KR101646002B1 - Pharmaceutical composition for prevention or treatment of colorectal cancer comprising ethylacetate fraction of jubak ethanol extract as an effective component and health functional food comprising the same - Google Patents

Abstract

The present invention relates to a pharmaceutical composition and a health functional food for the prevention or treatment of colorectal cancer containing an extract of Aspergillus oryzae as an active ingredient and more particularly to a pharmaceutical composition and a health functional food for preventing or treating colorectal cancer, The present invention relates to a pharmaceutical composition and a health functional food for preventing or treating colorectal cancer contained as an active ingredient. The pharmaceutical composition of the present invention and the ethyl acetate fraction of the alcoholic ethanol extract as an active ingredient of the health functional food inhibit the proliferation of the colon cancer cell line and inhibit the cancer suppressor genes NAG-1, ATF3, DDIT3 and / or p21 The above-mentioned effective ingredient can be applied to medicines and foods for prevention, improvement and treatment of colon cancer. In addition, the present invention may be usefully applied to research for the development of a therapeutic agent for colorectal cancer by providing a method of treating the colon cancer cells with the ethyl acetate fraction of the ethanol extract of the present invention to inhibit the proliferation of colon cancer cells .

Description

TECHNICAL FIELD The present invention relates to a pharmaceutical composition for the prevention or treatment of colorectal cancer, which comprises an ethyl acetate fraction of an ethanol extract of Alaska pollack as an active ingredient, and a health functional food. FUNCTIONAL FOOD COMPRISING THE SAME}

The present invention relates to a pharmaceutical composition and a health functional food for the prevention or treatment of colorectal cancer containing an extract of Aspergillus oryzae as an active ingredient and more particularly to a pharmaceutical composition and a health functional food for preventing or treating colorectal cancer, The present invention relates to a pharmaceutical composition and a health functional food for preventing or treating colorectal cancer contained as an active ingredient.

Makgeolli is made from starch, such as glutinous rice, and it is a unique Korean traditional liquor which is made by fermenting Takju mixed with fermentation agent. Makgeolli has a combination of two physico - chemical and microbiological changes, glycosylation and fermentation, which can not be seen in other liquors. When making makgeolli, the residues produced in the process of squeezing out the alcohol are called " Jukbu " or "Suljijimi", and the name "Makgeolli Lee , Ji-suk, is a major component of protein and starch, and contains alcohol, saccharides, yeast, organic acids and minerals. Respectively.

Cancer is a disease in which abnormal cells are uncontrolled and overproduced by proliferation. In Korea, according to the cancer registration statistics of the central registration office of the Ministry of Health and Welfare, the cancer occurrence status as of 2011 is the third (12.8%), followed by thyroid cancer (17.8%) and stomach cancer (14.9%), (19.6 percent), colon cancer (15.2 percent), lung cancer (14.2 percent), and liver cancer (11.5 percent). In particular, colon cancer is the second most common cancer in the world, and the incidence is quite high in Korea. The cause of colon cancer is the genetic factors such as excessive animal fat intake, fibrosis, calcium deficiency, inflammatory diseases and colon polyps due to the western type dietary change due to modernization. However, most of the anticancer drugs used as therapeutic agents induce apoptosis of cancer cells [6] as well as normal cell death. Therefore, in recent years, researches are being made on anticancer drugs that induce immune enhancement and cancer cell death using natural materials extracted from plants. In addition, patients with family history are encouraged to perform early detection of colon cancer.

DISCLOSURE OF THE INVENTION The present invention has been made in order to recover the useful physiologically active substance from the fermented bean sprout, to increase the utility of the fermented bee as a biomass, and to overcome the risk of side effects of existing anticancer drugs. A pharmaceutical composition for preventing or treating colorectal cancer containing an ethyl acetate fraction of an alcoholic extract of alcoholic beverages and an effective ingredient, and a health functional food containing the fraction. The present invention also provides a methodology for the development of a therapeutic agent for colorectal cancer by providing a method of treating the colon cancer cells in an in vitro environment with the ethacetate fraction of the ethanol extract of Zucchini ethanol of the present invention to inhibit the proliferation of the colon cancer cells I want to.

In order to solve the above-mentioned problems, the present invention provides a pharmaceutical composition for preventing or treating colon cancer containing an ethyl acetate fraction of an ethanol extract of Geranium japonica as an active ingredient.

