.-Y.; Yang, J.H.; Kim, HJ Linking a gene cluster to atranorin, a major cortical substance of lichens, through genetic dereplication and heterologous expression. mbio. 2021, 12, 10-1128.

[10] Yamada, T.; Iritani, M.; Ohishi, H.; Tanaka, K.; Minora, K.; Doi, M.; Numata, A. Pericosines, antitumour metabolites from the sea hare-derived fungus Periconia byssoides. Structures and biological activities. Org Biomol Chem. 2007, 5, 3979-3986.

[11] Numata, A.; Iritani, M.; Yamada, T.; Minora, K.; Matsumura, E.; Yamori, T.; Tsuruo, T. Novel antitumour metabolites produced by a fungal strain from a sea hare. Tetrahedron letters. 1997, 38, 8215-8218. The problem to be solved

[0008] The inventors heterogeneously express atr1 and atr2 A. rabiei We have newly discovered a previously unknown compound, pericodepside, which is a derivative of proatranorin III (1) produced by a strain, and have completed the present invention by discovering that the pericodepside compound downregulates the expression of EMT markers in addition to cytotoxicity and inhibits the motility of cancer cells.

[0009] Accordingly, one objective of the present invention is to provide a novel pericodeptide compound that inhibits the motility of cancer cells by downregulating EMT markers.

[0010] Another objective of the present invention is to provide a composition for inhibiting cancer metastasis and invasion comprising the novel pericodecid compound or a pharmaceutically acceptable salt of the compound as an active ingredient.

[0011] Another objective of the present invention is to provide a composition for treating cancer comprising the novel pericodepsid compound or a pharmaceutically acceptable salt of the compound as an active ingredient.

[0012] Another objective of the present invention is to provide a food composition for inhibiting cancer metastasis and invasion and for improving cancer, comprising the novel pericodeptide compound or a food-grade acceptable salt of the compound as an active ingredient. means of solving the problem

[0013] In one embodiment, the present invention provides a novel compound represented by the following chemical formula 2 or a pharmaceutically acceptable salt thereof.

[0014] [Chemical Formula 2]

[0015]

[0016] The compound represented by the above chemical formula 2 is a pericodepside compound that is a derivative of proatranorin III (1), and its molecular formula is C 26 H 25 O 13 ..., and the molecular weight is 545.1295. In this specification, the compound represented by the above chemical formula 2 is referred to as Compound 2.

[0017] The pericodepside compound represented by the above chemical formula 2 has cytotoxicity against cancer cells, inhibits cell invasion and motility, reduces the expression of N-cadherin and E-cadherin as EMT markers, and reduces the expression of Snail, Slug, Twist, and ZEB1 / 2, which are regulators of EMT markers, so it can not only improve or treat cancer but also inhibit cancer metastasis and invasion.

[0018] In one specific example, pericodepsid exhibited stronger inhibitory activity than proatranorin III (Compound 1) on cell viability, invasion, and cell growth. Additionally, pericodepsid downregulated the expression of β-catenin, phosphorylated STAT, and NF-кB, thereby inhibiting the expression of the EMT effector N-cadherin and its transcription factors Snail, Slug, Twist, and ZEB1 / 2, which inhibited cancer invasion and proliferation in U87, MCF-7, and PC3 cells (Fig. 8).

[0019] The pericodeptide compound represented by Chemical Formula 2 above may exist in the form of a pharmaceutically acceptable salt. As the salt, an acid addition salt formed by a pharmaceutically acceptable free acid is useful. The term "pharmaceutically acceptable salt" in the present invention refers to any organic or inorganic addition salt of the said compound at a concentration having a relatively non-toxic and harmless active effect on the patient, such that side effects caused by this salt do not impair the beneficial efficacy of the pericodeptide compound.

[0020] Acid addition salts are prepared by conventional methods, for example, by dissolving a compound in an excess amount of an aqueous acid solution and precipitating the salt using a water-miscible organic solvent, for example, methanol, ethanol, acetone, or acetonitrile. An equal molar amount of the compound and an acid or alcohol (e.g., glycol monomethyl ether) in water may be heated, and then the mixture may be dried by evaporation or the precipitated salt may be filtered by suction.

[0021] At this time, organic acids and inorganic acids may be used as free acids. Inorganic acids may include hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid, etc., and organic acids may include methanesulfonic acid, p-toluenesulfonic acid, acetic acid, trifluoroacetic acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, manderic acid, propionic acid, citric acid, lactic acid, glycolic acid, gluconic acid, galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, aspartic acid, ascorbic acid, carboxylic acid, vanillic acid, hydroiodic acid, etc., but are not limited to these.

[0022] In addition, pharmaceutically acceptable metal salts can be prepared using a base. Alkali metal salts or alkaline earth metal salts are obtained, for example, by dissolving a compound in an excess amount of an alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the undissolved compound salt, and then evaporating and drying the filtrate. At this time, it is pharmaceutically suitable to prepare sodium, potassium, or calcium salts as the metal salts, but is not limited to these. In addition, the corresponding silver salt can be obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (e.g., silver nitrate).

[0023] Pharmaceutically acceptable salts of the compounds of the present invention comprise salts of acidic or basic groups that may be present in the pericodecide compound of Formula 2, unless otherwise indicated. For example, pharmaceutically acceptable salts may include sodium, calcium, and potassium salts of the hydroxyl group, and other pharmaceutically acceptable salts of the amino group include hydrobromide, sulfate, hydrogen sulfate, phosphate, hydrogen phosphate, dihydrogen phosphate, acetate, succinate, citrate, tartrate, lactate, mandelate, methanesulfonate (mesylate), and p-toluenesulfonate (tosylate) salts, and may be prepared through methods for preparing salts known in the art.

