WO2024071464A1 - Anticancer composition comprising maleate metal salt - Google Patents

Anticancer composition comprising maleate metal salt Download PDF

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WO2024071464A1
WO2024071464A1 PCT/KR2022/014491 KR2022014491W WO2024071464A1 WO 2024071464 A1 WO2024071464 A1 WO 2024071464A1 KR 2022014491 W KR2022014491 W KR 2022014491W WO 2024071464 A1 WO2024071464 A1 WO 2024071464A1
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cancer
anticancer
metal salt
maleate
pharmaceutical composition
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PCT/KR2022/014491
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French (fr)
Korean (ko)
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정근영
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(주)바이오솔릭스
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Publication of WO2024071464A1 publication Critical patent/WO2024071464A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A61K31/194—Carboxylic acids, e.g. valproic acid having two or more carboxyl groups, e.g. succinic, maleic or phthalic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the present invention relates to the anticancer use of maleate metal salts. Specifically, the present invention relates to a pharmaceutical composition for preventing or treating cancer containing a maleate metal salt as an active ingredient, and a pharmaceutical composition for adjuvant anticancer use.
  • Malate is a basic metabolite of the TCA cycle (tricarboxylic acid cycle) and is derived from succinate.
  • TCA cycle tricarboxylic acid cycle
  • succinate and fumarate accumulate.
  • hypoxic cancer cells the relative abundance of several TCA cycle intermediates changes and changes in the expression of various enzymes involved in the TCA cycle are also observed.
  • MDH2 malate dehydrogenase 2
  • hypoxic cancer cells try to compensate for this. It mainly relies on biochemical actions utilizing glutamine. Specifically, cancer cells in a hypoxic environment induce an increase in aspartate in the cytoplasm through increased intracellular glutamine supply, and the increased aspartate is converted to oxaloacetate by activating aspartate aminotransferase, and the oxaloacetate level increases.
  • MDH1 malate dehydrogenase 1
  • Insulin is a hormone that affects the growth process of cells. If there is a problem with the insulin secretion function of the pancreas, blood sugar accumulates in the blood, leading to diabetes. Patients with diabetes maintain high blood sugar levels because their insulin control function does not function properly, which leads to continuous damage to DNA and causes cancer. On the other hand, cancer rapidly depletes energy during the growth phase, impairing the insulin regulation function, and cancer-like diabetes occurs due to damaged insulin-secreting cells. As such, cancer and diabetes have an inseparable relationship that works in both directions, so the need for treatment was required.
  • Malic acid enzymes are known to perform an important function in the secretion of insulin, so it is thought that controlling the activity of malic enzymes can solve the problem of insulin secretion or resistance induced by toxicity caused by various anticancer treatments.
  • malic acid has not been used effectively.
  • a drug that can control and treat cancer-related diabetes has not been developed.
  • the present inventor hypothesized that cancer cells could be effectively killed by depleting NAD + , an energy source supplied in excess to cancer cells in a hypoxic environment.
  • NAD + an energy source supplied in excess to cancer cells in a hypoxic environment.
  • the accumulation of intermediates such as succinate and fumarate due to changes in the TCA cycle were known to promote the growth and differentiation of cancer in a hypoxic environment (Nature Communications, 2020, 11: 102)
  • the present inventors used NAD + In order to deplete NAD + by reducing it to NADH, malate was supplied to the cells in the form of a metal salt, thereby accumulating malate within the cells.
  • One object of the present invention is to provide a pharmaceutical composition for preventing or treating cancer containing maleate metal salt as an active ingredient.
  • Another object of the present invention is to provide an adjuvant anti-cancer pharmaceutical composition that improves reactivity to a second anti-cancer agent or to radiation anti-cancer treatment, comprising the maleate metal salt as an active ingredient.
  • Another object of the present invention is to provide a method for preventing or treating cancer, including the step of administering the maleate metal salt to a subject in need.
  • Another object of the present invention is to provide a use of the maleate metal salt for the prevention or treatment of cancer and cancer-related diabetes.
  • Another object of the present invention is to provide a use of the maleate metal salt for the manufacture of a medicament for use in the prevention or treatment of cancer and cancer-related diabetes.
  • One aspect of the present invention for achieving the above object is a pharmaceutical composition for preventing or treating cancer containing a maleate metal salt as an active ingredient.
  • malate metal salt refers to a compound that can be produced or synthesized in the form of a metal ion bonded to maleate. Due to the nature of anticancer drugs that can be administered frequently and in large amounts, metal components that cannot be easily metabolized in the body are excluded from the scope of the present invention. The present inventor found that when cancer cells are treated with malate metal salt, the activity of malic enzymes such as malate dehydrogenase 2 (MDH2), ME2 (malic enzyme 2), and ME3 (malic enzyme 3) is increased, thereby producing NAD + It was expected to kill cancer cells by depleting .
  • MDH2 malate dehydrogenase 2
  • ME2 malic enzyme 2
  • ME3 malic enzyme 3
  • malate metal salt is used as a means to increase the concentration of malate in cancer cells by dissociating malate when administered into cancer cells, and as long as this purpose can be achieved, the type and combination of malate and There is no particular limitation on the ratio of metal ions.
  • the malate metal salt may be calcium malate, calcium dimaleate, zinc malate, magnesium malate, sodium malate, potassium malate, iron malate, chromium malate, copper malate, manganese malate, etc.
  • the pharmaceutical composition may further include DCA (Dichloroacetic acid) as an active ingredient in addition to the maleate metal salt.
  • DCA also called dichloroacetic acid
  • DCA is an acetic acid analogue in which two of the three hydrogen atoms of the methyl group of acetic acid are replaced with chlorine atoms. Since DCA activates pyruvate dehydrogenase, the present inventors predicted that pyruvate increased by MDH2 or malic enzymes in cancer cells could actively participate in the restoration of the TCA cycle by treatment with DCA.
  • treatment of various carcinomas with calcium malate and DCA increased the activity of MDH2 and malic enzymes to a higher level than treatment with malate metal salt (FIGS. 1 and 2), It was confirmed that cancer cells were effectively killed (FIGS. 6 and 12). Therefore, DCA may be additionally included to further increase the anti-cancer effect of the pharmaceutical composition of the present invention.
  • the term “cancer” refers to a disease related to the regulation of cell death, which is caused by excessive cell proliferation when the normal cell death balance is broken.
  • the cancer includes both malignant tumor and benign tumor, and may be solid cancer such as liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, and breast cancer.
  • it may be cancerous diabetes, but the type of cancer of the present invention is not limited to the above examples.
  • the composition for preventing or treating cancer of the present invention has a therapeutic effect on all cancers that can use intracellular NAD + as an energy source.
  • the term “treatment” refers to intervention to alter the natural processes of an individual or cell with a disease, which may be performed during the progression of the pathology or to prevent it.