The composition of the present invention can be used for the treatment of cancer such as NAG-1, ATF3, DNA damage-inducible transcript 3 (DDIT3) and / or cyclin-dependent kinase inhibitor 1A) gene expression.

The expression of p21 may be p53-dependent.

The present invention also provides a health functional food for preventing or ameliorating colorectal cancer containing an ethyl acetate fraction of an ethanol extract of Geranium japonica as an active ingredient.

Furthermore, the present invention in vitro (ex The present invention also provides a method for inhibiting the proliferation of colon cancer cells, which comprises treating the colorectal cancer cells with the ethyl acetate fraction of the ethanol extract of Guacause under vivo conditions.

The in vitro conditions in vitro (in in vitro conditions.

The treatment is preferably in a concentration range of 0.25 to 1 mg / ml.

Expression of NAG-1, ATF3, DDIT3 and / or p21 gene can be increased by the above treatment.

The pharmaceutical composition of the present invention and the ethyl acetate fraction of the alcoholic ethanol extract as an active ingredient of the health functional food inhibit the proliferation of the colon cancer cell line and inhibit the cancer suppressor genes NAG-1, ATF3, DDIT3 and / or p21 The above-mentioned effective ingredient can be applied to medicines and foods for prevention, improvement and treatment of colon cancer. In addition, the present invention can be usefully applied to research for the development of a therapeutic agent for colorectal cancer by providing a method of treating the colon cancer cells with the ethyl acetate fraction of the ethanol extract of the present invention to inhibit the proliferation of colon cancer cells.

(KSD-W6-5), an ethanol extract of KSD-E1-1 (KSD-W6-5) and a butanol fraction (KSD-W6-4) (KSD-E1-2), ethyl acetate fraction (KSD-E1-3), butanol fraction (KSD-E1-4) and water residue (KSD-E1- 5) treatment (2.5 mg / ml) in the human HCT116 cell line.
FIG. 2 is a graph showing the degree of inhibition of the survival rate of the human HCT116 cell line by the treatment with the ethyl acetate fraction (KSD-E1-3) of the ethanol extract of Vermicelli japonica.
FIG. 3 shows RT-PCR results showing the expression patterns of NAG-1, ATF3, DDIT3 and p21 genes on human HCT116 cell line by treatment with ethyl acetate fraction (KSD-E1-3) (1 mg / ml) .
4 shows the results of confirming whether p53-dependent increase in NAG-1, ATF3, DDIT3 and p21 gene expression on human HCT116 cell line by treatment with ethyl acetate fraction (KSD-E1-3) (1 mg / ml) to be.

Hereinafter, the present invention will be described in detail.

The inventors of the present invention have conducted experiments on various candidate groups in order to develop an anticancer substance of natural origin, and as a result, it has been confirmed that the ethyl acetate fraction of the ethanol extract of Jugaku inhibits the proliferation of human colon cancer cell line HCT116 cells in a concentration- , And the expression of NAG-1, ATF3, DDIT3 and p21 genes, which are cancer-related genes, is confirmed, and thus the ethyl acetate fraction of the above-mentioned extract of the alcoholic beverages can be applied as a pharmaceutical composition and a health functional food for the prevention or treatment of colon cancer Suggest a possibility.

Accordingly, the present invention provides a pharmaceutical composition for the prevention or treatment of colorectal cancer, which comprises an ethyl acetate fraction of an ethanol extract of Geranium japonica as an active ingredient.

The "bean sprout " of the present invention can be obtained from a brewing process such as sake, beer, sake, takju, or a fermentation product, but is not limited thereto. In addition, the present invention is not limited to the type of yeast used in the manufacture of brewing or fermentation stock, fermentation and aging conditions, and the like.

As a preferred embodiment, the ethyl acetate fraction of the water extract of the present invention has the effect of reducing the moisture content by pressing the collected water from the step of preparing the tabacum, extracting it with ethanol, filtering the extract with filter paper, Powder and sequentially fractionating the mixture with an organic solvent such as n-hexene, ethyl acetate and butanol to obtain an ethyl acetate fraction, which can be concentrated to a powder under reduced pressure.