[0024] As a salt of the pericodeptide compound of Formula 2 of the present invention, any pharmaceutically acceptable salt that exhibits pharmacological activity equivalent to that of the pericodeptide compound of Formula 2, for example, inhibits the migration and invasion of cancer cells to delay death caused by metastasis of cancer cells or treats cancer, may be used without limitation.

[0025] In addition, the pericodeptide compound of Formula 2 according to the present invention comprises, without limitation, its pharmaceutically acceptable salts as well as solvates such as possible hydrates that can be prepared therefrom and all possible stereoisomers. The solvates and stereoisomers of the pericodeptide compound of Formula 2 can be prepared from the pericodeptide compound of Formula 2 using methods known in the art.

[0026] Furthermore, the pericodecid compound of Formula 2 according to the present invention may be prepared in a crystalline or amorphous form, and if prepared in a crystalline form, it may be optionally hydrated or solvated. The present invention may include not only the stoichiometric hydrate of the pericodecid compound of Formula 2 but also compounds containing varying amounts of water. The solvates of the pericodecid compound of Formula 2 according to the present invention include both stoichiometric solvates and non-stoichiometric solvates.

[0027] The pericodecid compound of Formula 2 according to the present invention can be prepared by the following exemplary method, and a specific example is the reaction scheme described in the examples below.

[0028] The manufacturing method of the present invention may use commercially available compounds as reactants in the above reaction schemes, either by purchasing and using them as they are, or by synthesizing them by performing one or more reactions known in the art as they are or by appropriately modifying them. For example, the synthesis may be performed by carrying out one or more reactions in a series of sequences, taking into account the presence, type, and / or location of reactive functional groups and / or heteroelements included in the skeletal structure, but is not limited thereto.

[0029] In another aspect, the present invention provides a pharmaceutical composition for inhibiting cancer metastasis and invasion, or for treating cancer, comprising a pericodecid compound of Formula 2 or a pharmaceutically acceptable salt thereof as an active ingredient.

[0030] The pericodepsid compound of Formula 2, which is the active ingredient of the present invention, heterogeneously expresses atr1 and atr2. A. rabieiIt is a newly discovered compound isolated from a solid culture of a strain. The pericodepside compound is a depside-pericosine conjugate, the depside portion being composed of 3-methylorcelic acid and hematomic acid interconnected by ester bonds, and is a compound formed by the additional esterification of the depside portion and a pericosine derivative.

[0031] As a result of confirming the degree of cancer cytotoxicity, the degree of expression of EMT markers in cancer cells, and the inhibitory effect on cancer cell invasion and metastasis regarding the newly discovered pericodeptide compound above, the pericodeptide compound of the present invention has the effect of killing cancer cells, inhibiting the expression of EMT markers in cancer cells, and inhibiting cancer cell invasion and metastasis, and thus can be used for inhibiting cancer metastasis and invasion and for cancer treatment containing the same as an active ingredient.

[0032] The aforementioned cancer is a disease related to the regulation of cell death, referring to a disease that occurs when the balance of normal cell death (apoptosis) is disrupted, leading to excessive cell proliferation. These abnormally proliferated cells may, in some cases, invade surrounding tissues and organs to form a tumor, destroying or deforming normal structures within the body; this condition is called cancer.

[0033] The above types of cancer include brain and spinal cord tumors, brain cancer, head and neck cancer, lung cancer, breast cancer, thymoma, esophageal cancer, stomach cancer, colorectal cancer, liver cancer, pancreatic cancer, biliary tract cancer, kidney cancer, bladder cancer, prostate cancer, testicular cancer, germ cell tumors, ovarian cancer, cervical cancer, endometrial cancer, lymphoma, leukemia (e.g., acute or chronic leukemia), osteosarcoma, multiple myeloma, sarcoma, melanoma, malignant melanoma, and skin cancer. The anticancer composition of the present invention may be used without limitation on the type of cancer, but for the purposes of the present invention, it may be usefully used to treat one or more cancers selected from the group consisting of liver cancer, lung cancer, stomach cancer, pancreatic cancer, colorectal cancer, cervical cancer, breast cancer, prostate cancer, ovarian cancer, brain cancer, osteosarcoma, bladder cancer, head and neck cancer, kidney cancer, melanoma, leukemia, and lymphoma.

[0034] The term “treatment” in this invention refers to any act in which cancer symptoms are improved or benefited by the administration of the pharmaceutical composition of this invention.

[0035] The pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier, diluent, or excipient, and may be formulated into various forms according to conventional methods for each intended use, such as oral formulations like powders, granules, tablets, capsules, suspensions, emulsions, syrups, and aerosols, and injectable formulations like sterile injectable solutions, and may be administered orally or through various routes including intravenous, intraperitoneal, subcutaneous, rectal, and local administration. Examples of suitable carriers, excipients, or diluents that may be included in such compositions include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. Additionally, the composition of the present invention may further include fillers, anticoagulants, lubricants, wetting agents, fragrances, emulsifiers, preservatives, etc.

[0036] Solid dosage forms for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid dosage forms are formulated by mixing at least one excipient, such as starch, calcium carbonate, sucrose, lactose, gelatin, etc., into the above composition. In addition, in addition to simple excipients, lubricants such as magnesium stearate and talc may be used.

[0037] Examples of oral liquid formulations include suspensions, liquid formulations, emulsions, syrups, etc., and in addition to commonly used simple diluents such as water and liquid paraffin, various excipients, such as humectants, sweeteners, flavorings, and preservatives, may be included.

[0038] Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, and suppositories. As non-aqueous solvents and suspensions, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used. As bases for suppositories, Witepsol, Macrogol, Tween 61, cocoa oil, laurin oil, glycerogelatin, etc., may be used. Meanwhile, injectable preparations may contain conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers, and preservatives.

