Composition for inhibiting and treating cancer metastasis

Abstract

To provide a composition that has both anticancer activity against pancreatic cancer and the effect of inhibiting proliferation and metastasis of cancer cells.SOLUTION: Provided is a pharmaceutical composition for the prevention or treatment of pancreatic cancer and / or an anti-cancer supplement, containing chlorphenesin or a pharmaceutically acceptable salt thereof as an active ingredient.SELECTED DRAWING: Figure 18

Description

[Technical field]

[0001] The present invention relates to a composition for treating cancer and inhibiting metastasis, and relates to the anti-cancer and anti-metastasis effects of chlorphenesin, chloroquine and chloropyrazine, either alone or in combination. [Background technology]

[0002] The smallest unit that makes up the human body is called a cell, and normal cells maintain a balance in cell numbers by dividing, growing, or dying according to the intracellular regulatory function. If a cell is damaged by a certain cause, it can be treated and function as a normal cell, or if it does not recover, it will self-destruct. However, if a change occurs in the genes of a cell for various reasons, the cell will change abnormally and grow incompletely, and continue to divide without regulating the cell cycle. This is defined as cancer. Cancer is also characterized by invading and destroying surrounding tissues and organs, as well as spreading to other organs. Cancer mortality is the number one cause of death in Korea, and the number is increasing every year. Although there have been remarkable advances in the medical treatment of certain cancers, the overall five-year survival rate for all cancers has only improved by about 10% in the past 20 years. Cancer, or malignant tumors, metastasize and grow rapidly in an uncontrolled manner, making it extremely difficult to detect and treat them in a timely manner.

[0003] The large intestine is a long, tube-like digestive organ that starts at the end of the small intestine and continues to the anus, and cancer that occurs in this area is called colon cancer. Colon cancer is broadly divided into colon cancer and rectal cancer according to the location where it occurs. Patients with colon cancer generally show symptoms such as changes in bowel habits, bloody or mucus in the stool, thin stool, weight loss, abdominal discomfort, fatigue, and loss of appetite. Colon cancer mainly metastasizes to the liver and lungs, and more than 50% of colon cancer patients develop cancer metastasis. Conventional treatments for colon cancer include surgery and chemotherapy, and a representative targeted therapy is Cetuximab (Erbitux) injection. Cetuximab is a monoclonal antibody that targets the epidermal growth factor receptor (EGFR), and it specifically binds to EGFR on the surface of colon cancer cells, inhibiting a specific part of the signal transduction process that causes cancer cell proliferation, and suppressing the overall proliferation of cancer cells.

[0004] Pancreatic cancer is one of the most deadly forms of cancer. More than 40,000 people are diagnosed with pancreatic cancer each year in the United States, and of these, less than 5% survive more than five years after diagnosis. This low survival rate is mainly due to the fact that most pancreatic cancers are not diagnosed until they reach an advanced stage. Pancreatic cancer is usually asymptomatic in the early stages, but symptoms in later stages are non-specific and varied, making early diagnosis difficult. Treatment options for pancreatic cancer are limited. Surgery and radiation therapy may be performed for early-stage pancreatic cancer, but are less effective for advanced or recurrent pancreatic cancer. Weekly intravenous administration of gemcitabine has been found to be effective. It was approved for pancreatic cancer by the US FDA in 1998. Gemcitabine, the most commonly used anticancer drug for the treatment of pancreatic cancer, has been used in combination with other drugs such as oxalate and 5-fluorouracil (5-FU), but this has not had a significant impact on significantly increasing the survival rate of patients with pancreatic cancer. The standard of care for palliative treatment is gemcitabine, either alone or in combination with the EGFR tyrosine kinase inhibitor erlotinib. Alternative options include the combination of 5-fluorouracil, leucovorin, irinotecan, and oxaliplatin (also known as the FOLFIRINOX protocol), or the combination of gemcitabine and nab-paclitaxel, the latter of which shows superior efficacy compared to gemcitabine alone in the MPACT study (Von Hoff et al., 2013; S3-Leitlinie Exokrines Pankreaskarzinom, 2013). The US FDA has also approved the kinase inhibitor erlotinib for use in combination with gemcitabine for patients with advanced stage pancreatic cancer who have not previously received chemotherapy, but the median overall survival benefit derived from erlotinib was only less than 4 weeks (Moore et al., J. Clin. Oncol.,25(15):1960-6(2007)).

[0005] The bile duct is a tube that carries bile produced in the liver to the duodenum. Like the branches of a tree gathering toward one trunk, the bile ducts gradually merge and thicken from inside the liver, and when they leave the liver, the left and right bile ducts almost merge and become one. The bile duct is divided into the intrahepatic bile duct while running through the liver and the extrahepatic bile duct that runs from outside the liver to the duodenum. The pouch that temporarily stores and concentrates bile within the extrahepatic bile duct is called the gallbladder, and these intrahepatic and extrahepatic bile ducts and the gallbladder are collectively called the bile duct. The bile duct is a passageway for bile discharged from the liver, gradually thickening like the trunk of a tree, and opening into the duodenum. The gallbladder is the primary storage location for bile. Bile duct cancer and gallbladder cancer are collectively called biliary tract cancer, which occurs in the epithelial cells that surround the inside of the bile duct and gallbladder. Biliary tract cancer is one of the difficult-to-treat cancers, with 70-80% of cases at the time of diagnosis being progressive, surgery being possible in only 30-40%, and the 5-year survival rate being only around 7%. Although many anticancer drugs have been developed for various cancers up to now, only a few cancers can be cured completely with anticancer drugs alone. This is because when cancer is treated with anticancer drugs, cancer cells do not respond to the anticancer drugs, or tumors are effectively reduced in the early stages, but resistance to the anticancer drugs occurs during or after treatment. Therefore, for effective anticancer treatment, resistance to anticancer drugs, such as resistance of cancer cells to anticancer drugs, must be overcome. In the case of biliary tract cancer, anticancer drug resistance occurs frequently early on, the response rate to anticancer drugs is only 15%, and the recurrence rate after surgery reaches 85%, but there is no effective anticancer drug that can be used for pre- and post-operative adjuvant anticancer drug therapy.

[0006] In most cases, malignant tumors develop in one organ (lungs, liver, kidneys, stomach, colon, rectum, etc.) and then spread from the primary organ to other tissues, and this spread from the primary site to other tissues is called metastasis. Metastasis is a phenomenon that accompanies the progression of malignant tumors. As malignant tumor cells grow and the cancer progresses, they acquire new genetic traits necessary for metastasis, invade blood vessels and lymphatics, and while circulating along the blood and lymphatic vessels, they settle in other tissues and grow.

[0007] Currently, surgery, radiation therapy, chemotherapy, etc. are used to treat cancer. Among these, chemotherapy refers to a method of treating cancer using anticancer drugs. Today, about 60 different anticancer drugs are used, and as more knowledge about the development of cancer and the characteristics of cancer cells has been gained recently, research into the development of new anticancer drugs is actively progressing. In addition, current treatments focus on the death or removal of cancer cells, and there is a lack of research into drugs to prevent the proliferation and metastasis of cancer cells, which are the causes of cancer and are directly linked to the survival rate of cancer patients. Therefore, in order to improve cancer treatment and the survival rate of patients, there is an urgent need to develop a new concept drug that has both anticancer activity and the effect of inhibiting the proliferation and metastasis of cancer cells. [Prior art documents] [Non-patent literature]

[0008] [Non-Patent Document 1] Moore et al.,J. Clin. Oncol.,25(15):1960-6(2007) Summary of the Invention [Problem to be solved by the invention]

[0009] The inventors of the present invention have confirmed that chlorphenesin, chloroquine and chloropyrazine have anti-cancer effects and are effective in inhibiting the proliferation and metastasis of cancer cells, and that a combination of these has a synergistic effect, thereby completing the present invention. [Means for solving the problem]

[0010] To achieve the above object, the present invention provides a pharmaceutical composition for preventing or treating cancer, comprising one or more selected from chlorphenesin, chloroquine and chloropyrazine, or a pharma- ceutically acceptable salt thereof, as an active ingredient.

[0011] The present invention also provides a pharmaceutical composition for inhibiting cancer growth and metastasis, comprising as an active ingredient one or more selected from the group consisting of chlorphenesin, chloroquine and chloropyrazine, or a pharma- ceutical acceptable salt thereof.

[0012] The present invention also provides an anti-cancer supplement comprising, as an active ingredient, one or more compounds selected from the group consisting of chlorphenesin, chloroquine and chloropyrazine, or a pharma- ceutical acceptable salt thereof.