  • the desired therapeutic effects include preventing the occurrence or recurrence of the disease, alleviating symptoms, reducing all direct or indirect pathological consequences of the disease, preventing metastasis, reducing the rate of disease progression, and alleviating the disease state. Or it includes temporary relief, remission, or improvement of prognosis.
  • the present invention includes all actions to improve the course of cancer by administering a composition containing a maleate metal salt.
  • prevention refers to any action that inhibits or delays the onset of cancer by administering the composition.
  • the content of maleate metal salt included in the pharmaceutical composition is not particularly limited, but may generally be included in an amount of 0.0001 to 2000 mg/kg, specifically 0.001 to 1000 mg/kg.
  • the pharmaceutical composition of the present invention can be administered to individuals who have developed or are at risk of developing cancer.
  • the term “individual” refers to all animals, including humans.
  • the pharmaceutical composition of the present invention can be administered to a subject in a pharmaceutically effective amount.
  • administration refers to introducing the pharmaceutical composition of the present invention into a subject by any appropriate method, and the administration route can be administered through various routes such as oral or parenteral as long as it can reach the target tissue.
  • routes of administration include oral, intramuscular, intravenous, arterial, subcutaneous, peritoneal, pulmonary, and nasal, and may include, but are not limited to, subcutaneous, oral, or intravenous administration.
  • the term “pharmaceutically effective amount” means an amount sufficient to prevent and/or treat cancer with a reasonable benefit/risk ratio applicable to medical use.
  • the appropriate dosage and number of administrations can be selected according to methods known in the art, and the amount and number of administrations of the pharmaceutical composition of the present invention actually administered depend on the type of symptom to be treated, route of administration, gender, health condition, It can be appropriately determined by various factors such as diet, the individual's age, body weight, and the severity of the disease.
  • composition containing the maleate metal salt of the present invention as an active ingredient can be manufactured not only as a pharmaceutical composition but also in the form of a food composition with functionality for improving cancer.
  • the food composition of the present invention may contain additional ingredients that are commonly used in food to improve odor, taste, vision, etc.
  • additional ingredients that are commonly used in food to improve odor, taste, vision, etc.
  • food additives can be added. The additives are selected according to the type of food and used in an appropriate amount.
  • the food composition can be manufactured as a health functional food, where functional food (functional food) is the same term as food for special health use (FoSHU), and has an efficient bioregulatory function in addition to supplying nutrients. It refers to processed medicine and food with high medical effectiveness.
  • the health functional food can be manufactured in various forms such as tablets, capsules, powders, granules, liquids, pills, etc. to achieve useful effects in improving cancer.
  • Another aspect of the present invention is an auxiliary anti-cancer pharmaceutical composition that improves reactivity to a second anti-cancer agent and includes maleate metal salt as an active ingredient.
  • Another aspect of the present invention is a pharmaceutical composition for preventing or treating cancer, comprising a maleate metal salt and a second anticancer agent as active ingredients.
  • the maleate metal salt is as described above.
  • the composition containing the maleate metal salt of the present invention not only has an anti-cancer effect on its own, but also has an excellent effect as an auxiliary anti-cancer pharmaceutical composition for the purpose of enhancing reactivity to a second anti-cancer agent. Therefore, it can be used as a pharmaceutical composition for the prevention or treatment of cancer using the second anticancer agent along with the maleate metal salt as an active ingredient.
  • the term “second anticancer agent” refers to any drug having anticancer activity except for the maleate metal salt of the present invention.
  • the scope of the second anticancer agent is not particularly limited, and a person skilled in the art can select an appropriate type and use it for the purpose of curing, controlling, or alleviating symptoms, depending on the type and progression of cancer.
  • the second anticancer agent may be, for example, a cytotoxic anticancer agent, a targeted anticancer agent, an immune anticancer agent, or a metabolic anticancer agent, but is not limited thereto.
  • a cytotoxic anticancer agent is a drug that exhibits an anticancer effect by attacking cancer cells that divide indiscriminately at a faster rate than normal cells, and its meaning is the same as that commonly used in the technical field to which the present invention pertains.
  • the cytotoxic anticancer agents include alkylating agents, antimetabolites, and natural anticancer agents.
  • the alkylating agent is nitrogen mustard (e.g., cyclophosphamide, chlormethine, uramustine, melphalan, chlorambucil, ifosfamide, bendamustine, etc.), alkyl sulfonate (e.g., busulfan, procarbazine, etc.) ), nitrosoureas (e.g., carmustine, lomustine, streptozocin, etc.), platinum-based alkylating agents (e.g., cisplatin, carboplatin, dicycloplatin, eptaplatin, lobaplatin, myriplatin, daplatin, oxaliplatin, picoplatin, satraplatin, triplatin tetranitrate, etc.), but is not limited thereto. Alkylating agents can cause destruction of cancer cells by binding to the DNA within cancer cells and damaging the DNA structure.
  • nitrogen mustard e.g., cyclophosphamide, chlormethin
  • the antimetabolites include pyrimidine derivatives (e.g., 5-fluorouracil, capecitabine, cystarabine, gemcitabine, fludarabine, etc.), folate derivatives (e.g., methotrexate, pemetrexed, etc.), and purine derivatives (e.g., , mercaptopurine, etc.), but is not limited thereto.
  • Antimetabolites can induce cancer cell death by inhibiting metabolism required for DNA replication and cell survival.
  • the natural anticancer agents include topoisomerase inhibitors (e.g., camptothecin, epipodophyllotoxin, taxane-based drugs (docitaxel, paclitaxel)), antibiotics (e.g., dactinomycin, doxorubicin, daunorubicin, and mitomycin). , phleomycin, idarubicin, mitoxantrone HCl, etc.), but is not limited thereto.
  • topoisomerase inhibitors e.g., camptothecin, epipodophyllotoxin, taxane-based drugs (docitaxel, paclitaxel)
  • antibiotics e.g., dactinomycin, doxorubicin, daunorubicin, and mitomycin.
  • phleomycin, idarubicin, mitoxantrone HCl, etc. but is not limited thereto.
  • the targeted anticancer agent is an anticancer agent that induces death of cancer cells by inhibiting target proteins (receptors or enzymes) involved in cancer growth, and its meaning is the same as that commonly used in the technical field to which the present invention pertains.
  • the cytotoxic anticancer agents include small molecule compounds and monoclonal antibodies that inhibit target proteins (tyrosine kinases, etc.).
  • the targeted anticancer agent may be an antibody or tyrosine kinase inhibitor targeting one or more targets selected from the group consisting of VEGF-A, HER2, and EGFR.
  • the targeted anti-cancer agent that can be administered in combination with maleate metal salt is a VEGF-A inhibitor.
  • the VEGF-A inhibitor is monoclonal antibodies such as bevacizumab, ranibizumab, aflibercept, ramucirumab, sunitinib, and pazopar. Low molecular weight compounds such as pazopanib, sorafenib, and axitinib may be included, but are not limited thereto.
  • the targeted anti-cancer agent that can be administered in combination with the maleate metal salt is an EGFR inhibitor.