[0039] The above formulation can be prepared by conventional mixing, granulation, or coating methods and contains an active ingredient in an amount effective for medical treatment, specifically for inhibiting cancer metastasis and invasion or treating cancer.

[0040] At this time, the composition of the present invention is administered in a pharmaceutically effective amount. The term "pharmaceutically effective amount" in the present invention refers to an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment and that does not cause side effects. The effective dose level may be determined based on factors including the patient's health status, type and severity of the disease, drug activity, sensitivity to the drug, method of administration, time of administration, route of administration and elimination rate, duration of treatment, drugs used in combination or concurrently, and other factors well known in the medical field. The composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered as a single or multiple doses. It is important to administer an amount that obtains maximum effect with a minimum amount without side effects, taking all of the above-mentioned factors into consideration, and this can be easily determined by a person skilled in the art.

[0041] For example, since the dosage may be increased or decreased depending on the route of administration, severity of the disease, gender, body weight, age, etc., the above dosage does not limit the scope of the present invention in any way.

[0042] The preferred dosage of the compound of the present invention varies depending on the patient's condition and weight, the severity of the disease, the form of the drug, the route of administration, and the duration, but can be appropriately selected by those skilled in the art.

[0043] In another aspect, the present invention provides a method for inhibiting cancer metastasis and invasion, or treating cancer, comprising the step of administering the pharmaceutical composition of the present invention to an individual in need thereof.

[0044] The term "individual" in this invention refers to any animal including humans, monkeys, cattle, horses, sheep, pigs, chickens, turkeys, quails, cats, dogs, mice, rats, rabbits, or guinea pigs that have the potential for cancer metastasis and invasion, or have metastasized and invaded or have developed cancer. By administering the pharmaceutical composition of this invention to an individual, cancer metastasis and invasion can be inhibited or cancer can be effectively treated. Furthermore, since the pharmaceutical composition of this invention inhibits cancer metastasis and invasion and exhibits a therapeutic effect on cancer by suppressing the expression of EMT markers, a synergistic effect can be produced by administering it in combination with existing therapeutic agents.

[0045] The term "administration" in this invention means providing a specific substance to a patient by any appropriate method, and the route of administration of the composition of this invention may be administered through any general route as long as it can reach the target tissue. It may be administered intraperitoneally, intravenously, intramuscularly, subcutaneously, intradermally, orally, locally, intranasally, intrapulmonaryly, or rectally, but is not limited thereto. Additionally, the pharmaceutical composition of this invention may be administered by any device capable of delivering the active substance to target cells. Preferred modes of administration and formulations include intravenous injections, subcutaneous injections, intradermal injections, intramuscular injections, drip infusions, etc. Injectable preparations can be manufactured using aqueous solvents such as physiological saline solution and Ringer's solution, vegetable oils, higher fatty acid esters (e.g., ethyl oleate), alcohols (e.g., ethanol, benzyl alcohol, propylene glycol, glycerin), etc., and may include pharmaceutical carriers such as stabilizers to prevent deterioration (e.g., ascorbic acid, sodium bisulfite, sodium pyrosulfite, BHA, tocopherol, EDTA, etc.), emulsifiers, buffers to adjust pH, and preservatives to inhibit microbial growth (e.g., phenylmercury nitrate, thimerosal, benzalkonium chloride, phenol, cresol, benzyl alcohol, etc.).

[0046] In another aspect, the present invention provides a food composition for inhibiting cancer metastasis and invasion or for improving cancer, comprising a pericodeptide compound of Formula 2 or a food-acceptable salt thereof as an active ingredient.

[0047] Specific details regarding the structure, efficacy, uses, formulations, etc., of the above-mentioned pericodeptide compound, as well as specific details regarding cancer, are as described above and are therefore omitted below.

[0048] The food-grade acceptable salt of the present invention is preferably an acid addition salt formed by a food-grade acceptable free acid or a metal salt formed by a base. As an example, inorganic acids and organic acids may be used as free acids. Inorganic acids may include hydrochloric acid, sulfuric acid, bromic acid, sulfite, or phosphoric acid, and organic acids may include citric acid, acetic acid, maleic acid, fumaric acid, gluconic acid, methanesulfonic acid, etc. Additionally, alkali metal salts or alkaline earth metal salts, sodium, potassium, or calcium salts may be used as metal salts. However, they are not necessarily limited thereto.

[0049] The food composition of the present invention may be used by adding one or more components selected from the group consisting of the pericodepside compound and its food-acceptable salts as is, or together with other foods or food components, and may be appropriately used according to conventional methods.

[0050] The amount of the above ingredients can be appropriately determined according to the purpose of use (prevention, health, or therapeutic treatment). Generally, when manufacturing food or beverages, the above ingredients of the present invention are added in an amount of 10% by weight or less, preferably 5% by weight or less, relative to the raw materials. However, in the case of long-term consumption for the purpose of health and health control, the amount may be less than the above range, and since there are no issues regarding safety, the active ingredients may be used in an amount greater than the above range.

[0051] The food composition according to the present invention may include, in addition to one or more components selected from the group consisting of pericodecid compounds or food-grade acceptable salts thereof, components that are typically added during food production. For example, it includes proteins, carbohydrates, fats, nutrients, seasonings, and flavoring agents. The carbohydrates mentioned above are monosaccharides such as glucose and fructose, disaccharides such as maltose, sucrose, and oligosaccharides, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. The flavoring agents may include natural flavoring agents (thaumatin, stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.)) and synthetic flavoring agents (saccharin, aspartame, etc.).

[0052] There are no specific restrictions on the types of the above-mentioned foods. Examples of foods to which the above-mentioned ingredients may be added include meat, sausage, bread, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, chewing gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes, and include all health foods in the conventional sense.