[0013] Further, the present invention provides a food composition for preventing or ameliorating cancer, which contains one or more selected from the group consisting of chlorphenesin, chloroquine and chloropyrazine. Effect of the Invention

[0014] The present invention relates to an anti-cancer composition aimed at suppressing both the proliferation and metastasis of cancer cells, and by administering chlorphenesin, chloroquine and chloropyrazine, either individually or in combination, it is possible to suppress proliferation and metastasis extremely effectively. [Brief description of the drawings]

[0015] [Figure 1] 1 is a graph showing the cell viability of colon cancer cell lines CT26, HCT116, and SW480 in the presence of chlorphenesin (OC-201). [Diagram 2] 1 is a graph showing the cell viability of colon cancer cell lines CT26, HCT116, and SW480 in the presence of chloroquine (OC-202). [Diagram 3] 1 is a graph showing the cell viability of colon cancer cell lines CT26, HCT116 and SW480 in the presence of chloropyrazine (OC-203). [Figure 4] 1 is a graph showing cell viability of colon cancer cell lines CT26, HCT116, and SW480 by combined treatment with chlorphenesin and chloroquine. [Diagram 5]1 is a graph showing cell viability of colon cancer cell lines CT26, HCT116 and SW480 by combined treatment with chlorphenesin and chloropyrazine. [Figure 6] FIG. 1 is a graph confirming the degree of migration of SW480 cells depending on the concentration of chlorphenesin. [Figure 7] 1 is a graph showing the degree of migration of SW480 cells depending on the concentration of chlorphenesin. [Figure 8] FIG. 1 is a graph confirming the degree of migration of HCT116 cells depending on the concentration of chlorphenesin. [Figure 9] 1 is a graph showing the degree of migration of HCT116 cells depending on the concentration of chlorphenesin. [Figure 10] FIG. 1 is a graph confirming the degree of migration of CT26 cells depending on the concentration of chlorphenesin. [Figure 11] 1 is a graph showing the degree of migration of CT26 cells depending on the concentration of chlorphenesin. [Figure 12] FIG. 1 shows the degree of migration of SW480 cells following treatment with chlorphenesin, chloroquine, and chloropyrazine, either alone or in combination. [Figure 13] 1 is a graph showing the degree of migration of SW480 cells following treatment with chlorphenesin, chloroquine, and chloropyrazine, either alone or in combination. [Figure 14] FIG. 1 shows the degree of migration of HCT116 cells following treatment with chlorphenesin, chloroquine, and chloropyrazine, either alone or in combination. [Figure 15] 1 is a graph showing the degree of migration of HCT116 cells following treatment with chlorphenesin, chloroquine, and chloropyrazine, either alone or in combination. [Figure 16] FIG. 1 shows the degree of migration of CT26 cells after treatment with chlorphenesin, chloroquine, and chloropyrazine, either alone or in combination. [Figure 17] 1 is a graph showing the degree of migration of CT26 cells following treatment with chlorphenesin, chloroquine, and chloropyrazine, either alone or in combination. [Figure 18]FIG. 1 shows a synergistic effect of combined treatment of chlorphenesin with chloroquine or chloropyrazine at different concentrations on inhibition of SW480 cell migration. [Figure 19] FIG. 1 shows synergistic effects on the inhibition of migration of HCT116 cells by combined treatment with chlorphenesin and chloroquine or chloropyrazine at different concentrations. [Figure 20] FIG. 1 shows synergistic inhibition of migration of CT26 cells by combined treatment with chlorphenesin and chloroquine or chloropyrazine at different concentrations. [Figure 21] FIG. 13 is a graph showing the effect of chlorphenesin alone on inhibiting the migration of HCT116 cells, confirmed by a wound-healing assay. [Figure 22] FIG. 1 shows the effect of chlorphenesin alone on inhibiting the migration of HCT116 cells, confirmed by wound-healing assay. [Diagram 23] 1 is a graph showing the results of a wound-healing assay of HCT116 treated with chlorphenesin alone. [Figure 24] FIG. 1 shows the effect of treating alone with chloroquine (OC-202) or chloropyrazine (OC-203) on the inhibition of migration of HCT116 cells, as confirmed by a wound-healing assay. [Diagram 25] FIG. 1 shows the migration inhibitory effect of HCT116 cells treated with chlorphenesin in combination with chloroquine or chloropyrazine, confirmed by wound-healing assay. [Figure 26] 1 is a graph showing the results of a wound-healing assay of HCT116 cells treated with chloroquine or chloropyrazine alone, or in combination with chlorphenesin and chloroquine or chloropyrazine. [Figure 27] FIG. 1 shows the results of a colony formation assay of HCT116 cells depending on the treatment concentration of chlorphenesin. [Figure 28] FIG. 1 shows the results of colony formation assay of HCT116 cells treated with chlorphenesin, chloroquine, or chloropyrazine alone, or with chlorphenesin and chloroquine or chloropyrazine in combination at different concentrations. [Figure 29]FIG. 1 shows the results of a colony formation assay of CT26 cells treated with chlorphenesin, chloroquine, or chloropyrazine alone, or with chlorphenesin and chloroquine or chloropyrazine in combination at different concentrations. [Diagram 30] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 as a function of chlorphenesin treatment concentration. [Diagram 31] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 as a function of chloroquine treatment concentration. [Diagram 32] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 depending on the chloropyrazine treatment concentration. [Diagram 33] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 treated with 5 μM chlorphenesin in combination with 1 μM to 50 μM chloroquine. [Diagram 34] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 treated with a combination of 0.5 μM chloroquine and 1 μM to 50 μM chlorphenesin. [Diagram 35] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 treated with 1 μM chloroquine in combination with 1 μM to 50 μM chlorphenesin. [Diagram 36] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 treated with chloroquine at 5 μM in combination with chlorphenesin at concentrations of 1 μM to 50 μM. [Figure 37] 1 is a graph showing the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2, and Panc-1 treated with 5 μM chlorphenesin in combination with 1 μM to 25 μM chloropyrazine. [Figure 38] FIG. 1 shows the degree of migration of pancreatic cancer cell line Panc-1 treated with chlorphenesin, chloroquine, or chloropyrazine alone, or with chlorphenesin in combination with chloroquine or chloropyrazine. [Figure 39] FIG. 1 shows synergistic effects on cell migration inhibition at different concentrations of chlorphenesin and chloroquine or chloropyrazine in the pancreatic cancer cell line Panc-1 treated with the combination of chlorphenesin and chloroquine or chloropyrazine. [Diagram 40] FIG. 1 shows the degree of migration of pancreatic cancer cell line Aspc-1 treated with chlorphenesin, chloroquine, or chloropyrazine alone, or with chlorphenesin in combination with chloroquine or chloropyrazine. [Diagram 41] FIG. 13 is a graph confirming the synergistic effect on cell migration inhibition depending on the combined treatment concentration in the pancreatic cancer cell line Aspc-1 treated with chlorphenesin and chloroquine or chloropyrazine in combination. [Diagram 42] FIG. 1 shows the results of invasion analysis of pancreatic cancer cell line Panc-1 treated with chlorphenesin, chloroquine, or chloropyrazine alone, or with chlorphenesin in combination with chloroquine or chloropyrazine. [Diagram 43] FIG. 1 shows a graph confirming a synergistic effect on the inhibition of cell invasion depending on the combined treatment concentration in the pancreatic cancer cell line Panc-1 treated with combined treatment of chlorphenesin and chloroquine or chloropyrazine. [Diagram 44] FIG. 1 shows the results of invasion analysis of pancreatic cancer cell line MIACaPa2 treated with chlorphenesin, chloroquine, and chloropyrazine alone, or with chlorphenesin in combination with chloroquine or chloropyrazine. [Diagram 45] FIG. 13 is a graph confirming the synergistic effect on the inhibition of cell invasion depending on the combined treatment concentration in the pancreatic cancer cell line MIACaPa2 treated with chlorphenesin and chloroquine or chloropyrazine in combination. [Figure 46] 1 is a graph showing the cell viability of biliary tract cancer cells SNU1079 and SNU308 as a function of chlorphenesin concentration. [Figure 47] 1 is a graph showing the cell viability of biliary tract cancer cells SNU1079 and SNU308 as a function of chloroquine concentration. [Figure 48] 1 is a graph showing the cell viability of biliary tract cancer cells SNU1079 and SNU308 as a function of chloropyrazine concentration. [Figure 49] 1 is a graph showing the cell viability of biliary tract cancer cells SNU1079 and SNU308 according to the combined treatment concentration of chlorphenesin and chloroquine. [Figure 50] 1 is a graph showing the cell viability of biliary tract cancer cells SNU1079 and SNU308 as a function of combined treatment concentration of chlorphenesin and chloropyrazine. [Figure 51] FIG. 1 shows the degree of inhibition of migration of biliary tract cancer cells SNU1079 depending on the concentration of chlorphenesin. [Figure 52] 1 is a graph confirming the degree of inhibition of migration of biliary tract cancer cells SNU1079 depending on chlorphenesin concentration. [Figure 53] FIG. 1 shows the degree of inhibition of migration of biliary tract cancer cells SNU1079 treated with chlorphenesin, chloroquine, or chloropyrazine alone, or with chlorphenesin in combination with chloroquine or chloropyrazine. [Figure 54] FIG. 13 is a graph confirming the synergistic effect on cell migration inhibition depending on the combined treatment concentration in the biliary tract cancer cell line SNU1079 treated with chlorphenesin and chloroquine, or chloropyrazine in combination. [Figure 55] FIG. 1 shows the inhibitory effect of invasiveness on biliary tract cancer cells SNU1079 treated with chlorphenesin, chloroquine, or chloropyrazine alone, or with chlorphenesin and chloroquine, or with chloropyrazine in combination. [Figure 56] FIG. 13 is a graph confirming the synergistic effect on the inhibition of cell invasion depending on the combined treatment concentration in the biliary tract cancer cell line SNU1079 treated in combination with chlorphenesin and chloroquine, or chloropyrazine. [Figure 57] 1 is a graph showing the evaluation results of lorphenesin cytotoxicity. [Figure 58] 13 shows staining images of CT26 and HCR116 cells treated or not with chlorphenesin. [Figure 59] 13 shows stained images of monolayers of CT26 cells treated or not with chlorphenesin. [Figure 60]1 shows images of lungs collected from an animal model of cancer metastasis and a graph showing the number of nodules developed in the lungs. [Figure 61] 1 is a graph showing the results of measuring body weight and tumor size in a cancer metastasis animal model. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0016] Hereinafter, the present invention will be described in detail with reference to the accompanying drawings. However, the following examples are given as examples of the present invention, and if it is determined that a detailed description of a technique or configuration well known to those skilled in the art may unnecessarily obscure the gist of the present invention, the detailed description may be omitted and the present invention is not limited thereby. The present invention is subject to various modifications and applications within the scope of the claims and the scope of equivalents interpreted therefrom.