  • the EGFR inhibitor is osimertinib, gefitinib, erlotinib, afatinib, brigatinib, icotinib, vandetanib ( monoclonal antibodies such as cetuximab, panitumumab, zalutumumab, nimotuzumab, and matuzumab, along with small molecule compounds such as vandetanib, It is not limited to this.
  • the targeted anti-cancer agent that can be administered in combination with the maleate metal salt is a HER2 inhibitor such as trastuzumab, pertuzumab, lapatinib, neratinib, or It may be, but is not limited to, afatinib.
  • the targeted anticancer agent of the present invention also includes Bcr-Abl targeting anticancer agents such as imatinib, dasatinib, and nilotinib; Src-targeting anticancer drugs such as bosutinib; JAK-targeting anticancer drugs such as lestaurtinib, ruxolitinib, and pacritinib; MAP2 kinase targeting anticancer drugs such as cobimethinib, selumetinib, trametinib, and binimetinib; It includes MEL4-ALK targeting anticancer drugs such as ceritibin and crizotinib, and is not particularly limited to the type.
  • Bcr-Abl targeting anticancer agents such as imatinib, dasatinib, and nilotinib
  • Src-targeting anticancer drugs such as bosutinib
  • JAK-targeting anticancer drugs such as lest
  • the second anticancer agent of the present invention may be a combination of one or more targeted anticancer agents or immune anticancer agents, and they may be administered simultaneously or at different times.
  • the second anticancer drug sorafenib, trastuzumab (Herceptin), or osimertinib
  • osimertinib the second anticancer drug or It was confirmed that cancer cells were killed significantly compared to treatment with maleate metal salt (or addition of DCA) alone (FIG. 7).
  • the second anticancer agent of the present invention may be an immunological anticancer agent such as CAR-T, CAR-NK, immune checkpoint inhibitor, or anticancer vaccine, but is not limited thereto.
  • Immune cells experience energy depletion due to the rapid glycolysis of cancer cells in the hypoxic environment within tumor tissue, and this environment is one of the important causes of suppressed immune activity in relation to anticancer treatment.
  • T cells and NK cells were treated with maleate metal salt together with IL-2 or IL-12, the activity of immune cells was significantly improved compared to treatment with cytokines alone ( Figure 9 ). Therefore, maleate metal salt can have a synergistic effect even when used as a second anticancer drug with immunological anticancer drugs such as CAR-T, CAR-NK, immune checkpoint inhibitors, and anticancer vaccines.
  • the second anti-cancer agent of the present invention can be selected according to the type and progression of cancer, appropriate for curing, controlling, and alleviating symptoms of cancer.
  • the composition containing the maleate metal salt of the present invention has the effect of promoting depletion of NAD + in cancer cells and enhancing reactivity to the second anticancer agent, so it can be used as an anticancer adjuvant, so it can be used as an anticancer adjuvant, so it can be used as an anticancer adjuvant.
  • the type is not particularly limited.
  • the cancer may be liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, breast cancer, and cancerous diabetes, but is not limited thereto.
  • composition containing the maleate metal salt of the present invention can be co-administered in a form that exists independently of the second anticancer agent, as well as forming a physical/chemical bond with the second anticancer agent by any known method depending on the purpose. It can be administered at.
  • the composition containing the maleate metal salt may be used in a state directly bound to the second anticancer agent, or may be used in a state connected to the second anticancer agent through a known linker, and the composition containing the maleate metal salt of the present invention may be used as the second anticancer agent. 2.
  • There are no particular restrictions on the method of application as long as it works together with anticancer drugs to exhibit increased anticancer effects.
  • Another aspect of the present invention is an adjuvant anti-cancer pharmaceutical composition that improves responsiveness to radiation anti-cancer treatment, comprising the maleate metal salt as an active ingredient.
  • the maleate metal salt is as described above.
  • the term “radiation anti-cancer treatment” refers to the treatment of irradiating cancer cells or tumor tissue with radiation for the purpose of killing cancer cells. In general, it is a standard treatment for controlling unresectable or inoperable tumors or tumor metastases, and is based on the principle that radiation delivered to the target site causes the death of regenerative cells.
  • the radiation anti-cancer treatment may be ionizing radiation therapy, electromagnetic radiation therapy, brachytherapy, or external beam radiation therapy, but is not limited thereto.
  • composition containing maleate metal salt according to the present invention exhibits a synergistic anti-cancer effect when used in combination with radiotherapy, so it can be usefully used as an anti-cancer adjuvant for radiotherapy or as a radiosensitizer to improve radiosensitivity, and is applicable to cancer.
  • the type is not particularly limited.
  • the cancer may be liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, breast cancer, and cancerous diabetes, but is not limited thereto.
  • the composition for preventing or treating cancer containing malate metal salt as an active ingredient of the present invention has excellent anticancer effects by depleting NAD + , an energy source in cancer cells, and increasing immune activity, and increases insulin secretion by increasing malic acid activity. It can solve the problem of decreased insulin secretion or resistance caused by toxicity from various anticancer treatments. In addition, when administered in combination with other anticancer drugs or during radiotherapy, it improves responsiveness to anticancer drugs and radiotherapy, making it very effective as an anticancer adjuvant.
  • Figure 1 is a graph comparing the levels of MDH2, ME2, and ME3 when cancer cells are treated with CaMal, CaMal 2 , or CaMal-DCA.
  • Figure 2 is a graph comparing NAD + levels when cancer cells are treated with CaMal, CaMal 2 , or CaMal-DCA.
  • Figure 3 is a graph comparing the insulin secretion level according to the presence or absence of glucose in the pancreatic beta cells in the CaMal or CaMal 2 treatment group.
  • Figure 4 is a graph comparing the survival rate of cancer cells according to CaMal treatment concentration.
  • Figure 5 is a graph comparing the survival rate of cancer cells according to CaMal 2 treatment concentration.
  • Figure 6 is a graph comparing the survival rate of cancer cells according to CaMal-DCA treatment concentration.
  • Figure 7 is a graph comparing the survival rate of cells when the second anticancer drug and CaMal, CaMal 2 , or CaMal-DCA were administered alone or in combination to cancer cells.
  • Figure 8 is a graph comparing the survival rate of cancer cells treated with CaMal, CaMal 2 , or CaMal-DCA and radiation alone or in combination.
  • Figure 9 is a graph comparing the levels of activated T cells and NK cells by treatment with IL-2 alone or in combination with IL-2 or IL-12 and CaMal 2 .
  • Figure 10 is a graph showing the tumor growth inhibition effect of subcutaneous administration of CaMal in pancreatic cancer transplant mice.
  • Figure 11 is a graph showing the tumor growth inhibition effect of oral administration of CaMal 2 in liver cancer transplant mice.
  • Figure 12 is a graph showing the tumor growth inhibition effect of intravenous administration of CaMal-DCA in breast cancer transplant mice.