According to the study results of the present inventors, the ethyl acetate fraction of the alcoholic extract of the present invention can inhibit the proliferation of human colon cancer cell line HCT116 cells in a concentration-dependent manner.

The inventors of the present invention also found that the ethyl acetate fraction of the alcoholic extract of the present invention increased the expression of the NAG-1, ATF3, DDIT3 and / or p21 gene shown in Table 1 below, which is a gene involved in cancer development in human colon cancer cell lines Respectively.

Furthermore, it was confirmed that the expression of p21 gene was increased by p53.

The effect of inhibiting the proliferation of the human colon cancer cell line and the expression of the related gene of the ethyl acetate fraction of the extract of the present invention of the present invention can be confirmed by the fact that the ethyl acetate fraction of the present invention ethanol extract is effective for prevention and treatment of colon cancer It is possible.

The pharmaceutical composition containing the ethyl acetate fraction of the present invention may be formulated into oral preparations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols according to conventional methods, Sterile injectable solutions, and the like, and may be administered by various routes including oral administration or intravenous, intraperitoneal, subcutaneous, rectal, topical administration, and the like.

Such pharmaceutical compositions may further comprise carriers, excipients or diluents, and examples of suitable carriers, excipients or diluents that may be included include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, But are not limited to, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, amorphous cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, And the like.

In addition, the pharmaceutical composition of the present invention may further include a filler, an anti-coagulant, a lubricant, a wetting agent, a flavoring agent, an emulsifying agent, an antiseptic, and the like.

In a preferred embodiment, the solid preparations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient, for example starch, calcium carbonate, Sucrose, lactose, gelatin and the like are mixed and formulated. In addition to simple excipients, lubricants such as magnesium stearate, talc, and the like may also be used.

Examples of the oral liquid preparation include suspensions, solutions, emulsions, syrups and the like. In addition to water and liquid paraffin which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, Perfumes, preservatives, and the like.

As a preferable specific example, the preparation for parenteral administration includes sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-drying agents, suppositories, and the like. Examples of the non-aqueous solvent and suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. Injectables may include conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers, preservatives, and the like.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level will depend on the type of disease, severity, The sensitivity to the drug, the time of administration, the route of administration and the rate of release, the duration of the treatment, factors including co-administered drugs, and other factors well known in the medical arts. The pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered singly or multiply. It is important to take into account all of the above factors and to administer the amount in which the maximum effect can be obtained in a minimal amount without side effects, which can be easily determined by those skilled in the art.

In a preferred embodiment of the present invention, the effective amount of the extract of the present invention in the pharmaceutical composition of the present invention may vary depending on the age, sex and body weight of the patient. Generally, 1 to 5,000 mg, preferably 100 to 3,000 mg per kg of body weight, Or every other day or one to three times a day. However, the dosage may not be limited in any way because it may be increased or decreased depending on route of administration, severity of disease, sex, weight, age, and the like.

The pharmaceutical composition of the present invention can be administered to a subject through various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections.

In the present invention, "administration" means providing a predetermined substance to a patient by any suitable method, and the administration route of the pharmaceutical composition of the present invention is either oral or non-oral May be administered orally. The composition of the present invention may also be administered using any device capable of delivering an effective ingredient to a target cell.

In the present invention, the term "object" includes, but is not limited to, human, monkey, cow, horse, sheep, pig, chicken, turkey, quail, cat, dog, mouse, rat, rabbit or guinea pig , Preferably a mammal, more preferably a human.

The ethyl acetate fraction of the above described water-soluble ethanol extract of the present invention of the present invention can be provided not only as a pharmaceutical composition but also as a health functional food for preventing or improving colon cancer.

Accordingly, the present invention provides a health functional food for preventing or ameliorating colorectal cancer, which comprises the ethyl acetate fraction of the present invention ethanol extract.

The food containing the ethyl acetate fraction of the water extract of the present invention may be, for example, various foods, beverages, gums, tea, vitamin complexes, health supplements and the like, and may be a powder, a granule, a tablet, Can be used.

The ethacetate fraction of the water extract of the present invention may be added in an amount of 0.01 to 15% by weight of the total food, and the health beverage composition may be added in a proportion of 0.02 to 10 g, preferably 0.3 to 1 g, have.