[0017] In addition, the terminology used in this specification is used to appropriately express the preferred embodiment of the present invention, and may vary depending on the intention of the user or operator, or the practice of the field to which the present invention belongs. Therefore, the definition of the term should be based on the entire contents of this specification. In the entire specification, when a part "includes" a certain element, this does not exclude other elements, but means that other elements may be further included, unless otherwise specified to the contrary.

[0018] In one aspect, the present invention relates to a pharmaceutical composition for preventing or treating cancer, comprising one or more selected from chlorphenesin, chloroquine and chloropyrazine, or a pharma- ceutically acceptable salt thereof, as an active ingredient.

[0019] In one embodiment, chlorphenesin can be represented by the following formula 1, chloroquine can be represented by the following formula 2, and chloropyrazine can be represented by the following formula 3.

[0020] [ka]

[0021] [ka]

[0022] [ka]

[0023] In one embodiment, chlorphenesin can be chlorphenesin carbamate represented by the following formula 4:

[0024] [ka]

[0025] The chlorphenesin carbamate of the present invention is mainly used as a muscle relaxant, and is known to have sedative and anxiolytic effects as well as antifungal and antibacterial effects.

[0026] In one embodiment, the pharmaceutical composition of the present invention may contain chlorphenesin and chloroquine, chlorphenesin and chloropyrazine, chloroquine and chloropyrazine, or chlorphenesin, chloroquine and chlorphenesin, or pharma- ceutical acceptable salts thereof as active ingredients, and it is more preferable to contain chlorphenesin and chloroquine, or chlorphenesin and chloropyrazine, since they have a synergistic anti-cancer effect.

[0027] In one embodiment, the pharmaceutical composition of the present invention may contain 5-500 μM chlorphenesin, 0.5-25 μM chloroquine, or 1-100 μM chloropyrazine, and when chlorphenesin and chloroquine are contained together, the pharmaceutical composition may contain 5 μM (fixed concentration) chlorphenesin and 0.5-25 μM chloroquine, and when chlorphenesin and chloropyrazine are contained together, the pharmaceutical composition may contain 5 μM (fixed concentration) chlorphenesin and 25-50 μM chloropyrazine. In one embodiment of the present invention, the chlorphenesin, chloroquine, and / or chloropyrazine of the present invention inhibited the migration and invasion of cancer cells without serious cytotoxicity in cell experiments within the above concentration ranges.

[0028] In one embodiment, the cancer may be any one or more selected from the group consisting of brain tumor, melanoma, myeloma, non-small cell lung cancer, oral cancer, liver cancer, stomach cancer, colon cancer, breast cancer, lung cancer, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, endometrial cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, esophageal cancer, lymph node cancer, bladder cancer, biliary tract cancer (gallbladder and bile duct cancer), endocrine gland cancer, thyroid cancer, parathyroid cancer, adrenal gland cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, kidney or ureter cancer, renal cell carcinoma, renal pelvis cancer, central nervous system tumor, primary central nervous system lymphoma, spinal cord tumor, brain stem glioma, and pituitary adenoma, and more preferably, colon cancer, pancreatic cancer, or biliary tract cancer. In one embodiment of the present invention, the anticancer effects of chlorphenesin, chloroquine, and chlorphenesin, respectively, and the anticancer effects of combination treatment of these in mouse colon carcinoma cell line CT26, human colorectal carcinoma cell line HCT116, human colon carcinoma cell line SW480, human pancreatic carcinoma cell line Panc-1, human pancreatic cancer cell line Aspc-1, human pancreatic cancer cell line MIAPaCA2, human gallbladder carcinoma cell line SNU308, and human intrahepatic cholangiocarcinoma cell line SNU1079 were examined.

[0029] The present invention includes not only chlorphenesin, chloroquine, and chloropyrazine represented by chemical formulas 1 to 3, but also pharma- ceutically acceptable salts thereof, and solvates, hydrates, racemates, or stereoisomers thereof which can be prepared thereby.

[0030] Chlorphenesin, chloroquine and chloropyrazine represented by Chemical Formulas 1 to 3 of the present invention can be used in the form of a pharma- ceutically acceptable salt, and useful salts are acid addition salts formed with pharma- ceutically acceptable free acids. Acid addition salts are obtained from inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, nitrous acid or phosphorous acid, and non-toxic organic acids such as aliphatic mono- and dicarboxylates, phenyl-substituted alkanoates, hydroxyalkanoates and alkanedioates, aromatic acids, and aliphatic and aromatic sulfonic acids. Such pharma- ceutically non-toxic salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate chloride, bromide, iodide, fluoride, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caprate, heptanoate, propionate, oxalate, malonate, succinate, suberate, sebacate, fumarate, malate, butyrate-1,4-dioate, hexaphosphate ... The benzoate, benzoyl peroxide, benzoic acid, benzoate, benzoyl peroxide, benzoic acid, benzoyl peroxide ...

[0031] The acid addition salts according to the present invention can be prepared by a conventional method, for example, by dissolving chlorphenesin, chloroquine, and chloropyrazine represented by Chemical Formulas 1 to 3 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. Alternatively, the mixture can be dried by evaporating the solvent and excess acid, or the precipitated salt can be suction filtered to prepare the salt.

[0032] Also, a pharma- ceutically acceptable metal salt can be prepared using a base. An alkali metal or alkaline earth metal salt can be obtained, for example, by dissolving a compound in a solution of an excess of an alkali metal hydroxide or an alkaline earth metal hydroxide, filtering the undissolved compound salt, and evaporating and drying the remaining liquid. Here, it is pharma- ceutical appropriate to prepare a sodium, potassium, or calcium salt as the metal salt. Also, 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).

[0033] The pharmaceutical compositions of the present invention may further contain known anti-cancer drugs in addition to chlorphenesin, chloroquine and chlorphenesin as active ingredients, and may be used in combination with other known therapies for the treatment of these diseases, including, but not limited to, chemotherapy, radiation therapy, hormone therapy, bone marrow transplantation, stem-cell replacement therapy, other biological therapies, immunotherapy, etc.

[0034] In the present invention, the term "prevention" means any action of suppressing or delaying the occurrence, spread, and recurrence of cancer by administering the pharmaceutical composition according to the present invention, and "treatment" means any action of killing cancer cells or improving or beneficially changing the symptoms of cancer by administering one or more selected from chlorphenesin, chloroquine, and chloropyrazine, or a pharma- ceutically acceptable salt thereof, or a composition containing the same. A person having ordinary skill in the art to which the present invention pertains can refer to materials presented by the Korean Medical Association, etc., to know the exact criteria for diseases for which the composition of the present application is effective, and can judge the degree of improvement, enhancement, and treatment.

[0035] In the present invention, the term "therapeutically effective amount" used in combination with an active ingredient means an amount effective for preventing or treating a target disease, and the therapeutically effective amount of the composition of the present invention may vary depending on several factors, such as the administration method, the target site, and the condition of the patient. Therefore, the dosage when used in the human body must be determined by considering both safety and efficacy. It is also possible to estimate the amount to be used in humans from the effective amount determined through animal experiments. These matters to be considered when determining the effective amount are described, for example, in Hardman and Limbird, eds., Goodman and Gilman's The Pharmacological Basis of Therapeutics, 10th ed. (2001), Pergamon Press; and EW Martin ed,, Remington's Pharmaceutical Sciences, 18th ed, (1990), Mack Publishing Co.

[0036] The pharmaceutical composition of the present invention is administered in a pharmacologic effective amount. The term "pharmacologic effective amount" as used herein means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to any medical treatment and without causing side effects, and the effective dose level can be determined based on factors including the patient's health condition, type of cancer, severity, drug activity, drug sensitivity, administration method, administration time, administration route, and excretion rate, treatment period, co-administered or co-administered drugs, and other factors well known in the medical field. The composition 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 multiple times. Taking all of the above factors into consideration, it is important to administer an amount that can obtain maximum effect at a minimum amount without side effects, which can be easily determined by those skilled in the art.