  • liver cancer cells, breast cancer cells, and lung cancer cells were treated with CaMal, CaMal 2 , or CaMal-DCA, respectively, the amounts of MDH2, ME2, and ME3 were confirmed to significantly increase compared to the untreated control group ( Figure 1; ** p ⁇ 0.001 ).
  • liver cancer HepG2
  • breast cancer MDA-MB-231
  • lung cancer H1975
  • 5mM CaMal 2.5mM CaMal 2
  • 1mM CaMal-DCA 1mM CaMal-DCA
  • liver cancer cells, breast cancer cells, and lung cancer cells were treated with CaMal, CaMal 2 , or CaMal-DCA, respectively, the level of NAD + was confirmed to be significantly reduced compared to the untreated control group ( Figure 2; ** p ⁇ 0.001).
  • 3 x 103 colon cancer (HCT-116) or brain cancer (U-87MG) cells were cultured for 24 hours, then treated with 0.5, 1, 2.5, or 5 mM CaMal and cultured again for 96 hours. Afterwards, 10 ⁇ l of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 ⁇ l of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
  • kidney cancer Caki-1
  • pancreatic cancer Aspc-1
  • the survival rate of cancer cells decreased in a CaMal 2 concentration-dependent manner, and this result was found to be significant (Figure 5; * * p ⁇ 0.001 vs 0.5, 1 mM groups. ## p ⁇ 0.001 vs 0.5, 1, 2.5 mM groups).
  • 3 x 10 liver (HepG2), lung (H1975), pancreatic (Aspc-1), or breast (MDA-MB-231) cells were cultured for 24 h and then incubated with 0.5, 1, 2.5, or 5 mM CaMal-DCA. The cells were treated and cultured again for 96 hours. Afterwards, 10 ⁇ l of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 ⁇ l of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
  • liver cancer HepG2
  • lung cancer H1975)
  • pancreatic cancer Aspc-1
  • breast cancer MDA-MB-231
  • Figure 6 ** p ⁇ 0.001 vs 0.5 group, ## p ⁇ 0.001 vs 0.5, 1 mM groups; $$ p ⁇ 0.001 vs 0.5, 1, 2.5 mM groups).
  • liver cancer HepG2
  • breast cancer MDA-MB-231
  • lung cancer H1975 cells
  • liver cancer cells were treated with 1 ⁇ M sorafenib and 5 mM CaMal alone or in combination
  • breast cancer cells were treated with 2.5 ⁇ M Herceptin and 2.5 mM CaMal 2 alone or in combination
  • lung cancer cells were treated with 1 nM Oshi.
  • Osimertinib and 1mM CaMal-DCA were treated alone or in combination and cultured for another 96 hours.
  • maleate metal salt is an existing anticancer drug known to have excellent anticancer effects even when administered alone. It showed an effect equal to or better than that of other anticancer drugs, and in particular, when each anticancer drug and maleate metal salt were treated in combination, a synergistic effect was observed and almost all cancer cells were confirmed to be killed.
  • 3 x 103 liver cancer (HepG2), breast cancer (MDA-MB-231), or lung cancer (H1975) cells were cultured for 24 hours. Afterwards, each cell was treated alone or in combination with a dose of 2 Gy of radiation and 5 mM CaMal, 2.5 Mm CaMal 2 or 1 mM CaMal-DCA and cultured for 48 hours. Afterwards, 10 ⁇ l of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 ⁇ l of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
  • DMSO 2-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent
  • liver cancer HepG2
  • breast cancer MDA-MB-231
  • lung cancer H1975
  • maleate metal salt killed cancer cells more effectively than 2 Gy of radiation.
  • treatment with maleate metal salt in combination with radiation showed increased anticancer efficacy.
  • T cells were isolated from the spleen and lymphoid tissue of C57bl/6 mice using a mouse CD8 isolation kit (ThermoFisher). The T cells were treated with 20 units of IL-2 alone or in combination with 2.5mM CaMal 2 , and then CD25 + and CD69 + cells were separated by FACS to quantify activated T cells.
  • NK cells were isolated from the spleen of c57bl/6 mice using a mouse NK cell isolation kit (ThermoFisher).
  • the NK cells were treated with 2 ng/ml IL-12 alone or in combination with 2.5 mM CaMal 2 , and CD16 - and CD56 + cells were sorted by FACS to quantify activated NK cells.
  • the number of activated T cells in the group treated simultaneously with IL-2 and malate metal salt significantly increased compared to the group treated with IL-2 alone (left side of Figure 9), and the number of activated T cells was significantly increased compared to the group treated with IL-2 alone (left side of Figure 9).
  • the group treated simultaneously with late metal salts was confirmed to have a significant increase in the number of activated NK cells compared to the group treated with IL-12 alone (right side of Figure 9; ** p ⁇ 0.001 vs low dose IL-2 or IL-12 group).
  • liver cancer HepG2
  • CaMal 2 CaMal 2
  • mice transplanted with breast cancer (MDA-MB-231) cells were treated with CaMal-DCA, a decrease in tumor size was clearly observed with the naked eye compared to the untreated control group (left side of Figure 12), and this result was statistically significant. (right side of Figure 12; ** p ⁇ 0.001 vs Control).

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Abstract

The present invention relates to anticancer uses of a maleate metal salt. Particularly, the present invention relates to a pharmaceutical composition for preventing or treating cancer and an anticancer adjuvant pharmaceutical composition, the compositions comprising a maleate metal salt as an active ingredient. The composition for preventing or treating cancer, comprising a maleate metal salt as an active ingredient, of the present invention has excellent anticancer effects by exhausting NAD+, which is an energy source in cancer cells, and increasing immunoactivity, increases insulin secretion through an increase in malic acid activity, and thus solves problems of decreased insulin secretion or resistance induced by toxicity from various anticancer treatments. In addition, when co-administered during the administration of other anticancer drugs or anticancer radiotherapy, the present invention increases responsiveness to the anticancer drugs and radiotherapy and thus also has very excellent effects as an anticancer adjuvant.

Description

말레이트 금속염을 포함하는 항암용 조성물Anticancer composition containing maleate metal salt
본 발명은 말레이트 금속염의 항암 용도에 관한 것이다. 구체적으로, 본 발명은 말레이트 금속염을 유효성분으로 포함하는 암의 예방 또는 치료용 약학적 조성물, 및 항암 보조용 약학적 조성물에 관한 것이다.The present invention relates to the anticancer use of maleate metal salts. Specifically, the present invention relates to a pharmaceutical composition for preventing or treating cancer containing a maleate metal salt as an active ingredient, and a pharmaceutical composition for adjuvant anticancer use.