[0053] When the food of the present invention is a beverage, it may contain various flavoring agents or natural carbohydrates as additional ingredients, as in conventional beverages. The above natural carbohydrates may include monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, natural sweeteners such as dextrin and cyclodextrin, or synthetic sweeteners such as saccharin and aspartame.

[0054] In addition to the ingredients described above, the above food may contain various nutritional supplements, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. Furthermore, the health food of the present invention may contain fruit pulp for the production of natural fruit juices, beverages, and vegetable beverages. These ingredients may be used independently or in combination. Effects of the invention

[0055] The pericodecid compound of Formula 2 of the present invention has cytotoxicity against cancer cells and downregulates the expression of β-catenin, phosphorylated STAT, and NF-к in cancer cells, thereby inhibiting the expression of N-cadherin, an EMT effector, and its transcription factors Snail, Slug, Twist, and ZEB1 / 2, and inhibits the migration and invasion of cancer cells; thus, it can be usefully used as an active ingredient in a composition for inhibiting cancer metastasis and invasion and for treating or improving cancer. Brief explanation of the drawing

[0056] Figure 1 is a figure showing the HPLC chromatogram of pericodecid denoted as compound 2. Figure 2 shows the HRESIMS spectrum of pericodecid denoted as compound 2. FIG. 3 is for pericodecid (2). 1 H- 1 This is a figure showing H correlation spectroscopy and major heteronuclear multiple bond correlations. Figure 4 shows the effects of proatranorin III (1) and pericodecside (2) on the viability of various cancer cell lines. (a) Results of measuring the relative viability of A549, U87, Caco-2, MCF-7, AGS, and PC3 cells treated with proatranorin III (1) at 3.13-100 μg / mL for 48 hours using an MTT assay. (b) Results of measuring the relative viability of A549, U87, Caco-2, MCF-7, AGS, and PC3 cells treated with pericodecside (2) at 3.13-100 μg / mL using an MTT assay. Data represent mean ± standard deviation. n=3. *p < 0.05; **p < 0.01***; p < 0.001; NS, no significant difference compared to cells treated with DMSO in each group. Figure 5 illustrates the effects of proatranorin III (1) and pericodecside (2) on cell invasion and proliferation in various cancer cell lines. (a) Representative images of A549, U87, Caco-2, MCF-7, AGS, and PC3 cells invading through a 1% gelatin-coated transwell after treatment with non-toxic concentrations (5 μg / mL) of proatranorin III (1) and pericodecside (2) for 24 hours. Scale bar = 100 μm. (b) Quantitative analysis results of the number of invading cells in each group. (c) Representative images of a clonogenic assay in various cancer cell lines treated with 5 μg / mL of proatranorin III (1) and pericodecside (2), respectively, for 14 days. Representative images of three independent experiments are shown (n=3). Scale bar = 35 mm. (d) Quantitative analysis of colony areas in various cancer cell lines treated with proatranorin III (1) and pericodepsid (2) at 5 μg / mL each for 14 days. Representative images of three independent experiments are shown (n=3). Scale bar = 35 mm. Data represent mean ± standard deviation. *p < 0.05; **p < 0.01; NS, no significant difference compared to DMSO-treated cells in each group. Figure 6 illustrates the effect of pericodecside (2) on the protein and mRNA expression of EMT markers. (a) U87, MCF-7, and PC3 cells were treated with pericodecside (2) at concentrations of 5 μg / mL or 1 μg / mL, cultured for 24 hours, then lysed and total protein analyzed by immunoblot. Representative images and quantitative analysis of the relative protein levels of N-cadherin and Twist in cells treated with compound 2 show that compound 2 reduces protein levels in a concentration-dependent manner. β-actin was used as a loading control. (b) Relative mRNA levels of EMT markers (E-cadherin, N-cadherin) and their regulators (Snail, Slug, Twist, ZEB1, and ZEB2) were analyzed by qRT-PCR. Data are expressed as mean ± standard deviation (n=3 for each group). *p < 0.05; **p < 0.01; ***p < 0.001; NS: No significant difference compared to DMSO-treated cells. Figure 7 shows the effect of pericodecside (2) on the expression of transcription factors associated with the Wnt, AP-1, STAT, and NF-κB signaling pathways. (a) HEK293T cells were co-transfected with the pRL-TK (Renilla) plasmid and the pTOPFLASH-luc / β-catenin-luc, pAP-1-luc / Kitenin-luc, pSTAT-luc, or pNF-κB-luc reporter plasmids (firefly). After 12 hours, the transfected cells were treated with 5 μg / mL or 1 μg / mL of pericodecside (2) for an additional 48 hours. Relative luciferase activity was then quantified. (b) U87, MCF-7, and PC3 cells treated with 5 μg / mL or 1 μg / mL of pericodecside (2) for 24 hours were lysed, and total protein was immunoblotted with the labeled antibody. (c) Relative protein levels of β-catenin, cyclin-D1, p-STAT, and p-NF-κB were quantified in cells treated with Compound 2, demonstrating that Compound 2 reduces protein levels in a concentration-dependent manner. β-actin was used as a loading control. Relative protein levels of phospho-STAT and phospho-NF-κB were compared to the total expression of STAT and NF-κB. Data are presented as mean ± standard deviation (n=3 for each group). *p < 0.05; **p < 0.01; ***p < 0.001; NS: No significant difference compared to DMSO-treated cells. FIG. 8 is a schematic diagram illustrating the mechanism of action of pericodeptide according to the present invention. Pericodeptide (2) inhibits the activation of Wnt, STAT, and NF-кB, thereby inhibiting N-cadherin, an EMT effector, and its transcription factors Snail, Slug, Twist, and ZEB1 / 2, thereby inhibiting cancer cell invasion and proliferation. Specific details for implementing the invention

[0057] Hereinafter, the present invention will be described in detail with reference to examples and the like to aid in understanding the invention. However, the embodiments according to the present invention may be modified in various different forms, and the scope of the present invention should not be interpreted as being limited to the following embodiments. The embodiments of the present invention are provided to more completely explain the invention to those with average knowledge in the art.