[0037] The pharmaceutical composition of the present invention may contain carriers, diluents, excipients, or combinations of two or more thereof that are commonly used in biological preparations. The term "pharmaceutical acceptable" as used herein means that the composition exhibits the property of being non-toxic to cells or humans exposed to the composition. The carrier is not particularly limited as long as it is suitable for in vivo delivery of the composition, and may be, for example, a compound listed in the Merck Index, 13th ed., Merck & Co. Inc., saline, sterile water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol, or a mixture of one or more of these components, and other commonly used additives such as antioxidants, buffers, bacteriostatic agents, etc. may be added as necessary. In addition, the composition may be formulated into an injectable preparation, such as an aqueous solution, suspension, emulsion, pill, capsule, granule, or tablet, by additionally adding a diluent, dispersant, surfactant, binder, and lubricant. Furthermore, formulations can be suitably prepared according to each disease or ingredient using a method appropriate to the art or a method disclosed in Remington's Pharmaceutical Sciences (Mack Publishing Company, Easton PA, 18th, 1990).

[0038] In one embodiment, the pharmaceutical composition may be one or more formulations selected from the group including oral formulations, topical preparations, suppositories, sterile injection solutions and sprays, with oral or injectable formulations being more preferred.

[0039] The term "administration" as used in the present invention means providing a given substance to an individual or patient in any suitable manner, and can be administered parenterally (e.g., intravenously, subcutaneously, intraperitoneally or locally in an injection preparation) or orally depending on the intended method, and the dosage range varies depending on the patient's weight, age, sex, health condition, diet, administration time, administration method, excretion rate, and disease severity. Liquid preparations for oral administration of the composition of the present invention include suspensions, oral liquids, emulsions, syrups, etc., which may contain various excipients, so-called wetting agents, sweeteners, flavoring agents, preservatives, etc., in addition to water and liquid paraffin, which are commonly used simple diluents. Preparations for parenteral administration include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories, etc. The pharmaceutical composition of the present invention can also be administered by any device that can transfer the active substance to target cells. Preferred administration methods and preparations are intravenous injections, subcutaneous injections, intradermal injections, intramuscular injections, drip injections, etc. Injectables can be prepared using aqueous solvents such as physiological saline and Ringer's solution, or non-aqueous solvents such as vegetable oils, higher fatty acid esters (e.g., ethyl oleate, etc.), and alcohols (e.g., ethanol, benzyl alcohol, propylene glycol, glycerin, etc.), and can contain pharmaceutical carriers such as stabilizers to prevent deterioration (e.g., ascorbic acid, sodium bisulfite, sodium pyrosulfite, BHA, tocopherol, EDTA, etc.), emulsifiers, buffers for pH adjustment, and preservatives to prevent microbial growth (e.g., phenylmercuric nitrate, thimerosal, benzalkonium chloride, phenol, cresol, benzyl alcohol, etc.).

[0040] The term "individual" as used in the present invention means all animals including monkeys, cows, horses, sheep, pigs, chickens, turkeys, quails, cats, dogs, mice, rats, rabbits, and guinea pigs, including humans, who are affected or may be affected by the above-mentioned cancers, and the above-mentioned diseases can be effectively prevented or treated by administering the pharmaceutical composition of the present invention to an individual. The pharmaceutical composition of the present invention can be administered in parallel with a conventional therapeutic agent.

[0041] The pharmaceutical composition of the present invention may further include a pharma- ceutically acceptable additive, and examples of the pharma-ceutically acceptable additive include starch, gelatinized starch, microcrystalline cellulose, lactose, povidone, colloidal silicon dioxide, calcium hydrogen phosphate, lactose, mannitol, candy, gum arabic, pregelatinized starch, corn starch, powdered cellulose, hydroxypropyl cellulose, Opadry, sodium starch glycolate, carbonauba wax, synthetic aluminum silicate, stearic acid, magnesium stearate, aluminum stearate, calcium stearate, sucrose, dextrose, sorbitol, and talc. The pharma-ceutically acceptable additive according to the present invention is preferably included in an amount of 0.1 to 90 parts by weight based on the composition, but is not limited thereto.

[0042] In one aspect, the present invention relates to a pharmaceutical composition for inhibiting cancer growth and metastasis, comprising one or more selected from chlorphenesin, chloroquine and chloropyrazine, or a pharma- ceutically acceptable salt thereof, as an active ingredient.

[0043] In one embodiment, chlorphenesin can be represented by the following formula 5, chloroquine can be represented by the following formula 6, and chloropyrazine can be represented by the following formula 7.

[0044] [ka]

[0045] [ka]

[0046] [ka]

[0047] In one embodiment, chlorphenesin can be chlorphenesin carbamate represented by the following formula 8:

[0048] [ka]

[0049] In one embodiment, the pharmaceutical composition of the present invention may contain chlorphenesin and chloroquine, chlorphenesin and chloropyrazine, chloroquine and chloropyrazine, or chlorphenesin, chloroquine and chlorphenesin, or pharma- ceutical acceptable salts thereof as active ingredients, and it is more preferable to contain chlorphenesin and chloroquine, or chlorphenesin and chloropyrazine, since this provides a synergistic effect of inhibiting metastasis and invasion.

[0050] In one embodiment, the pharmaceutical composition of the present invention may contain 5-500 μM chlorphenesin, 0.5-25 μM chloroquine, or 1-100 μM chloropyrazine, and when both chlorphenesin and chloroquine are contained, the pharmaceutical composition may contain 5 μM (fixed concentration) chlorphenesin and 0.5-25 μM chloroquine, and when both chlorphenesin and chloropyrazine are contained, the pharmaceutical composition may contain 5 μM (fixed concentration) chlorphenesin and 25-50 μM chloropyrazine. In one embodiment of the present invention, within the above concentration ranges, the migration and invasion of cancer cells were inhibited without serious cytotoxicity.

[0051] The chlorphenesin of the present invention can inhibit only the proliferation and metastasis of cancer cells at a low concentration of 0.1 μM to 10 mM, but not the death of cancer cells. For example, the composition of the present invention can contain a low concentration of chlorphenesin in the range of 1 μM to 1 mM. When the chlorphenesin is at a concentration of less than 1 μM, the effect of inhibiting the proliferation and metastasis of cancer cells is reduced compared to 1 μM, and when the chlorphenesin is at a concentration exceeding 1 mM, particularly 10 mM or more, it can exhibit cytotoxicity.

[0052] In one embodiment, the cancer may be colorectal cancer, pancreatic cancer, or biliary tract cancer. In one embodiment of the present invention, the cancer cell metastasis and invasion inhibitory effects of chlorphenesin, chloroquine, and chlorphenesin on mouse-derived colon cancer cell line CT26, human-derived colorectal cancer cell line HCT116, human-derived colon cancer cell line SW480, human-derived pancreatic cancer cell line Panc-1, human-derived pancreatic cancer cell line Aspc-1, human-derived pancreatic cancer cell line MIAPaCA2, human-derived gallbladder cancer cell line SNU308, and human-derived intrahepatic cholangiocarcinoma cell line SNU1079, and the cancer cell metastasis and invasion inhibitory effects of combination treatment of these compounds were confirmed.

[0053] In one aspect, the present invention relates to an anti-cancer supplement comprising one or more selected from chlorphenesin, chloroquine, and chloropyrazine, or a pharma- ceutically acceptable salt thereof, as an active ingredient.

[0054] In one embodiment, the chlorphenesin of the present invention can suppress only the proliferation and metastasis of cancer cells, not the death of cancer cells, at low concentrations, so that when administered in combination with an anticancer drug that already has cytotoxicity, the toxicity of cells can be minimized.For example, the composition of the present invention can contain a low concentration of chlorphenesin in the range of 1 μM to 1 mM.When the concentration of chlorphenesin is less than 1 μM, it does not have the effect of suppressing cancer proliferation and metastasis, and when it is more than 1 mM, especially 10 mM or more, it can show cytotoxicity.

[0055] In one embodiment, chlorphenesin and chloroquine, or chlorphenesin and chloropyrazine may be included as active ingredients. In one embodiment of the present invention, it was confirmed that the size and metastasis of tumors induced in mouse-derived colon carcinoma cell line CT26 were significantly suppressed by co-treatment with chlorphenesin anticancer drug.

[0056] Examples of anti-cancer agents that can be included in the pharmaceutical compositions of the present invention include DNA alkylating agents such as mechloethamine, chlorambucil, phenylalanine, mustard, cyclophosphamide, ifosfamide, carmustine (BCNU), lomustine (CCNU), streptozotocin, busulfan, thiotepa, cisplatin, and carboplatin; anti-cancer antibiotics such as dactinomycin (actinomycin A); D), doxorubicin (adriamycin), daunorubicin, idarubicin, mitoxantrone, plicamycin, mitomycin C, and bleomycin; and plant alkaloids such as vincristine, vinblastine, paclitaxel, docetaxel, etoposide, teniposide, topotecan, and irinotecan.

[0057] In one aspect, the present invention relates to a food composition for preventing or ameliorating cancer, comprising one or more selected from chlorphenesin, chloroquine and chloropyrazine.