The health functional food of the present invention may contain, as an additional ingredient, a food-acceptable food-aid additive such as natural carbohydrates and various flavors, in addition to containing the above-mentioned compound as an essential ingredient in the indicated ratio.

Examples of the natural carbohydrate include sugar sugars such as glucose, monosaccharides such as fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol and erythritol. The ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the health functional food of the present invention.

Examples of the flavoring agents include natural flavoring agents such as tautatin, rebaudioside A and stiglycerin such as glycyrrhizin, and synthetic flavoring agents such as saccharin and aspartame.

In addition to the above, the health functional food of the present invention may contain various kinds of nutrients, vitamins, minerals, flavors such as synthetic flavors and natural flavors, colorants and heavy stabilizers, pectic acid and its salts, alginic acid and its salts, Thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated drinks, and the like.

In addition, the health functional food of the present invention may contain flesh for producing natural fruit juice, fruit juice drink, vegetable drink and the like. These components may be used independently or in combination. The ratio of such additives is generally selected in the range of 0.01 to 20 parts by weight per 100 parts by weight of the extract of the present invention.

The present invention also other (ex vivo The present invention also provides a method for inhibiting the proliferation of colon cancer cells, which comprises treating the colorectal cancer cells with the ethyl acetate fraction of the ethanol extract of Guacause under vivo conditions.

The in vitro conditions is meant the state in which there is a colon cancer cells are physically completely separated from a living body, in vitro (in in vitro conditions.

The colorectal cancer cells may be primary cells, subcultured cells, colon cancer cells, or the like extracted from a patient through surgical operation or the like.

The treatment can be carried out, for example, by adding the ethyl acetate fraction of the alcoholic ethanol extract to a medium containing colon cancer cells in a cell culture dish, and the treatment concentration is preferably 1 to 25 mg / ml.

When the ethyl acetate fraction of the ethanol extract of the present invention is treated with the colon cancer cells, the proliferation of colon cancer cells can be effectively inhibited, and the expression of NAG-1, ATF3, DDIT3 and / or p21 gene Can be effectively increased. In particular, p21 shows p53-dependent expression pattern.

As described above, the method of treating the colon cancer cells with the ethyl acetate fraction of the extract of the present invention from the extract of the present invention in vitro is a process for the development of a therapeutic agent for colon cancer using the ethyl acetate fraction of the extract of the present invention Can be used as an efficient methodology.

Hereinafter, the present invention will be described in more detail with reference to specific examples. The following examples are only a preferred embodiment of the present invention and are not to be construed as limiting the scope of the present invention.

[ Example ]

1. Materials and Methods

1.1. The  Extract preparation

The spruce used in the present invention was supplied and used by Kunsoo-dang Co., Ltd. The specific preparation method of the extract and its fractions are as follows. 1 kg of water was extracted with 1 L of water at 100 ° C. for 30 minutes, filtered (Whatman No. 2), and concentrated under reduced pressure at 70 ° C. to prepare powder. The ethanol extract was prepared by adding 6 L of 95% ethanol to 1.5 kg of the sample, extracting it twice at room temperature for 3 days, filtering it, and concentrating it under reduced pressure at 60 ° C. 4 g of each of the extracted extracts was suspended in water and sequentially fractionated using n-hexane, ethylacetate and butanol, and the water residue was recovered. The butanol fraction and the water The n-hexene fraction, ethyl acetate fraction, butanol fraction and water residue of the residue and the ethanol extract of the alcohol were obtained. The extracts and fractions were dissolved in DMSO (dimethyl sulfoxide) and stored at -20 ° C.

1.2. Cell culture and materials

The colon cancer cell line HCT116 was purchased from the American Type Culture Collection (ATCC, Frederick MD, USA) and the p53 null HCT116 (HCT116 p53 - / -) cell line was sold by Dr Bert Vogelstein from Johns Hopkins Medical School. Dulbecco's Modified Eagle Medium (DMEM, Gibco, USA) containing 10% FBS (Fetal Bovine Serum, Gibco, USA), 1% penicillin and streptomycin (WelGene, Korea) was used for both cell line cultures. The number of cells was maintained by subculturing while observing the degree of cell growth.