말레이트는 TCA 주기 (tricarboxylic acid cycle)의 기본 대사 산물로서 석시네이트에서 파생된다. 그러나 생체 내에서 암 세포가 성장하게 되면 종양 조직 내 미세환경은 저산소 상태가 되며, 저산소 상태의 암 세포는 TCA 주기가 많이 변화되며 석시네이트와 푸말레이트가 축적된다. 즉, 저산소 암세포에서는 여러 TCA 주기 중간체의 상대적 존재비가 변화하며 TCA 주기에 관여하는 다양한 효소의 발현 변화도 관찰된다.Malate is a basic metabolite of the TCA cycle (tricarboxylic acid cycle) and is derived from succinate. However, when cancer cells grow in vivo, the microenvironment within the tumor tissue becomes hypoxic. In hypoxic cancer cells, the TCA cycle changes significantly and succinate and fumarate accumulate. In other words, in hypoxic cancer cells, the relative abundance of several TCA cycle intermediates changes and changes in the expression of various enzymes involved in the TCA cycle are also observed.
TCA 주기에서 파생되는 중간체의 활용성이 변경됨에 따라 암 세포 내 말레이트의 수준이 감소하게 되면 말레이트 탈수소효소 2 (malate dehydrogenase 2, MDH2)의 활성이 저하되고, 저산소 암 세포는 이를 보완하기 위해 글루타민을 활용한 생화학적 작용에 주로 의존하게 된다. 구체적으로, 저산소 환경의 암 세포는 세포 내 글루타민 공급 증가를 통해 세포질 내 아스파테이트 증가를 유도하며, 증가된 아스파테이트는 아스파테이트아미노전달효소를 활성화하여 옥살로아세테이트로 변환되고, 옥살로아세테이트 수준이 세포질에서 증가하면 옥살로아세테이트를 말레이트로 환원시키고 NADH를 NAD+로 산화시키는 말레이트 탈수소효소 1 (malate dehydrogenase 1, MDH1)에 매우 의존적이게 된다. 결과적으로 MDH1의 활성화는 암 세포에서 지속적인 에너지원으로 사용되는 NAD+를 공급하여 암 세포 생존에 유리한 종양 미세환경을 조성하게 된다. When the level of malate in cancer cells decreases as the availability of intermediates derived from the TCA cycle changes, the activity of malate dehydrogenase 2 (MDH2) decreases, and hypoxic cancer cells try to compensate for this. It mainly relies on biochemical actions utilizing glutamine. Specifically, cancer cells in a hypoxic environment induce an increase in aspartate in the cytoplasm through increased intracellular glutamine supply, and the increased aspartate is converted to oxaloacetate by activating aspartate aminotransferase, and the oxaloacetate level increases. When increased in the cytoplasm, it becomes highly dependent on malate dehydrogenase 1 (MDH1), which reduces oxaloacetate to malate and oxidizes NADH to NAD + . As a result, activation of MDH1 supplies NAD + , which is used as a continuous energy source in cancer cells, creating a tumor microenvironment favorable for cancer cell survival.
한편, 암과 당뇨는 밀접한 관련을 가지고 있는 것으로 알려져 있다. 인슐린은 세포의 성장 과정에 영향을 끼치는 호르몬으로, 췌장의 인슐린 분비 기능에 문제가 생기면 혈액에 혈당이 쌓이게 되고, 당뇨병에 이르게 된다. 당뇨병을 가진 환자는 인슐린 조절 기능이 제대로 작동하지 않아 높은 혈당을 유지하게 되고, 이는 DNA의 지속적인 손상으로 이어지며, 암을 발생시키는 원인이 된다. 다른 한편으로, 암은 성장 단계에서 에너지를 빠르게 고갈시켜 인슐린 조절 기능을 손상시키는데, 손상된 인슐린 분비 세포로 인하여 암성 당뇨병이 발생하게 된다. 이처럼 암과 당뇨는 양방향으로 작동하는 불가분적 관계를 지니고 있어 치료의 필요성이 요구되고 있었다. 말산 효소들은 인슐린의 분비에 중요한 기능을 수행하는 것으로 알려져 있어, 말산 효소의 활성 조절이 다양한 항암 치료에 의한 독성으로 유도되는 인슐린 분비나 저항성의 문제를 해결할 수 있을 것으로 생각되나, 현재까지 말산을 효과적으로 조절하여 암성 당뇨를 치료할 수 있는 약물은 개발되지 않은 실정이다. Meanwhile, cancer and diabetes are known to be closely related. Insulin is a hormone that affects the growth process of cells. If there is a problem with the insulin secretion function of the pancreas, blood sugar accumulates in the blood, leading to diabetes. Patients with diabetes maintain high blood sugar levels because their insulin control function does not function properly, which leads to continuous damage to DNA and causes cancer. On the other hand, cancer rapidly depletes energy during the growth phase, impairing the insulin regulation function, and cancer-like diabetes occurs due to damaged insulin-secreting cells. As such, cancer and diabetes have an inseparable relationship that works in both directions, so the need for treatment was required. Malic acid enzymes are known to perform an important function in the secretion of insulin, so it is thought that controlling the activity of malic enzymes can solve the problem of insulin secretion or resistance induced by toxicity caused by various anticancer treatments. However, to date, malic acid has not been used effectively. A drug that can control and treat cancer-related diabetes has not been developed.
상술한 배경 하에, 본 발명자는 저산소 환경의 암 세포에 과량으로 공급되는 에너지원인 NAD+를 소진시키면 암 세포를 효과적으로 사멸시킬 수 있을 것이라고 가설을 설정하였다. 앞서, TCA 주기의 변화로 인한 중간체인 석시네이트, 푸마레이트 등의 축적은 저산소 환경의 암의 성장과 분화를 촉진하는 것으로 알려져 있었으나 (Nature Communications, 2020, 11: 102), 본 발명자는 NAD+를 NADH로 환원시킴으로써 NAD+를 고갈시키고자 말레이트를 금속염 형태로 세포에 공급하여 오히려 세포 내 말레이트를 축적시키고자 하였다. 그 결과, 놀랍게도 말레이트 금속염 처리로 말산 효소들을 활성시킴으로써 인슐린 분비가 증가되었고, 암 세포에서 MDH2 및 말산 효소들의 활성을 유도하여 TCA 주기가 복구되었으며, 에너지원인 NAD+가 고갈되어, 말레이트 금속염이 우수한 항암 효과를 가짐을 확인함으로써 본 발명을 완성하였다.Under the above-described background, the present inventor hypothesized that cancer cells could be effectively killed by depleting NAD + , an energy source supplied in excess to cancer cells in a hypoxic environment. Previously, the accumulation of intermediates such as succinate and fumarate due to changes in the TCA cycle were known to promote the growth and differentiation of cancer in a hypoxic environment (Nature Communications, 2020, 11: 102), but the present inventors used NAD + In order to deplete NAD + by reducing it to NADH, malate was supplied to the cells in the form of a metal salt, thereby accumulating malate within the cells. As a result, surprisingly, insulin secretion was increased by activating malic enzymes through malate metal salt treatment, the TCA cycle was restored by inducing the activity of MDH2 and malic enzymes in cancer cells, and NAD + , an energy source, was depleted, resulting in malate metal salt. The present invention was completed by confirming that it has excellent anticancer effect.