[0058] When the composition of the present invention is used as a food composition, the chlorphenesin, chloroquine or chlorphenesin can be added as it is or can be used together with other foods or food ingredients, and can be used appropriately according to a conventional method. The composition can contain a food-scientifically acceptable food supplement additive in addition to the active ingredient, and the amount of the active ingredient mixed can be appropriately determined based on the purpose of use (prevention, health or therapeutic treatment).

[0059] The term "nutritional supplement additive" used in the present invention means a component that can be added supplementarily to food, which is added to produce the health functional food of each formulation, and can be appropriately selected and used by those skilled in the art. Examples of nutritional supplements include various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants, and fillers, pectinic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, and carbonation agents used in carbonated drinks, but the above examples do not limit the types of food supplement additives of the present invention.

[0060] The food composition of the present invention may include a functional health food. The term "functional health food" used in the present invention refers to a food manufactured and processed in the form of tablets, capsules, powders, granules, liquids, and pills using raw materials and ingredients having useful functionality for the human body. Here, "functionality" means that nutrients are adjusted for the structure and function of the human body, and useful effects for health purposes such as physiological action are obtained. The functional health food of the present invention can be manufactured by a method commonly used in the ordinary technical field, and during the manufacturing, raw materials and ingredients commonly added in the ordinary technical field can be added to manufacture. In addition, the formulation of the functional health food can also be manufactured without restrictions as long as it is a formulation recognized as a functional health food. The food composition of the present invention can be manufactured in various forms of formulation, and unlike general medicines, it has the advantage of being made of food as a raw material and having no side effects that may occur when taking medicines for a long period of time, and is highly portable, and the functional health food of the present invention can be taken as a supplement to enhance the effect of anticancer drugs.

[0061] In addition, there is no limit to the type of health food that the composition of the present invention can be used in. In addition, the composition of the present invention containing chlorphenesin, chloroquine or chlorphenesin as an active ingredient can be prepared by mixing other suitable auxiliary ingredients that can be contained in health functional foods and known additives according to the selection of a person skilled in the art. Examples of foods that can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, tea, energy drinks, alcoholic beverages and vitamin-containing preparations, and can be prepared by adding the extract of the present invention to soups, teas, jellies and juices that are prepared as the main ingredient.

[0062] In one aspect, the present invention relates to a method of treating cancer comprising administering to an individual suffering from cancer a pharma- tically effective amount of one or more selected from the group consisting of chlorphenesin, chloroquine and chloropyrazine, or a pharma- tically acceptable salt thereof.

[0063] In one embodiment, chlorphenesin and chloroquine, chlorphenesin and chloropyrazine, chloroquine and chloropyrazine, or chlorphenesin, chloroquine and chlorphenesin, or pharma- ceutically acceptable salts thereof may be administered, with administration of chlorphenesin and chloroquine, or chlorphenesin and chloropyrazine together being more preferred as they provide a synergistic anti-cancer effect.

[0064] In one embodiment, the pharmaceutical composition of the present invention may contain 5-500 μM chlorphenesin, 0.5-25 μM chloroquine, or 1-100 μM chloropyrazine, and when both chlorphenesin and chloroquine are contained, the pharmaceutical composition may contain 5 μM (fixed concentration) chlorphenesin and 0.5-25 μM chloroquine, and when both chlorphenesin and chloropyrazine are contained, the pharmaceutical composition may contain 5 μM (fixed concentration) chlorphenesin and 25-50 μM chloropyrazine. In one embodiment of the present invention, within the above concentration ranges, the migration and invasion of cancer cells were inhibited without serious cytotoxicity.

[0065] In one embodiment, the cancer is brain cancer, melanoma, myeloma, non-small cell lung cancer, oral cancer, liver cancer, stomach cancer, colon cancer, breast cancer, lung cancer, bone cancer, pancreatic cancer, skin cancer, head or neck cancer, cervical cancer, ovarian cancer, colon cancer, small intestine cancer, rectal cancer, fallopian tube cancer, anal cancer, endometrial cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, esophageal cancer, lymph node cancer, bladder cancer, biliary tract cancer (gallbladder and bile duct cancer), endocrine gland cancer, thyroid cancer, parathyroid cancer, The cancer may be any one or more selected from the group consisting of adrenal gland cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, kidney or ureter cancer, renal cell carcinoma, renal pelvis cancer, central nervous system tumor, primary central nervous system lymphoma, spinal cord tumor, brain stem glioma and pituitary adenoma, and is more preferably colon cancer, pancreatic cancer or biliary tract cancer.

[0066] In one aspect, the present invention relates to the use of one or more selected from the group consisting of chlorphenesin, chloroquine and chloropyrazine, or a pharma- ceutically acceptable salt thereof, for use in the manufacture of a pharmaceutical composition for the prevention and treatment of cancer.

[0067] The present invention will be described in more detail with reference to the following examples, which are provided for illustrative purposes only and are not intended to limit the scope of the present invention. EXAMPLES

[0068] Confirmation of anti-cancer and metastasis-suppressing effects against colon cancer 1-1. Confirmation of cell viability 1-1-1. Confirmation of cell viability by single administration To confirm the effect of single administration of chlorphenesin (OC-201), chloroquine (OC-202), and chloropyrazine (OC-203) on the viability of colon cancer cells, the viability of colon cancer cell lines CT26, HCT116, and SW480 was evaluated by MTT assay (Promega, Ltd.) according to the manufacturer's protocol. Each colon cancer cell line was cultured at 5 × 10 cells per well. 3 Cells were seeded in a 96-well plate at a density of 100 μM (control group DMSO treatment), 10 μM, 25 μM, 50 μM, 100 μM, 250 μM, 500 μM and 1 mM for 24 h, 48 h or 72 h. The cells were pretreated with chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203), respectively, at concentrations of 0 μM (control group DMSO treatment), 10 μM, 25 μM, 50 μM, 100 μM, 250 μM, 500 μM and 1 mM, respectively, and incubated with 5 mg / mL MTT for 4 hours. After that, the medium was removed, and 150 μL of solubilization solution and suspension solution were added, followed by incubation at 30 °C for 4 hours. The absorbance of the reaction solution was measured at 570 nm. The cell viability was calculated using the following formula: (Number 1) Cell viability = experimental group absorbance (570 nm) / control group absorbance (570 nm) × 100 (%)

[0069] As a result, as shown in Figures 1 to 3, it was found that OC-201 showed toxicity at concentrations exceeding 500 μM, OC-202 showed toxicity at concentrations exceeding 10 uM, and OC-203 showed no toxicity at concentrations of 100 uM or less.

[0070] 1-1-2. Confirmation of cell viability by combined administration To confirm the viability of colon cancer cells after combined treatment with chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203), cell viability was evaluated for colon cancer cell lines CT26, HCT116 and SW480 by MTT assay (Promega, Ltd.) according to the manufacturer's protocol. Each colon cancer cell line was cultured at 5 × 10 per well. 3 Cells were seeded into 96-well plates at a density of 100 μM each, and treated with control (DMSO treatment), chlorphenesin 5 μM, chlorphenesin and chloroquine (5 μM+500 nM, 5 μM+1 μM, 5 μM+5 μM, 5 μM+10 μM, 5 μM+25 μM and 5 μM+50 μM), or chlorphenesin and chloropyrazine (5 μM+1 μM, 5 μM+5 μM, 5 μM+10 μM, 5 μM+25 μM, 5 μM+50 μM and 5 μM+100 μM) for 24 h, 48 h or 72 h, respectively, and incubated with 5 mg / mL MTT for 4 h. Then, the medium was removed, and 150 μL of solubilization solution and suspension solution were added, followed by incubation at 30 °C for 4 h. The absorbance of the reaction solution was measured at 570 nm. The cell viability was calculated using the above (Equation 1).

[0071] As a result, when chlorphenesin 5 μM was combined with chloroquine at 25 μM or more, it was toxic to colon cancer cells (Figure 4), whereas when chlorphenesin 5 μM was combined with chloropyrazine at 100 μM or less, it was not toxic to colon cancer cells (Figure 5).

[0072] 1-2. Confirmation of cell migration 1-2-1.Migration assay 1-2-1-1. Chlorphenesin monotherapy Since cancer cell metastasis is related to cell motility, the migration of colon cancer cell lines SW480, HCT116, and CT26 cell lines by the treatment concentration of chlorphenesin (OC-201) was confirmed using a migration analysis method. Specifically, colon cancer cell lines CT26, HCT116, and SW480 cell lines were suspended in serum-free RPMI, and then 1x10 cells per well were placed in the upper chamber of a 24-well transwell chamber with a polycarbonate membrane (8.0 μM pore size, Costar). 5 Laminin (10 μg / ml) was placed in the lower well, and each well was treated with chlorphenesin (OC-201) at 0 μM (DMSO-treated control), 5 μM, 10 μM, 25 μM, 50 μM, 100 μM, 250 μM, 500 μM, 1 mM, and 2 mM. The cells were incubated at 37°C in CO 2 The cells were cultured in an incubator for 18 hours to allow migration. Afterwards, the cells were fixed with 70% methyl alcohol in PBS for 30 minutes and washed three times with PBS. The cells were stained with hematoxylin (Sigma) for 10 minutes and washed with distilled water. Non-migrating cells were removed from the upper surface of the membrane with a cotton swab. The membrane was excised from the chamber and fixed with Gel Mount (Biomeda, Foster City, CA). Migrated cells (cells attached to the lower surface of the membrane) were counted under a high-powered (x20) scope randomly selected from the control.