1.3. cell Growth rate  Measure( MTS assay )

Cell viability assay was performed to investigate the effect of hot - water extract, ethanol extract and fraction thereof on cell viability in colon cancer cell line HCT116. First, cells were inoculated in a 96-well plate at 1 × 10 ³ cells / well. After 24 hours, the medium was replaced with serum-free medium without FBS, and the extracts and fractions were dissolved in DMSO to give 2.5 mg / ml Respectively. After 24 hours, MTS (3- (4,5-dimethylthia-zol-2-yl) 5- (3-carboxy-methoxyphenyl) -2- (4-sulfophenyl) -2H-tetrazolium ) Solution (Promega, USA) was added at 20 μl per well, and incubated at 37 ° C in a 5% CO ₂ incubator for 4 hours. After the reaction, the absorbance was measured at 580 nm using NanoQuant Plate ^™ (Tecan Trading AG, Switzerland). Experimental results were obtained by analyzing the values in 4 independent wells using the Microsoft EXCEL program.

1.4. Total RNA  extraction

Total RNA extraction was performed according to the manufacturer's manual using a RNeasy mini kit (Qiagen, USA) from a colorectal cancer cell line treated with extracts and fractions. Finally, purified total RNA was quantified by measuring absorbance at 260 nm and 280 nm using NanoQuant Plate ^{™ and} then used for oligo DNA Microarray and RT-PCR.

1.5. Oligo DNA Microarray  Experiment

Oligo DNA microarray experiments and data analysis were performed by Genomictree, Inc, Korea. The microarray used was Agilent Human GE 4 X 44K (V2) arrays (Agilent Techonlogies, Palo Alto, USA) from Agilent.

1.6. Reverse transcription - PCR

Using ^{PrimeScript TM RT-PCR Kit (TaKaRa} , Japan) to 2μg total RNA extracted from the harvested cell line as template, cDNA was synthesized according to the manual of the manufacturer. The synthesized cDNA was used as a template and polymerase chain reaction (PCR) was performed using a gene-specific oligo primer (Table 2).

The final PCR product was electrophoresed on 1.5% agarose gel, stained with ethidium bromide (EtBr, Bioneer, Korea) and photographed using Gel Image Analysis System (CoreBio, Korea).

2. Results and discussion

2.1. The Heat number  Extracts and ethanol extracts and their The fraction  Effect on cell survival rate

(KSD-W6-1), its butanol fraction (KSD-W6-4) and water residue (KSD-W6-5) were investigated in order to investigate the effect of the extracts and fractions on the survival rate of colon cancer cell line HCT116. (KSD-E1-1), the n-hexane fraction (KSD-E1-2), the ethyl acetate fraction (KSD-E1-3), the butanol fraction (KSD-E1-4) and the water residue KSD-E1-5) at a concentration of 2.5 mg / ml and then subjected to a cell viability assay. DMSO was used as a control substance of the extracts and fractions.

As a result, it was confirmed that the cell viability was significantly reduced by the ethyl acetate fraction (KSD-E1-3) of the ethanol extract of Chenpai (FIG. 1). The ethylacetate fraction (KSD-E1-3) of the alcoholic extract of the present invention was used for further experiments.

2.2. Selected The  Ethyl acetate of ethanol extract Fraction  Measurement of cell viability by concentration

The effect of ethyl acetate fraction (KSD-E1-3) on the cell death of HCT116 cell line was investigated. The viability of the HCT116 cell line was determined by measuring the cell viability of the HCT116 cell line treated with 0.25, 0.5, and 1 mg / ml of the ethyl acetate fraction of the ethanol extract. As a result, it was confirmed that the ethylacetate fraction of the selected ethanol extract of Chenpai showed cell viability decreased in a concentration-dependent manner (FIG. 2).

2.3. Oligo DNA microarray Analysis of Gene Expression at Genome Level and Analysis of Anticancer-Related Gene Expression Using

(KSD-E1-3) of the ethanol extract of succinic acid selected to have the effect of reducing the cell survival of the colon cancer HCT116 cell line was examined in terms of gene expression level. The ethyl acetate fraction KSD-E1-3) was treated with HCT116 cell line and oligo DNA microarray analysis was performed. DNA microarray experiments, or 4 tumor suppressor gene which is a 2-fold increase over the expression of genes associated with cancer (Non-steroid Anti-inflammatory drug -activated Gene-1 [NAG -1], Activating Transcription Factor 3 [ATF3], DNA -Damage-Inducible Transcript3 [ DDIT3 ], and Cyclin-Dependent Kinase Inhibitor 1A [ CDKN1A , p21 ]) were selected and RT-PCR was performed (Table 2).