[0059] Experimental Materials and Statistical Analysis

[0060] The strain used in the following experiments is a genetically modified strain, similar to the one described in a previous paper (Non-patent Literature 9) published by some of the inventors. Ascochyta rabiei It is a strain. This modified strain is Stereocaulon alpinum Replicated from the atranorin biosynthetic gene cluster isolated from (GenBank PRJNA693574) atr1 class atr2 Two biosynthetic genes of lost the ability to produce solanapyrones A. rabiei It is a strain introduced into the strain. Ascochyta rabiei The engineered strain (Sta04643-pII95-T2) is preserved at the Korean Lichen and United Biological Resource Center at Suncheon National University in Korea.

[0061] All experiments were performed three times. All statistical analyses were performed using IBM SPSS (IBM Statistical Package for Social Science) version 22 and SigmaPlot 12.5. Data were expressed as mean ± standard deviation. The Student t-test was used to determine significant differences between groups, and a p-value < 0.05 was considered significant.

[0063] Example 1: Extraction and Purification

[0064] To perform structural identification and biological evaluation, atr1 class atr2 Genetically modified expressing A. rabiei Compound 1 represented by Chemical Formula 1 and Compound 2 represented by Chemical Formula 2 were purified from the culture extract of the strain.

[0065] [Chemical Formula 1]

[0066]

[0067] [Chemical Formula 2]

[0068]

[0069] In a previous study (Non-patent Literature 9), the inventors discovered that the major product synthesized by this strain was Compound 1. Subsequent experiments confirmed the production of Compound 2 by the same strain, albeit in small quantities. To obtain a sufficient amount of Compound 2, large-scale culture of the strain was performed. The strain was first grown on PDA medium at 20°C for 3 days. Subsequently, agar was cut into small blocks (0.5 cm in diameter), inoculated directly onto freshly prepared PDA, and cultured at room temperature for 3 weeks. Four 0.5 L bottles containing PDA culture medium were autoclaved, and 20 PDA plates were prepared per 0.5 L batch, resulting in a total of 80 plates. The strain was inoculated onto the entire medium of all 80 PDA culture plates. After 3 weeks of culture, the 80 PDA culture plates were harvested, immersed in ethyl acetate, and extracted by ultrasonic treatment at least three times. The ethyl acetate was filtered and stored for the subsequent preparation of a series of crude extracts.

[0070] Culture extracts are evaporated using a rotary evaporator (Rotavapor ® It was evaporated and dried in ) and reconstituted with methanol, after which it was analyzed using high-performance liquid chromatography (HPLC). Separation was performed in a semi-preparative HPLC using a Kromasil C18 column (250×10 mm, 5 μm, 10 nm; 40 °C) at a flow rate of 2.1 mL / min and UV monitoring at 254 nm. Compound 1 was purified using 80% acetonitrile (t R = 9.60 min; 18.2 mg), Compound 2 was purified using 55% acetonitrile solvent (t R = 16.27 min; 9.4 mg). 0.1% trifluoroacetic acid was added to the solvent used. The purity of Compound 2 was less than 90%. Subsequently, a second separation process was performed using 50% methanol buffered with 0.1% trifluoroacetic acid as the mobile phase. This separation was carried out on a YMC-Pack ODS-A column (150×4.6 mm, 5 μm, 12 nm) at 40°C at a flow rate of 1 mL / min, yielding 7.2 mg of Compound 2 (t R = retention time of 18.90 min). Purified Compound 2 was analyzed by NMR spectroscopic analysis. 1D and 2D NMR spectra for Compound 2 in CD3OD were acquired using a Bruker Avance III HD500 spectrometer (Bruker, Billerica, MA, USA). In addition, DMSO- using a JEOL 400 MHz NMR spectrometer (JEOL, Ltd., Tokyo, Japan) d 6 1D and 2D NMR spectra of Compound 2 were acquired. HPLC chromatography was performed using a Waters VION IMS QTOF mass spectrometer (Waters MS Technologies, Manchester, UK). HPLC chromatography (Fig. 1) showed the purity of Compound 2. Compound 2 had a purity exceeding 90%, making it suitable for biological activity.

[0071] Proatranorin III (1): white solid; 1 ¹H NMR (400 MHz, CDCl3): δ H12.54 (s, OH), 12.45 (s, OH), 11.77 (s, OH), 10.35 (s, CHO), 6.54 (s, 1H), 6.40 (s, 1H), 2.68 (s, 3H), 2.59 (s, 3H), 2.08 (s, 3H). HRESIMS m / z 359.0760 [MH] - (C 18 H 15 Calculated for O8, 359.0767); MS / MS spectrum is stored in the GNPS spectrum library.

[0072] Pericodeptide (2): light purple powder; UV (MeOH) λ max (log ε ) 253 (4.57), 323 (0.48); 1 ¹H NMR (500 MHz, CD3OD), 13 13C NMR (125 MHz, CD3OD) data, 1 ¹H NMR (400 MHz, DMSO- d 6 ), and 13 13C NMR (100 MHz, DMSO- d 6 ) data, see Table 1; HRESIMS m / z 545.1292 [MH] - (C 26 H 25 O 13 Calculated for, 545.1295). MS / MS spectra are stored in the GNPS spectrum library.