[0073] As a result, in the SW480 cell line, cell migration was significantly reduced when treated with 25 μM or more of chlorphenesin (OC-201) (Figures 6 and 7). In the HCT116 cell line, cell migration was reduced when treated with chlorphenesin (OC-201), and was particularly significantly reduced when treated with 250 μM or more (Figures 8 and 9). In addition, cell migration was also reduced in the CT26 cell line when treated with chlorphenesin (OC-201), and was particularly significantly reduced when treated with 250 μM or more (Figures 10 and 11).

[0074] 1-2-1-2. Combined use The migration of colon cancer cell lines CT26, HCT116 and SW480 was examined when treated with chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203) alone, and when treated with chlorphenesin and chloroquine or chloropyrazine in combination.

[0075] Specifically, colon cancer cell lines CT26, HCT116, or SW480 were treated with a control group (DMSO treatment), chlorphenesin (5μM), chloroquine (5μM, 10μM, or 25μM), chloropyrazine (25μM or 50μM), chlorphenesin and chloroquine (5μM+5μM, 5μM+10μM, 5μM+25μM), and chlorphenesin and chloropyrazine (5μM+25μM, 5μM+50μM), and the degree of cell migration was confirmed in the same manner as in the previous examples. In addition, the synergistic effect during combination treatment was calculated through the combination index (CI) according to the combination treatment concentration of chlorphenesin and chloroquine or chloropyrazine using Compusyn software.

[0076] As a result, it was confirmed that the migration of colon cancer cell lines CT26, HCT116 and SW480 was reduced when chlorphenesin and chloroquine or chlorphenesin and chloropyrazine were co-treated compared to when chloroquine (OC-202) or chloropyrazine (OC-203) were co-treated alone (Figures 12 to 17). In addition, synergistic effects were observed when chlorphenesin and chloroquine or chloropyrazine were co-treated, and in particular, synergistic effects were observed in all cell lines when chlorphenesin and chloroquine were co-treated (Figures 18 to 20).

[0077] 1-2-2. Wound healing assay 1-2-2-1. Chlorphenesin monotherapy Since cancer cell metastasis is a relationship that should be premised on cell motility, the degree of migration of colon cancer cell line HCT116 when treated alone with chlorphenesin (OC-201) was confirmed by a wound healing assay. Specifically, colon cancer cell line HCT116 was added to RPMI supplemented with 10% FBS, and after 24 hours, it was seeded into a 24-well tissue culture plate at a concentration at which it reached 70-80% confluence in a monolayer. A scratch was carefully and slowly made on the tomography with a new 200 μL yellow pipette tip across the center of the well. The resulting gap distance was made equal to the outer diameter of the end of the tip. After making the scratch, the dish was carefully washed twice with medium to remove the detached cells. Thereafter, chlorphenesin was applied at 0 μM (DMSO), 5 μM, 10 μM, 25 μM, 50 μM, 250 μM, 500 μM or 1 mM, and after incubation for 0, 8 and 24 hours, the cells were examined under a microscope and graphed.

[0078] As a result, the migration of colon cancer cells was reduced when treated with chlorphenesin at 25 μM or more (FIGS. 21 to 23).

[0079] 1-2-2-2. Combined use The migration degree of colon cancer cell line HCT116 was confirmed when treated with chloroquine (OC-202) and chloropyrazine (OC-203) alone and in combination with chlorphenesin (OC-201). Specifically, colon cancer cell line HCT116 was seeded into 24-well tissue culture plates at a concentration where the monolayer reached 70-80% confluence after 24 h in RPMI supplemented with 10% FBS. A scratch was carefully and slowly made in the tomography with a new 200 μL yellow pipette tip across the center of the well. The resulting gap distance was made equal to the outer diameter of the end of the tip. After making the scratch, the dish was carefully washed twice with medium to remove the detached cells. Thereafter, the cells were treated with chloroquine alone (5, 10, or 25 μM), chloropyrazine alone (25 or 50 μM), chlorphenesin and chloroquine in combination (5 μM + 5 μM, 5 μM + 10 μM, 5 μM + 25 μM), and chlorphenesin and chloropyrazine in combination (5 μM + 25 μM, 5 μM + 50 μM), and the degree of cell migration was confirmed at 0 hours, 8 hours, or 24 hours after treatment in the same manner as in Example 1-2-2-1.

[0080] The results showed that the migration of colon cancer cells was reduced when treated with chloroquine (OC-202) or chloropyrazine (OC-203) together with chlorphenesin compared to when treated alone (FIGS. 24 to 26).

[0081] 1-3. Non-adherent growth analysis 1-3-1.Chlorphenesin monotherapy Anchorage independent growth is an important trait that distinguishes normal cells from cancer cells. Normal cells require anchorage to grow, but cancer cells can survive and grow without attachment. That is, normal cells cannot grow if they are not attached to a culture plate, but cancer cells can grow in a floating state without cell adhesion, such as in soft agar. Taking advantage of this characteristic, we confirmed the non-adherent growth ability through a soft agar colony formation assay. First, we performed a colony formation assay to confirm the non-adherent growth ability of colon cancer cell lines by administering chlorphenesin alone. Specifically, 3,000 colon cancer cell lines HCT116 were mixed with soft agar and dispensed into 6-well plates, and then treated with chlorphenesin at 0μM (DMSO), 5μM, 10μM, 25μM, 50μM, 100μM, 250μM, 500μM, 1mM, or 2mM. Thereafter, chlorphenesin was also added whenever the cell culture medium was changed, and the cells were observed 3 weeks after division.

[0082] As a result, as shown in FIG. 27, it was shown that the colony forming ability was reduced when treated with 250 μM or more of chlorphenesin.

[0083] 1-3-2. Concomitant use To confirm whether the combined treatment of chlorphenesin with chloroquine (OC-202) or chloropyrazine (OC-203) suppresses the non-adherent proliferation of colon cancer cell lines compared to single treatment, colon cancer cell lines HCT116 and CT26 were treated with chlorphenesin (5 μM), chloroquine (10 or 25 μM), chloropyrazine (10 μM), chlorphenesin and chloroquine (5 μM + 10 μM or 5 μM + 25 μM), and chlorphenesin and chloropyrazine (5 μM + 10 μM, 5 μM + 25 μM, or 5 μM + 50 μM), respectively, and colony formation assays were performed.

[0084] As a result, colony formation was reduced by the combined treatment with chlorphenesin and chloroquine compared to the treatment with chloroquine or chloropyrazine alone (FIGS. 28 and 29). EXAMPLES

[0085] Confirmation of efficacy against pancreatic cancer 2-1. Confirmation of cell viability 2-1-1. Confirmation of cell viability by single administration To confirm the effect of each of the single administrations of chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203) on the viability of pancreatic cancer cells, the cell viability of the pancreatic cancer cell lines Aspc-1, MIAPaCA2 and Panc-1 was evaluated by MTT assay (Promega, Ltd.) according to the manufacturer's protocol. Each pancreatic cancer cell line was cultured at 5 × 10 per well. 3 Cells were seeded in 96-well plates at a density of 1000 μM and pretreated with chlorphenesin (OC-201) at 0 μM (control DMSO treatment), 5 μM, 10 μM, 25 μM, 50 μM, 100 μM, 250 μM, 500 μM, and 1 mM (1000 μM), chloroquine (OC-202) at 0 μM (control DMSO treatment), 0.5 μM, 1 μM, 5 μM, 10 μM, 25 μM, 50 μM, and 100 μM, and chloropyrazine (OC-203) at 0 μM (control DMSO treatment), 1 μM, 5 μM, 10 μM, 25 μM, 50 μM, and 100 μM, respectively, for 24 h, 48 h, or 72 h. The cells were then incubated with 5 mg / mL MTT for 4 h. Then, the medium was removed, 150 μL of solubilization solution and suspension solution were added, and the mixture was incubated at 30° C. for 4 hours. The absorbance of the reaction solution was measured at 570 nm. The cell viability was calculated using the above (Equation 1).

[0086] As a result, as can be seen from Figures 30 to 32, in the cases of the groups containing chlorphenesin and chloropyrazine alone, no cytotoxicity was observed even at large doses, whereas in the case of chloroquine, cytotoxicity was observed at concentrations of 50 µM or more.

[0087] 2-1-2. Confirmation of cell viability by combined administration To confirm the viability of pancreatic cancer cells upon combined treatment with a combination of chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203), the cell viability of pancreatic cancer cell lines Aspc-1, MIAPaCA2 and Panc-1 was evaluated by MTT assay (Promega, Ltd.) according to the manufacturer's protocol. Each pancreatic cancer cell line was cultured at 5 × 10 per well. 3 Cells were seeded in 96-well plates at a density of 1000 x 1000 and treated with control (DMSO treatment), chlorphenesin 5 μM, chlorphenesin and chloroquine (combination of chlorphenesin 5 μM and chloroquine 1, 5, 10, 25, or 50 μM, combination of chloroquine 0.5 μM and chlorphenesin 1, 5, 10, 25, or 50 μM, combination of chloroquine 1 μM and chlorphenesin 1, 5, 10, 25, or 50 μM, or combination of chloroquine 5 μM and chlorphenesin 1, 5, 10, 25, or 50 μM), or chlorphenesin and chloropyrazine (combination of chlorphenesin 5 μM and chloropyrazine 1, 5, 10, 25, or 50 μM) for 24 h, 48 h, or 72 h, respectively, and then the cells were incubated with 5 mg / mL MTT for 4 h. Then, the medium was removed, 150 μL of solubilization solution and suspension solution were added, and the mixture was incubated at 30° C. for 4 hours. The absorbance of the reaction solution was measured at 570 nm. The cell viability was calculated using the above (Equation 1).