본 발명의 하나의 목적은, 말레이트 금속염을 유효성분으로 포함하는 암의 예방 또는 치료용 약학적 조성물을 제공하는 것이다.One object of the present invention is to provide a pharmaceutical composition for preventing or treating cancer containing maleate metal salt as an active ingredient.
본 발명의 다른 하나의 목적은, 상기 말레이트 금속염을 유효성분으로 포함하는, 제2항암제에 대한 반응성을 증진시키거나 방사선 항암 치료에 대한 반응성을 증진시키는 항암 보조용 약학적 조성물을 제공하는 것이다.Another object of the present invention is to provide an adjuvant anti-cancer pharmaceutical composition that improves reactivity to a second anti-cancer agent or to radiation anti-cancer treatment, comprising the maleate metal salt as an active ingredient.
본 발명의 또 다른 하나의 목적은 상기 말레이트 금속염을 필요로 하는 개체에 투여하는 단계를 포함하는 암의 예방 또는 치료방법을 제공하는 것이다.Another object of the present invention is to provide a method for preventing or treating cancer, including the step of administering the maleate metal salt to a subject in need.
본 발명의 또 다른 하나의 목적은 암 및 암성 당뇨의 예방 또는 치료를 위한 상기 말레이트 금속염의 용도를 제공하는 것이다.Another object of the present invention is to provide a use of the maleate metal salt for the prevention or treatment of cancer and cancer-related diabetes.
본 발명의 또 다른 하나의 목적은 암 및 암성 당뇨의 예방 또는 치료에 사용하기 위한 약제(medicament)의 제조에 사용하기 위한, 상기 말레이트 금속염의 용도를 제공하는 것이다.Another object of the present invention is to provide a use of the maleate metal salt for the manufacture of a medicament for use in the prevention or treatment of cancer and cancer-related diabetes.
이를 구체적으로 설명하면 다음과 같다. 한편, 본 발명에서 개시된 각각의 설명 및 실시형태는 각각의 다른 설명 및 실시 형태에도 적용될 수 있다. 즉, 본 발명에서 개시된 다양한 요소들의 모든 조합이 본 발명의 범주에 속한다. 또한, 하기 기술된 구체적인 서술에 의하여 본 발명의 범주가 제한된다고 볼 수 없다.This is explained in detail as follows. Meanwhile, each description and embodiment disclosed in the present invention may also be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present invention fall within the scope of the present invention. Additionally, the scope of the present invention cannot be considered limited by the specific description described below.
상기 목적을 달성하기 위한 본 발명의 하나의 양태는, 말레이트 금속염을 유효성분으로 포함하는 암의 예방 또는 치료용 약학적 조성물이다. One aspect of the present invention for achieving the above object is a pharmaceutical composition for preventing or treating cancer containing a maleate metal salt as an active ingredient.
본 발명에서, 용어 "말레이트 금속염"은 말레이트에 금속이온이 결합된 형태로 생성 또는 합성될 수 있는 화합물을 말한다. 빈번하게 다량으로 투여될 수 있는 항암제의 특성상 체내에서 용이하게 대사될 수 없는 금속 성분은 본 발명의 범주에서 제외된다. 본 발명자는 암 세포에 말레이트 금속염을 처리할 경우 말레이트 탈수소효소 2 (malate dehydrogenase 2, MDH2) 및 ME2 (malic enzyme 2), ME3 (malic enzyme 3)와 같은 말산 효소들의 활성을 증가시켜 NAD+를 고갈시킴으로써 암 세포를 사멸시킬 것으로 예상하였다. 본 발명의 구체적인 일 실시예에서는 다양한 암 세포주에 칼슘 말레이트 (calcium malate) 또는 칼슘 디말레이트 (calcium dimalate)를 처리한 결과 미처리 대조군에 비해 MDH2, ME2, ME3의 활성이 현저히 증가하는 것을 확인하였으며 (도 1), 놀랍게도 암 세포 내 NAD+가 급격히 고갈되어 (도 2) 결과적으로 암 세포가 사멸되는 것을 확인하였다 (도 4, 도 5, 도 10 및 도 11). 따라서 본 발명의 말레이트 금속염을 포함하는 약학적 조성물은 암의 예방 또는 치료용도로 매우 유용하게 사용될 수 있다.In the present invention, the term “malate metal salt” refers to a compound that can be produced or synthesized in the form of a metal ion bonded to maleate. Due to the nature of anticancer drugs that can be administered frequently and in large amounts, metal components that cannot be easily metabolized in the body are excluded from the scope of the present invention. The present inventor found that when cancer cells are treated with malate metal salt, the activity of malic enzymes such as malate dehydrogenase 2 (MDH2), ME2 (malic enzyme 2), and ME3 (malic enzyme 3) is increased, thereby producing NAD + It was expected to kill cancer cells by depleting . In a specific example of the present invention, as a result of treating various cancer cell lines with calcium malate or calcium dimalate, it was confirmed that the activities of MDH2, ME2, and ME3 were significantly increased compared to the untreated control group ( Figure 1), surprisingly, it was confirmed that NAD + in cancer cells was rapidly depleted (Figure 2), resulting in the death of cancer cells (Figures 4, 5, 10, and 11). Therefore, the pharmaceutical composition containing the maleate metal salt of the present invention can be very useful for preventing or treating cancer.
본 발명에서 “말레이트 금속염”은 암 세포 내에 투여될 경우 말레이트가 해리되어 암 세포 내 말레이트의 농도를 증가시키는 수단으로 사용되며, 이와 같은 목적을 달성시킬 수 있는 한 그 종류 및 말레이트와 금속이온의 비율에 특별히 제한되지 않는다. 예컨대, 상기 말레이트 금속염은 칼슘 말레이트, 칼슘 디말레이트, 아연 말레이트, 마그네슘 말레이트, 나트륨 말레이트, 칼륨 말레이트, 철 말레이트, 크롬 말레이트, 구리 말레이트, 망간 말레이트 등일 수 있다. In the present invention, “malate metal salt” is used as a means to increase the concentration of malate in cancer cells by dissociating malate when administered into cancer cells, and as long as this purpose can be achieved, the type and combination of malate and There is no particular limitation on the ratio of metal ions. For example, the malate metal salt may be calcium malate, calcium dimaleate, zinc malate, magnesium malate, sodium malate, potassium malate, iron malate, chromium malate, copper malate, manganese malate, etc.