[0073] Example 2: Identification of Compound 2

[0074] Compound 2 in the High Resolution Mass Spectrometry (HRESIMS) spectrum m / z 545.1292 [MH] - (C 26 H 25 O 13Calculated by. Since it showed a deprotonated molecular peak at 545.1295) (Fig. 2), the molecular formula of Compound 2 is C with a degree of unsaturation of 14. 26 H 26 O 13 It was determined to be. This molecular formula 13 It was supported by 26 resonances indicated in the C NMR spectrum.

[0075] Compound 2 1 H and 13 The 1C NMR spectrum was similar to that of Compound 1, except for a few additional signals. Compound 2's 13 C-7'' in the C NMR spectrum δ C Additional carbonyl signals were observed at 166.9), and at C-2'', C-3'', C-4'', C-5'', and C-6'' (each δ H / δ C Five more methane signals were recorded at (with chemical shifts of 7.12 / 147.4, 4.43 / 69.1, 3.74 / 72.4, 4.15 / 71.7, and 5.99 / 71.9). In addition, δ C C-8'' with sp2 hybridized interquartile carbon signal at 126.8(C-1'')( δ H / δ C A methylation signal was observed at 3.76 / 52.5) ​​(Table 1).

[0076] 1 H- 1The spin system of H-2''-H-3''-H-4''-H-5''-H-6'' was confirmed through COSY data. Chemical shifts of C-1'' and C-2'' suggested that they are part of the olefin group, which was confirmed by the heteronuclear multiple bond correlation (HMBC) from H-2'' to C-1''. Chemical shifts of H-3'', H-4'', H-5'', and H-6'' implied that they are oxygenated, which was supported by the inferred molecular formula. The HMBCs from H-6'' to C-1'' and from H-6'' to C-7'' suggested that this ring system is a moiety with a planar structure similar to pericosines (Fig. 3). From H-6'' to C-7' ( δ C HMBC to 168.7) placed a pericosine-like moiety at C-7' via an ester bond. This result was supported by a downfield chemical shift at H-6''. HMBC from H-8'' to C-7'' suggested the presence of a methyl ester group at C-7''.

[0077] In conclusion, the planar structure of Compound 2 was determined to be the structure shown in Chemical Formula 2. Stereochemical identification of Compound 2 was difficult. According to previous reports, pericosine is Periconia byssoidesOUPS-N133 is a C7 cyclohexenoid metabolite of the fungus [Non-patent Literature 10 and 11], and in that report, the planar structures of pericosines B, C, and E were nearly identical to one of the hydroxylated cyclohexenyl moiety groups of Compound 2. Although the absolute and relative configurations of pericosines were characterized based on those determined by the overall synthesis, determining the configurations of pericosine analogs remained difficult due to the ambiguity of the cyclohexenyl ring conformation. Unfortunately, attempts to purify pericosines from the acid hydrolysate of Compound 2 were unsuccessful. The relative configurations were determined by comparing coupling constants with literature values ​​(3'' R* , 4'' R* , 5'' R* , 6'' S* It was tentatively identified as ), and this was identical to that of pericosin B. A. rabiei phylogenetically P. byssoides It is related to, and since both belong to the order Pleosporales, the discovery of pericodepside is A. rabiei This suggests the possibility of producing pericosine B. However, it is considered that additional experimental evidence is needed to confirm this configuration. Here, the inventors named compound 2 pericodepside.

[0078]

[0079] C represents the NMR solvent (CD3OD) used for HMBC data acquisition, D is the NMR solvent (DMSO-) used for HMBC data acquisition d It represents 6).

[0080] Example 3: Cell Culture

[0081] Human cancer cell lines A549 (lung carcinoma cells), U87 (glioblastoma cells), Caco-2 (colorectal adenocarcinoma cells), MCF-7 (breast cancer cells), AGS (gastric cancer cells), and PC3 (prostate adenocarcinoma cells) were cultured in DMED or RPMI medium (GenDepot, USA) supplemented with 10% fetal bovine serum (FBS; Gen Depot, USA) and 1% penicillin-streptomycin (Penicillin-Streptomycin; Gen Depot, USA) in a humidified environment containing 5% CO2 at 37°C.

[0082] Example 4: MTT Analysis

[0083] Cell viability was investigated using A549, U87, Caco-2, MCF-7, AGS, and PC3 cell lines based on the colorimetric quantification of the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Briefly, 2 × 10⁶ 4 Cells were seeded into 96-cell plates at a concentration of 3.3–100 μg / mL and cultured with compounds 1 and 2 for 48 hours. After incubation with MTT solution for 4 hours, the cells were lysed with DMSO. Absorbance values ​​at 540 nm were measured using a microplate reader and analyzed using Gen 5 (2.03.1) (BioTek, VT, USA). The results are shown in Table 2 and Figure 4.

[0084] The viability of A549, U87, MCF-7, and AGS cells was unaffected when treated with Compound 1 at concentrations below 100 μg / mL, but cell viability significantly decreased when treated with Compound 1 at concentrations above 100 μg / mL. Additionally, the viability of Caco-2 and PC3 cells was unaffected when treated with Compound 1 at concentrations below 25 μg / mL, but decreased when treated with Compound 1 at concentrations between 25 and 100 μg / mL (Fig. 4a). Treatment with Compound 2 at concentrations between 0.78 and 12.5 μg / mL did not affect the viability of any cell lines, but treatment with Compound 2 at concentrations between 12.5 and 100 μg / mL significantly reduced the viability of all cell lines in a concentration-dependent manner (Fig. 4b). Therefore, these results indicate that Compound 2 possesses stronger biological activity than Compound 1 in reducing cancer cell viability. IC50 of Compounds 1 and 2 50 The values ​​were 81–300 μg / mL and 27–120 μg / mL for the tested cell lines, respectively, indicating no cytotoxicity (IC). 50 It is considered a cytotoxic compound only when the value is < 10 μg / mL). In subsequent experiments, it was used at a non-toxic concentration (5 μg / mL).