[0088] As a result, when chlorphenesin 5 μM was combined with chloroquine at 1 μM to 50 μM (Figure 33), when chloroquine 0.5 μM was combined with chlorphenesin at 1 μM to 50 μM (Figure 34), when chloroquine 1 μM was combined with chlorphenesin at 1 μM to 50 μM (Figure 35), and when chloroquine 5 μM was combined with chlorphenesin at 1 μM to 50 μM (Figure 36), no serious cytotoxicity was observed in the pancreatic cancer cell lines up to 72 hours, and when chlorphenesin 5 μM was combined with chloropyrazine at 1 μM to 25 μM (Figure 37), no serious cytotoxicity was observed in the pancreatic cancer cell lines up to 24 hours.

[0089] 2-2.Cell migration assay 2-2-1. Comparison of single and combined treatments The degree of migration of pancreatic cancer cell lines Panc-1 and Aspc-1 was confirmed by the same method as in Example 1-2-1 when chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203) were treated alone, and when chlorphenesin and chloroquine or chloropyrazine were treated in combination, respectively. Specifically, the pancreatic cancer cell lines Panc-1 and Aspc-1 were treated with a control group (DMSO), chlorphenesin (5 μM), chloroquine (5 μM, 10 μM or 25 μM), chloropyrazine (25 μM or 50 μM), chlorphenesin and chloroquine (5 μM + 5 μM, 5 μM + 10 μM, 5 μM + 25 μM), and chlorphenesin and chloropyrazine (5 μM + 25 μM, 5 μM + 50 μM), and the degree of cell migration was confirmed by the same method as in Example 1-2-1. In addition, the synergistic effect of combined treatment was calculated using the combination index (CI) based on the combined treatment concentration of chlorphenesin and chloroquine or chloropyrazine using Compusyn software.

[0090] As a result, the migration of Panc-1 cells was reduced in the groups administered chlorphenesin, chloroquine, and chloropyrazine alone (Figures 38 and 39), and the groups administered chlorphenesin 5μM and chloroquine 10μM in combination and the groups administered chlorphenesin 5μM and chloropyrazine 25μM or more in combination showed an increasing effect (Figure 39). In addition, the migration of cells was reduced in the Aspc-1 cell line from the groups administered chlorphenesin, chloroquine, and chloropyrazine alone (Figures 40 and 41), and all groups administered chlorphenesin and chloroquine or chloropyrazine in combination showed an increasing effect on the reduction of cell migration (Figure 41).

[0091] 2-3. Infiltration analysis In order to confirm that the chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203) of the present invention inhibit the characteristic of cancer cells to invade and metastasize to other sites by opening holes in the thin membrane surrounding the cell tissue or by decomposing the extracellular matrix filling the spaces between the cells, an invasion assay was performed using Matrigel, which mimics the extracellular matrix. Specifically, pancreatic cancer cell lines Panc-1 and MIACaPa2 were suspended in serum-free RPMI and then placed in the upper chamber of a 24-well transwell chamber with a polycarbonate membrane (8.0 μM pore size, Costar) at 1 x 10 cells per well. 5 Matrigel (10 μg / ml) was placed in the lower well, and the cells were treated with control (DMSO), chlorphenesin (5 μM), chloroquine (5 μM, 10 μM, or 25 μM), chloropyrazine (25 μM or 50 μM), chlorphenesin and chloroquine (5 μM + 5 μM, 5 μM + 10 μM, 5 μM + 25 μM), and chlorphenesin and chloropyrazine (5 μM + 25 μM, 5 μM + 50 μM). The cells were then incubated at 37 °C in CO 2 The cells were cultured in an incubator for 18 hours. Then, the cells were fixed with 70% methyl alcohol in PBS for 30 minutes and washed three times with PBS. The cells were stained with hematoxylin (Sigma) for 10 minutes and washed with distilled water. Non-migrated cells were removed from the upper surface of the membrane with a cotton swab. The membrane was excised from the chamber and fixed with Gel Mount (Biomeda, Foster City, CA). Migrated cells (cells attached to the lower surface of the membrane) were counted with a high-power scope randomly selected from the x20 range. In addition, the synergistic effect of combination treatment was calculated through the combination index (CI) according to the combination treatment concentration of chlorphenesin and chloroquine or chloropyrazine using Compusyn software.

[0092] As a result, as shown in Figures 42 and 44, it was confirmed that invasion of Panc-1 and MIACaPa2 cells was suppressed in the groups administered chlorphenesin, chloroquine, or chloropyrazine alone, and both pancreatic cancer cell lines showed a synergistic effect when chlorphenesin was treated with chloroquine or chloropyrazine in combination (Figures 43 and 45). EXAMPLES

[0093] Efficacy against biliary tract cancer confirmed 3-1. Confirmation of cell viability 3-1-1. Confirmation of cell viability by single administration To confirm the effect of single administration of chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203) on the viability of biliary tract cancer cells, the cell viability of the biliary tract cancer cell lines SNU1079 and SNU308 was evaluated by MTT assay (Promega, Ltd.) according to the manufacturer's protocol. Each biliary tract cancer cell line was cultured at 5 × 10 per well. 3 Cells were seeded in a 96-well plate at a density of 0 μM (control group DMSO treatment), 5 μM, 10 μM, 25 μM, 50 μM, 100 μM, 250 μM, 500 μM and 1 mM (1000 μM) with chlorphenesin (OC-201), and chloroquine (OC-202) and chloropyrazine (OC-203) at 0 μM (control group DMSO treatment), 1 μM, 5 μM, 10 μM, 25 μM, 50 μM and 100 μM, respectively, for 24 h, 48 h or 72 h. The cells were then incubated with 5 mg / mL MTT for 4 h. The medium was then removed, and 150 μL of solubilization solution and suspension solution were added, followed by incubation at 30 °C for 4 h. The absorbance of the reaction solution was measured at 570 nm. The cell viability was calculated using the above (Equation 1).

[0094] As a result, as shown in Figures 46 to 48, chlorphenesin showed clear cytotoxicity at 1 mM or more after 48 hours, chloroquine showed cytotoxicity at a concentration of 50 μM or more after 24 hours and at a concentration of 25 μM or more after 48 hours in the case of the SNU1079 cell line, and showed cytotoxicity at a concentration of 50 μM or more after 48 hours in the case of the SNU308 cell line. In the case of the chloropyrazine alone group, no cytotoxicity was shown even at large doses.

[0095] 3-1-2. Confirm cell viability by combined administration To confirm the viability of biliary tract cancer cells upon combined treatment with a combination of chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203), cell viability was evaluated for biliary tract cancer cell lines SNU1079 and SNU308 by MTT assay (Promega, Ltd.) according to the manufacturer's protocol. Each biliary tract cancer cell line was cultured at 5 × 10 per well. 3 The cells were seeded in a 96-well plate at a density of 1000 x 1000 and treated with either the control group (DMSO treatment), chlorphenesin (OC-201) 5 μM treatment, chlorphenesin and chloroquine (OC-202) combined treatment (chlorphenesin 5 μM + chloroquine 1, 5, 10, 25, or 50 μM combined treatment), or chlorphenesin and chloropyrazine (OC-203) combined treatment (chlorphenesin 5 μM + chloropyrazine 1, 5, 10, 25, 50, or 100 μM combined treatment), respectively, and then incubated with 5 mg / mL MTT for 4 hours after 24 h, 48 h, or 72 h. The medium was then removed, and 150 μL of solubilization solution and suspension solution were added, followed by incubation at 30 ° C for 4 hours. The absorbance of the reaction solution was measured at 570 nm. The cell viability was calculated using the above (Equation 1).

[0096] As a result, when chlorphenesin 5 μM and chloroquine 50 μM or more were treated in combination (FIG. 49), and when chlorphenesin 5 μM and chloropyrazine 100 μM were treated in combination (FIG. 50), cytotoxicity was observed in the biliary tract cancer cell line.