일 실시양태로, 상기 약학적 조성물은 말레이트 금속염 외에 유효성분으로 DCA (Dichloroacetic acid)를 추가로 포함할 수 있다. DCA는 디클로로아세트산으로도 명명되는 아세트산 유사체로서, 아세트산의 메틸 그룹의 3 개의 수소 원자 중 2 개가 염소 원자로 대체된 물질이다. DCA는 피루브산 탈수소효소 (pyruvate dehydrogenase)를 활성화시키기 때문에, 본 발명자는 암 세포 내에서 MDH2나 말산 효소들에 의해 증가된 피루브산이 DCA의 처리로 TCA 주기의 복구에 적극 참여할 수 있을 것으로 예상하였다. 본 발명의 구체적 일 실시예에서는 칼슘 말레이트와 DCA를 다양한 암종에 처리한 결과, 말레이트 금속염을 처리한 것보다 더 높은 수준으로 MDH2 및 말산 효소들의 활성을 증가시키고 (도 1 및 도 2), 암 세포를 효과적으로 사멸시키는 것을 확인하였다 (도 6 및 도 12). 따라서 DCA는 본 발명의 약학적 조성물의 항암 효과를 더욱 증대시키기 위해 추가적으로 포함될 수 있다.In one embodiment, the pharmaceutical composition may further include DCA (Dichloroacetic acid) as an active ingredient in addition to the maleate metal salt. DCA, also called dichloroacetic acid, is an acetic acid analogue in which two of the three hydrogen atoms of the methyl group of acetic acid are replaced with chlorine atoms. Since DCA activates pyruvate dehydrogenase, the present inventors predicted that pyruvate increased by MDH2 or malic enzymes in cancer cells could actively participate in the restoration of the TCA cycle by treatment with DCA. In one specific embodiment of the present invention, treatment of various carcinomas with calcium malate and DCA increased the activity of MDH2 and malic enzymes to a higher level than treatment with malate metal salt (FIGS. 1 and 2), It was confirmed that cancer cells were effectively killed (FIGS. 6 and 12). Therefore, DCA may be additionally included to further increase the anti-cancer effect of the pharmaceutical composition of the present invention.
본 발명에서 용어 "암"은 세포의 사멸 조절과 관련된 질병으로서, 정상적인 세포 사멸 균형이 깨지는 경우 세포가 과다 증식하게 됨으로써 생기는 질병을 말한다. 본 발명에서 상기 암은 악성 종양 (malignant tumor) 및 양성 종양 (benign tumor)을 모두 포함하며, 예를 들어 간암, 폐암, 대장암, 뇌암, 신장암, 췌장암 및 유방암과 같은 고형암일 수 있고, 예를 들어 암성 당뇨일 수 있으나, 상기 예들에 의해 본 발명의 암의 종류가 한정되는 것은 아니다. 본 발명의 암의 예방 또는 치료용 조성물은 세포 내 NAD+를 에너지원으로 사용할 수 있는 모든 암에 치료 효과를 가진다.In the present invention, the term “cancer” refers to a disease related to the regulation of cell death, which is caused by excessive cell proliferation when the normal cell death balance is broken. In the present invention, the cancer includes both malignant tumor and benign tumor, and may be solid cancer such as liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, and breast cancer. For example, it may be cancerous diabetes, but the type of cancer of the present invention is not limited to the above examples. The composition for preventing or treating cancer of the present invention has a therapeutic effect on all cancers that can use intracellular NAD + as an energy source.
본 발명에서 용어, "치료"는 질병을 갖는 개개인 또는 세포의 천연 과정을 변경시키기 위해 개입하는 것을 지칭하고, 이는 병리 상태가 진행되는 동안 또는 이를 예방하기 위해 수행될 수 있다. 목적하는 치료 효과에는 질병의 발생 또는 재발을 예방하고, 증상을 완화시키며, 질병에 따른 모든 직접 또는 간접적인 병리학적 결과를 저하시키며, 전이를 예방하고, 질병 진행 속도를 감소시키며, 질병 상태를 경감 또는 일시적 완화시키며, 차도시키거나 예후를 개선시키는 것이 포함된다. 특히, 본 발명에서는 말레이트 금속염을 포함하는 조성물의 투여로 암의 경과를 호전시키는 모든 행위를 포함한다. 또한, 용어, "예방"은 상기 조성물의 투여로 암의 발병을 억제 또는 지연시키는 모든 행위를 말한다.In the present invention, the term “treatment” refers to intervention to alter the natural processes of an individual or cell with a disease, which may be performed during the progression of the pathology or to prevent it. The desired therapeutic effects include preventing the occurrence or recurrence of the disease, alleviating symptoms, reducing all direct or indirect pathological consequences of the disease, preventing metastasis, reducing the rate of disease progression, and alleviating the disease state. Or it includes temporary relief, remission, or improvement of prognosis. In particular, the present invention includes all actions to improve the course of cancer by administering a composition containing a maleate metal salt. Additionally, the term “prevention” refers to any action that inhibits or delays the onset of cancer by administering the composition.
상기 약학적 조성물에 포함되는 말레이트 금속염의 함량은 특별히 이에 제한되지 않으나, 일반적으로 0.0001 내지 2000 mg/kg의 양, 구체적으로는 0.001 내지 1000 mg/kg의 양이 포함될 수 있다.The content of maleate metal salt included in the pharmaceutical composition is not particularly limited, but may generally be included in an amount of 0.0001 to 2000 mg/kg, specifically 0.001 to 1000 mg/kg.
본 발명의 약학적 조성물은 암이 발병했거나 발병할 위험을 가지는 개체에 투여될 수 있다. 본 발명에서 용어, "개체"는 인간을 포함한 모든 동물을 의미한다. The pharmaceutical composition of the present invention can be administered to individuals who have developed or are at risk of developing cancer. In the present invention, the term “individual” refers to all animals, including humans.
본 발명의 약학적 조성물은 대상 개체에 약학적으로 유효한 양으로 투여될 수 있다. 본 발명에서 용어, "투여"는 어떠한 적절한 방법으로 대상 개체에 본 발명의 약학적 조성물을 도입하는 것을 말하며, 투여 경로는 목적 조직에 도달할 수 있는 한 경구 또는 비경구의 다양한 경로를 통하여 투여될 수 있다. 투여 경로의 예로는 경구, 근육, 정맥, 동맥, 피하, 복강, 폐, 및 비강이 포함되며, 예컨대 피하, 경구 또는 정맥 내로 투여되는 것일 수 있으나 이에 제한되는 것은 아니다. The pharmaceutical composition of the present invention can be administered to a subject in a pharmaceutically effective amount. In the present invention, the term "administration" refers to introducing the pharmaceutical composition of the present invention into a subject by any appropriate method, and the administration route can be administered through various routes such as oral or parenteral as long as it can reach the target tissue. there is. Examples of routes of administration include oral, intramuscular, intravenous, arterial, subcutaneous, peritoneal, pulmonary, and nasal, and may include, but are not limited to, subcutaneous, oral, or intravenous administration.