[0085] Tissue Source Cell lines IC for Compound 1 50 (μg / mL) IC for Compound 2 50 (μg / mL) Human lung carcinoma cell A549 109.05±2.04 55.59±1.75 Human glioblastoma cell U87 303.64±2.49 120.42±2.08 Human colorectal adenocarcinoma cell Caco-2 119.85±2.08 48.81±1.69 Human breast cancer cell MCF-7 262.40±2.42 75.15±1.88 Human gastric cancer cell AGS 99.04±1.99 37.69±1.58 Human prostatic adenocarcinoma PC3 81.37±1.91 27.73±1.44

[0086] Example 5: Analysis of cell invasiveness

[0087] 5-1. Invasion assay

[0088] The transwell invasion assay was performed using an 8 μm pore polycarbonate membrane transwell chamber (Corning, New York, USA) pre-coated with 1% gelatin. Briefly, A549, U87, Caco-2, MCF-7, AGS, and PC3 cells were 1–2 × 10⁶ in DMEM or RPMI medium containing 0.2% bovine serum albumin (BSA). 6Cells were plated in the upper compartment of the chamber at cells / well. In the lower chamber, culture medium containing 10 μg / mL of fibronectin was placed as a chemoattractant. After 12 hours of incubation, the cells were treated with DMSO (0.01%), 5 μg / mL of Compound 1, or 5 μg / mL of Compound 2. After culturing for 24 hours, the cells in the upper chamber were fixed using a Diff Quik kit (Sysmex, Kobe, Japan) and counted using an optical microscope (5 fields per chamber).

[0089] 5-2. Clonogenic Assay

[0090] A549, U87, Caco-2, MCF-7, AGS, and PC3 cells in 2 mL of DMEM or RPMI medium at a volume of 0.5–2 × 10⁶ 3 Cells were inoculated at a density of cells / well and treated with 0.01% DMSO and 5 μg / mL of Compound 1 or 2 3 hours later. After incubation at 37°C for 7–14 days, cells were stained with 0.01% crystal violet. Pixel intensity of colony areas was measured across the entire microscope field of each plate using IMT iSolution FL software (IMT i-Solution Inc., Northampton, NJ, USA). Data were presented as the average of three experiments.

[0091] 5-3. Analysis Results

[0092] Cell invasiveness plays a significant role in metastatic disease and is a major cause of death in cancer patients. To determine the effects of non-toxic concentrations of compounds 1 and 2 on cell motility and proliferation, invasiveness and clonogenicity assays were performed on cell lines treated with 5 μg / mL of compound 1 or 2. Treatment with compound 1 did not inhibit cell invasion or reduce colony numbers in any cell line except for MCF-7 cells. However, imaging and quantitative analysis revealed that treatment with compound 2 significantly reduced the invasiveness of A549, U87, MCF-7, and PC3 cells by approximately 12%, 38%, 31%, and 38%, respectively (Figs. 5a and 5b), and reduced the colony numbers of U87, MCF-7, and PC3 cells by approximately 12%, 27%, and 32%, respectively (Figs. 5c and 5d). These results indicate that treatment with 5 μg / mL of compound 2 inhibits both cell invasion and proliferation in U87, MCF-7, and PC3 cells.

[0093] Example 6: Investigation of protein and mRNA expression levels of EMT markers

[0094] EMT facilitates the invasion and metastasis of cancer cells and plays an important role in tumor progression. E-cadherin and N-cadherin, as well as transcription factors Snail, Twist, and ZEB1 / 2, are known as EMT markers. Since Compound 2 significantly inhibited cell invasion and proliferation in U87, MCF-7, and PC3 cells, the inventors investigated the protein and mRNA expression levels of EMT markers to determine how Compound 2 affects cell motility.

[0095] 6-1. Western Blot

[0096] Specifically, U87, MCF-7, and PC3 cells were treated with 5 μg / mL of Compound 1 or 2 for 24 hours. 20 μg of extracted protein was separated by 8–10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis. EMT markers were detected using antibodies against N-cadherin (Cell Signaling Technology) and the Twist (Abcam) primary antibody. Various target molecules were detected using primary antibodies against anti-β-catenin (Cell Signaling Technology), anti-cyclin-D1 (Calbiochem), phospho-STAT, and phospho-NF-κB (Cell Signaling Technology). The β-Actin (Santa Cruz Biotechnology) primary antibody was used as an internal standard. Bands were measured for each sample using a Multi Gauge 3.0. Values ​​were expressed in densitometer units corresponding to signal intensity.

[0097] 6-2. Quantitative Real-Time PCR (qRT-PCR).

[0098] Total RNA was isolated from U87, MCF-7, and PC3 cells after treatment with 5 μg / mL of Compound 1 or 2 in DMSO (0.01%) for 24 hours using RNAiso Plus (TaKaRa, Otsu, Japan) according to the manufacturer's instructions. Total RNA (1 μg) from each treated cell group was converted to cDNA using the M-MLV Reverse Transcriptase Kit (Invitrogen, Carlsbad, USA) and SYBR green (En-zynomics, Seoul, Korea). The forward and reverse sequences used for qRT-PCR are shown in Table 3. qRT-PCR reactions and analysis were performed using CFX (Bio-Rad, Hercules, USA).