[0097] 3-2. Confirmation of cell migration 3-2-1.Chlorphenesin monotherapy The mobility of the biliary tract cancer cell line SNU1079 depending on the treatment concentration of chlorphenesin (OC-201) was confirmed by migration analysis. Specifically, the biliary tract cancer cell line SNU1079 was suspended in serum-free RPMI and then placed in the upper chamber of a 24-well transwell chamber with a polycarbonate membrane (8.0 μM pore size, Costar) at 1 x 10 cells per well. 5 Laminin (10 μg / ml) was placed in the lower well and treated with 5 μM, 10 μM, 25 μM, 50 μM, 100 μM, 250 μM, 500 μM, 1 mM, or 2 mM chlorphenesin. The cells were incubated at 37° C. in CO 2 The cells were cultured in an incubator for 18 hours to allow migration. Afterwards, the cells were fixed with 70% methyl alcohol in PBS for 30 minutes and washed three times with PBS. The cells were stained with hematoxylin (Sigma) for 10 minutes and washed with distilled water. Non-migrating cells were removed from the upper surface of the membrane with a cotton swab. The membrane was excised from the chamber and fixed with Gel Mount (Biomeda, Foster City, CA). Migrated cells (cells attached to the lower surface of the membrane) were counted under a high-powered (x20) scope randomly selected from the control.

[0098] As a result, the migration of biliary tract cancer cells treated with chlorphenesin at 25 μM or more was reduced, and particularly at a concentration of 100 μM or more, the reduction in cell migration was remarkable (FIGS. 51 and 52).

[0099] 3-2-2. Concomitant use When chlorphenesin and chloroquine or chloropyrazine were treated in combination, the mobility of biliary tract cancer cell line SNU1079 was confirmed by a migration analysis method. Specifically, the biliary tract cancer cell line SNU1079 was treated with a control group (DMSO), chlorphenesin (5 μM), chloroquine (5 μM, 10 μM or 25 μM), chloropyrazine (25 μM or 50 μM), chlorphenesin and chloroquine (5 μM + 5 μM, 5 μM + 10 μM, 5 μM + 25 μM), and chlorphenesin and chloropyrazine (5 μM + 25 μM, 5 μM + 50 μM), and the degree of cell mobility was confirmed by the same method as in Example 3-2-1. In addition, the synergistic effect during combination treatment was calculated through the combination index (CI) according to the combination treatment concentration of chlorphenesin and chloroquine or chloropyrazine using Compusyn software.

[0100] As a result, it was shown that when chlorphenesin (5 μM) was co-treated with chloroquine (5 μM or 10 μM), the inhibition of migration of biliary tract cancer cells was synergistically increased (synergistic effect) (FIGS. 53 and 54).

[0101] 3-3. Infiltration analysis In order to confirm that the chlorphenesin (OC-201), chloroquine (OC-202) and chloropyrazine (OC-203) of the present invention inhibit the property of cancer cells to invade and metastasize to other sites when treated alone or in combination, the biliary tract cancer cell line SNU1079 was treated with a control group (DMSO), chlorphenesin (5 μM), chloroquine (5 μM, 10 μM or 25 μM), chloropyrazine (25 μM or 50 μM), chlorphenesin and chloroquine (5 μM + 5 μM, 5 μM + 10 μM, 5 μM + 25 μM), and chlorphenesin and chloropyrazine (5 μM + 25 μM, 5 μM + 50 μM), and an invasion assay was performed as described in Example 2-3. In addition, the synergistic effect of the combined treatment was calculated through the combination index (CI) according to the combined treatment concentration of chlorphenesin and chloroquine or chloropyrazine using Compusyn software.

[0102] As a result, as shown in FIG. 55, it was confirmed that the invasion of bile duct cancer cells was inhibited, and in particular, when chlorphenesin and chloroquine were used in combination, a synergistic effect was observed (FIG. 56). EXAMPLES

[0103] Confirmation of the effect of suppressing cancer metastasis at low concentrations 4-1. Confirmation of the effect of chlorphenesin in inhibiting metastasis of colon cancer 4-1-1. Determination of low concentration without cytotoxicity To determine the non-cytotoxic concentration of chlorphenesin, cell viability was assessed by MTT assay (Promega, Ltd.) according to the manufacturer's protocol. CT26 and HCT-116 cell lines were cultured at 5 × 10 per well. 3Cells were seeded in a 96-well plate at a density of 100 μM, 1 μM, 10 μM, 100 μM, 1 mM, and 10 mM. The cells were then pretreated with or without chlorphenesin (100 ppm, 1 μM, 10 μM, 100 μM, 1 mM, and 10 mM), and then incubated with 5 mg / mL MTT for 4 hours. The medium was then removed, and 150 μL of solubilization solution and suspension solution were added, followed by incubation at 30°C for 4 hours. The absorbance of the reaction solution was measured at 570 nm. The cell viability was calculated using the above formula (1). As can be seen from Figure 57, it was confirmed that there was no cytotoxicity in either CT26 cells or HCT-116 cells in the concentration range of 1 μM to 1 mM.

[0104] Since a chlorphenesin concentration of 1 μM is below the concentration that exhibits muscle relaxant effects and is also a concentration that is not cytotoxic, subsequent experiments were conducted at this concentration to evaluate whether low concentrations of chlorphenesin that have no anticancer activity have an effect of inhibiting cancer cell metastasis.

[0105] 4-1-1. Confirmation of cell migration inhibition 4-1-1-1. Movement analysis Cell migration was assessed using a 24-well transwell chamber with a polycarbonate membrane (8.0 μM pore size, Costar). CT26 and HCT116 colon cancer cells were suspended in serum-free RPMI and then cultured at 1 × 10 per well. 5 Cells were added to the upper chamber. Laminin (10 μg / ml) was placed in the lower well, and the cells were incubated at 37°C in CO 2 The cells were allowed to migrate for 8 h in an incubator. The cells were fixed with 70% methyl alcohol in PBS for 30 min and washed three times with PBS. The cells were stained with hematoxylin (Sigma) for 10 min and washed with distilled water. Non-migrated cells were removed from the upper surface of the membrane with a cotton swab. The membrane was excised from the chamber and fixed with Gel Mount (Biomeda, Foster City, CA). Migrated cells (cells attached to the lower surface of the membrane) were counted under a high-powered (x20) scope randomly selected from the rest of the cells.

[0106] As a result, as shown in FIG. 58, it was confirmed that when treated with a low concentration of chlorphenesin (1 μM), the permeability and migration ability of the cells were significantly reduced compared to the control group.

[0107] 4-1-1-2. Wound healing assay A wound-healing assay was performed to measure cell motility. First, CT26 colon cancer cell lines were seeded into 24-well tissue culture plates at a concentration of 100 mM sodium dodecyl sulfate in RPMI supplemented with 10% FBS, at which point, after 24 h, the monolayer reached 70–80% confluence. A careful and slow scratch was made on the tomography with a new 200 μL yellow pipette tip across the center of the well. The resulting gap distance was made equal to the outer diameter of the end of the tip. After making the scratch, the dish was carefully washed twice in medium to remove detached cells. After 24 h of incubation of the cells in the presence or absence of chlorphenesin (1 μM), pictures of the stained tomography were taken under a microscope.

[0108] As a result, as shown in Figure 59, it was confirmed that the motility of the CT26 cell line was reduced by treatment with chlorphenesin (1 μM). This result also proves that even a low concentration that cannot kill cancer cells can have an effect on suppressing metastasis of cancer cells.

[0109] 4-1-2. Confirmation of in vivo inhibition of cancer metastasis The tumor growth inhibitory effect of chlorphenesin was confirmed in an animal experiment. Specifically, 100 μL of a CT26 cell suspension (1 x 10 7Xenograft animal models were prepared by subcutaneously inoculating cancer cells once with 1000 cells / mL of chlorphenesin to induce tumors. After cancer cell inoculation, chlorphenesin was administered either alone or in combination with fluorouracil (5'FU), an anticancer drug, for 5 weeks. First, 11 animal models were administered 25mg / kg fluorouracil (5 times a week) and 10mg / kg chlorphenesin (3 times a week) intraperitoneally for 5 weeks in the combined administration with fluorouracil. For the chlorphenesin alone administration group, 10 animal models were administered 10mg / kg chlorphenesin intraperitoneally 3 times a week for 5 weeks, and 14 animal models were administered 20mg / kg chlorphenesin orally 5 times a week for 5 weeks. As a negative control, 14 animal models were administered the same volume of PBS as that of chlorphenesin, and as a positive control, 14 animal models were administered 25 mg / kg of fluorouracil, an anticancer drug, alone, five times a week. The body weight of the animal models was measured once a week, and the tumor size was measured once a week from the day of drug administration. After 6 weeks, the animal models were anesthetized with ether, and the tumors and lungs were collected.

[0110] As a result, as shown in Figure 60, it was confirmed that there was no metastasis to the lungs in the chlorphenesin-treated group compared to the control group. Also, as shown in Figure 61, it was confirmed that the tumor size was smaller in the animals treated with 20 mg / kg chlorphenesin compared to the control group. Therefore, it can be seen that chlorphenesin negatively regulates both tumor growth and metastatic ability.

Claims

1. A pharmaceutical composition for preventing or treating pancreatic cancer, comprising chlorphenesin or a pharma- ceutically acceptable salt thereof as an active ingredient.

2. The composition according to claim 1 , wherein the chlorphenesin is represented by the following formula 1: 【Chemistry 1】

3. A pharmaceutical composition for inhibiting metastasis of pancreatic cancer, comprising chlorphenesin or a pharma- ceutically acceptable salt thereof as an active ingredient.

4. An anti-cancer supplement comprising chlorphenesin or a pharma- ceutically acceptable salt thereof as an active ingredient, wherein the cancer is pancreatic cancer.

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