As a result jubak ethanol, ethyl acetate fraction (KSD-E1-3) gene by treatment NAG -1, ATF3, DDIT3, and p21 of the extract was found that both up-regulation (FIG. 3). These results were consistent with the oligo DNA microarray experiment results.

 2.4. p53 null HCT116  Using cell lines p53  Dependence gene validation

To confirm whether the expression of genes identified by Oligo DNA microarray experiments is dependent on p53 expression, HCT116 cell line with null p53 gene was used. As a result, the expression of p53 was increased by the ethyl acetate fraction (KSD-E1-3) of the p53 wild type cell line and the p53 null cell line was not expressed. The gene expression of four (NAG -1, ATF3, DDIT3, p21) who perform the RT-PCR using this cell line was confirmed. As a result, it was confirmed that the genes other than p21 were up-regulated regardless of the presence of p53 gene, whereas the expression of p21 gene was up-regulated only when p53 was present (Fig. 4) .

<110> PHARMACEUTICAL COMPOSITION COMPRISING THE LEES EXTRACT OF KOREAN RICE WINE AS AN EFFECTIVE COMPONENT FOR PRE OR OR TREATMENT OF COLORECTAL CANCER AND HEALTH FUNCTIONAL FOOD COMPRISING THE SAME          Kook Soon Dang Co., Ltd. <120> PHARMACEUTICAL COMPOSITION COMPRISING THE LEES EXTRACT OF KOREAN          RICE WINE AS AN EFFECTIVE COMPONENT FOR PREVENTION OR TREATMENT          OF COLORECTAL CANCER AND HEALTH FUNCTIONAL FOOD COMPRISING THE          SAME <130> PP14-266 <160> 12 <170> Kopatentin 2.0 <210> 1 <211> 20 <212> DNA <213> GAPDH forward primer <400> 1 ctgacctgcc gtctagaaaa 20 <210> 2 <211> 20 <212> DNA <213> GAPDH reverse primer <400> 2 gagcttgaca aagtggtcgt 20 <210> 3 <211> 20 <212> DNA <213> NAG-1 forward primer <400> 3 ctctcagatg ctcctggtgt 20 <210> 4 <211> 20 <212> DNA <213> NAG-1 reverse primer <400> 4 gaatcttccc agctctggtt 20 <210> 5 <211> 20 <212> DNA <213> ATF3 forward primer <400> 5 tggtgtttga ggattttgct 20 <210> 6 <211> 20 <212> DNA <213> ATF3 reverse primer <400> 6 atttctttct cgtcgcctgt 20 <210> 7 <211> 20 <212> DNA <213> DDIT3 forward primer <400> 7 cattgccttt ctccttcggg 20 <210> 8 <211> 20 <212> DNA <213> DDIT3 reverse primer <400> 8 tgctggttct ggctcctcct 20 <210> 9 <211> 20 <212> DNA <213> CDKN1A forward primer <400> 9 cgatggaact tcgactttgt 20 <210> 10 <211> 20 <212> DNA <213> CDKN1A reverse primer <400> 10 gtccacatgg tcttcctctg 20 <210> 11 <211> 20 <212> DNA <213> p53 forward primer <400> 11 ctcaccatca tcacactgga 20 <210> 12 <211> 20 <212> DNA <213> p53 reverse primer <400> 12 gagaggagct ggtgttgttg 20

Claims (8)

The ethyl acetate fraction of the ethanol extract of Lactobacillus japonica was treated with colon cancer cells under the ex vivo condition to prepare a group consisting of DNA damage-inducible transcript 3 (DDIT3) and p21 (cyclin-dependent kinase inhibitor 1A) A method for increasing the expression of a cancer-related gene selected from one or more genes selected from the group consisting of: 2. The method of claim 1, wherein the expression of p21 is p53-dependent. The method according to claim 1, wherein the colorectal cancer cell is a human colon cancer cell line. 4. The method according to any one of claims 1 to 3, wherein said in vitro condition is an in vitro condition. delete delete delete delete

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