[0099] Target marker order E-cadherin Forward 5'-cagaaagttttccaccaaag-3' Reverse 5'-aaatgtgagcaattctgctt-3' N-cadherin Forward 5'-ctcctat gagtggaacagggaacg-3' Reverse 5'-ttggatcaatgtcataatcaagtgctgta-3' Snail Forward 5'-tcccgggcaatttaacaatg-3' Reverse 5'-tgggagacacatcggtcga-3' Slug Forward 5'-cgaactggacacacatacagtg-3' Reverse 5'-ctgaggatctctggttgtggt-3' Twist Forward 5'-cgggagtccgcagtctta-3' Reverse 5'-tgaatcttgctcagcttgtc-3' ZEB1 Forward 5'-atgacacaggaaaggaagg-3' Reverse 5'-agcagtgtcttgttgtag-3' ZEB2 Forward 5'-caagaggcgcaaacaagcc-3' Reverse 5'-ggttggcaataccgtcatcc-3' β-Actin Forward 5'-attgtgaactttgggggatg-3' Reverse 5'-gatgagattggcatggcttt-3'

[0100] 6-3. Analysis Results

[0101] Treatment with 5 μg / mL or 1 μg / mL of compound 2 significantly reduced the mRNA expression of N-cadherin, Snail, Slug, Twist, and ZEB1 / 2 in PC3 cells, in addition to the protein expression of N-cadherin and Twist. Similarly, compound 2 inhibited the protein and mRNA expression of EMT markers in U87 and MCF-7 cells, specifically inhibiting N-cadherin expression in U87 cells and Twist expression in MCF-7 cells in a concentration-dependent manner (Figures 6a and 6b). These results suggest that compound 2 regulates the expression of the EMT effector N-cadherin by inhibiting the transcription factors of Snail, Slug, Twist, and ZEB1 / 2, thereby inhibiting motility in U87, MCF-7, and PC3 cells.

[0102] Example 7: Reporter Assay

[0103] To identify the signaling pathways involved in the inhibition of EMT by compound 2, reporter analysis was performed using HEK293T cells transfected with a vector containing TOPFLASH, AP-1, STAT, or NF-κ promoter.

[0104] HEK293T cells were transfected with TOPFLASH-luc / β-catenin-luc, AP-1-luc / Kitenin-luc, STAT-luc, and NF-κB-luc-conjugated firefly plasmids along with the Renilla-luc (pRL-TK) plasmid using GENE 9 DNA transfection reagent (Roche, Werk Penzberg, Germany). After 12 hours of transfection, cells were treated with 5 μg / mL of compound 1 or 2 in DMSO (0.01%) and cultured for 48 hours. Luciferase activity was measured using a Dual-Luciferase reporter assay kit (Promega, Madison, WI, USA), and transfection efficiency was controlled by normalizing firefly activity to Renilla luciferase activity.

[0105] As shown in Figure 7a, Compound 2 at 5 μg / mL significantly inhibited β-catenin-mediated TOPFLASH activity. Compound 2 reduced STAT and NF-κB transcriptional activity in a concentration-dependent manner. However, chitenin-mediated AP-1 activity by Compound 2 was unaffected. To investigate whether Compound 2 affects the activation of signaling pathways such as Wnt, STAT, and NF-κB, their target molecules were measured by Western blot in U87, MCF-7, and PC3 cells. Compound 2 reduced the protein levels of β-catenin, a target of Wnt signaling, and its downstream target, cyclin-D1. Additionally, Compound 2 significantly reduced the protein levels of phosphorylated STAT and NF-κB in MCF-7 and PC3 cells in a concentration-dependent manner, but did not affect total protein levels. In addition, Compound 2 reduced phosphorylated STAT levels in U87 cells and significantly reduced phosphorylated NF-κB as well as total NF-κB levels (Fig. 7b). Quantitative analysis showed that the levels of these target proteins were more robustly inhibited in PC3 cells than in other cell lines (Fig. 7c). These findings demonstrate that Compound 2 inhibits the invasion and proliferation of U87, MCF-7, and PC3 cells by inhibiting the activation of Wnt, STAT, and NF-κB signaling, suggesting that Compound 2 has potential for the treatment of brain tumors, breast cancer, and prostate cancer.

Claims

Claim 1 delete Claim 2 A pharmaceutical composition for inhibiting cancer metastasis and invasion comprising a pericodepsid compound or a pharmaceutically acceptable salt thereof. Claim 3 A pharmaceutical composition for inhibiting cancer metastasis and invasion according to claim 2, characterized in that the cancer is one or more cancers selected from the group consisting of liver cancer, lung cancer, stomach cancer, pancreatic cancer, colorectal cancer, cervical cancer, breast cancer, prostate cancer, ovarian cancer, brain cancer, osteosarcoma, bladder cancer, head and neck cancer, kidney cancer, melanoma, leukemia, and lymphoma. Claim 4 A pharmaceutical composition for treating cancer comprising a pericodepsid compound or a pharmaceutically acceptable salt thereof. Claim 5 A pharmaceutical composition for treating cancer according to claim 4, characterized in that the cancer is one or more cancers selected from the group consisting of liver cancer, lung cancer, stomach cancer, pancreatic cancer, colorectal cancer, cervical cancer, breast cancer, prostate cancer, ovarian cancer, brain cancer, osteosarcoma, bladder cancer, head and neck cancer, kidney cancer, melanoma, leukemia, and lymphoma. Claim 6 delete Claim 7 delete Claim 8 A food composition for improving cancer comprising a pericodeptide compound or a salt thereof. Claim 9 A food composition for improving cancer, characterized in that, in claim 8, the cancer is one or more cancers selected from the group consisting of liver cancer, lung cancer, stomach cancer, pancreatic cancer, colorectal cancer, cervical cancer, breast cancer, prostate cancer, ovarian cancer, brain cancer, osteosarcoma, bladder cancer, head and neck cancer, kidney cancer, melanoma, leukemia, and lymphoma.

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