본 발명에서 용어, "약학적으로 유효한 양"은 의학적 용도에 적용 가능한 합리적인 수혜/위험 비율로 암을 예방 및/또는 치료하기에 충분한 양을 의미한다. 당업계에 공지된 방법에 따라 적절한 투여 량 및 투여 횟수가 선택될 수 있으며, 실제로 투여되는 본 발명의 약학적 조성물의 양 및 투여 횟수는 치료하고자 하는 증상의 종류, 투여 경로, 성별, 건강 상태, 식이, 개체의 연령, 체중 및 질환의 중증도와 같은 다양한 인자에 의해 적절하게 결정될 수 있다.As used herein, the term “pharmaceutically effective amount” means an amount sufficient to prevent and/or treat cancer with a reasonable benefit/risk ratio applicable to medical use. The appropriate dosage and number of administrations can be selected according to methods known in the art, and the amount and number of administrations of the pharmaceutical composition of the present invention actually administered depend on the type of symptom to be treated, route of administration, gender, health condition, It can be appropriately determined by various factors such as diet, the individual's age, body weight, and the severity of the disease.
본 발명의 말레이트 금속염을 유효성분으로 포함하는 조성물은 약학적 조성물 뿐만 아니라 암을 개선시키기 위한 기능성을 가진 식품 조성물의 형태로도 제조될 수 있다.The composition containing the maleate metal salt of the present invention as an active ingredient can be manufactured not only as a pharmaceutical composition but also in the form of a food composition with functionality for improving cancer.
본 발명의 조성물이 식품 조성물의 형태로 제조될 경우, 상기 식품 조성물은 식품에 통상 사용되어 냄새, 맛, 시각 등을 향상시킬 수 있는 추가 성분을 포함할 수 있다. 예를 들어, 식품 첨가물 (food additives)을 첨가할 수 있다. 상기 첨가물은 식품의 종류에 따라 선별되고 적절한 양으로 사용된다.When the composition of the present invention is prepared in the form of a food composition, the food composition may contain additional ingredients that are commonly used in food to improve odor, taste, vision, etc. For example, food additives can be added. The additives are selected according to the type of food and used in an appropriate amount.
상기 식품 조성물은 건강기능성 식품으로 제조될 수 있으며, 여기서 건강기능성 식품 (functional food)이란 특정보건용 식품 (food for special health use, FoSHU)과 동일한 용어로, 영양 공급 외에도 생체조절기능이 효율적으로 나타나도록 가공된 의학, 의료효과가 높은 식품을 의미한다. 상기 건강기능성 식품은 암의 개선에 유용한 효과를 얻기 위하여 정제, 캡슐, 분말, 과립, 액상, 환 등의 다양한 형태로 제조될 수 있다.The food composition can be manufactured as a health functional food, where functional food (functional food) is the same term as food for special health use (FoSHU), and has an efficient bioregulatory function in addition to supplying nutrients. It refers to processed medicine and food with high medical effectiveness. The health functional food can be manufactured in various forms such as tablets, capsules, powders, granules, liquids, pills, etc. to achieve useful effects in improving cancer.
본 발명의 다른 하나의 양태는, 말레이트 금속염을 유효성분으로 포함하는, 제2항암제에 대한 반응성을 증진시키는 항암 보조용 약학적 조성물이다.Another aspect of the present invention is an auxiliary anti-cancer pharmaceutical composition that improves reactivity to a second anti-cancer agent and includes maleate metal salt as an active ingredient.
본 발명의 또 다른 하나의 양태는, 말레이트 금속염 및 제2항암제를 유효성분으로 포함하는, 암의 예방 또는 치료용 약학적 조성물이다.Another aspect of the present invention is a pharmaceutical composition for preventing or treating cancer, comprising a maleate metal salt and a second anticancer agent as active ingredients.
말레이트 금속염에 대해서는 상술한 바와 같다.The maleate metal salt is as described above.
본 발명의 말레이트 금속염을 포함한 조성물은 단독으로 항암 효과를 가지는 것에서 더 나아가, 제2항암제에 대한 반응성을 증진시키기 위한 목적으로 항암 보조용 약학적 조성물로서도 매우 우수한 효과를 갖는다. 따라서 말레이트 금속염과 함께 제2항암제를 유효성분으로 하여 암의 예방 또는 치료용 약학적 조성물로 사용될 수 있다. The composition containing the maleate metal salt of the present invention not only has an anti-cancer effect on its own, but also has an excellent effect as an auxiliary anti-cancer pharmaceutical composition for the purpose of enhancing reactivity to a second anti-cancer agent. Therefore, it can be used as a pharmaceutical composition for the prevention or treatment of cancer using the second anticancer agent along with the maleate metal salt as an active ingredient.
본 발명에서 용어, "제2항암제"는 본 발명의 말레이트 금속염을 제외한 임의의 항암 활성을 갖는 약물을 말한다. 본 발명에서 상기 제2항암제의 범위는 특별히 제한되지 않고, 암의 종류와 진행 정도에 따라 암의 완치, 조절, 증상 완화 등의 목적에 따라 당업자가 적절한 종류를 선택하여 사용할 수 있다. 제2항암제는 예컨대 세포독성 항암제, 표적 항암제, 면역 항암제 또는 대사 항암제일 수 있으나, 이에 제한되지 않는다.In the present invention, the term “second anticancer agent” refers to any drug having anticancer activity except for the maleate metal salt of the present invention. In the present invention, the scope of the second anticancer agent is not particularly limited, and a person skilled in the art can select an appropriate type and use it for the purpose of curing, controlling, or alleviating symptoms, depending on the type and progression of cancer. The second anticancer agent may be, for example, a cytotoxic anticancer agent, a targeted anticancer agent, an immune anticancer agent, or a metabolic anticancer agent, but is not limited thereto.
본 발명에서, 세포독성 항암제는 정상 세포에 비해 빠른 속도로 무분별하게 분열하는 암 세포를 공격하여 항암 효과를 나타내는 약물로, 그 의미는 본 발명이 속한 기술분야에서 통상적으로 사용되는 것과 같다. 상기 세포독성 항암제는 알킬화제, 항대사제 (antimetabolite), 천연물 항암제를 포함한다.In the present invention, a cytotoxic anticancer agent is a drug that exhibits an anticancer effect by attacking cancer cells that divide indiscriminately at a faster rate than normal cells, and its meaning is the same as that commonly used in the technical field to which the present invention pertains. The cytotoxic anticancer agents include alkylating agents, antimetabolites, and natural anticancer agents.
상기 알킬화제는 질소 머스타드 (예컨대, 시클로포스파미드, 클로르메틴, 우라머스틴, 멜팔란, 클로람부실, 이포스파미드, 벤다머스틴 등), 알킬 설포네이트 (예컨대, 부설판, 프로카바진 등), 니트로소우레아 (예컨대, 카르무스틴, 로무스틴, 스트렙토조신 등), 백금 기반 알킬화제 (예컨대, 시스플라틴, 카보플라틴, 디시클로플라틴, 엡타플라틴, 로바플라틴, 미리플라틴, 네다플라틴, 옥살리플라틴, 피코플라틴, 사트라플라틴, 트리플라틴 테트라니트레이트 등)를 들 수 있으나, 이에 제한되지 않는다. 알킬화제는 암 세포 내 DNA에 결합하여 DNA 구조를 손상시킴으로써 암 세포의 파괴를 유발할 수 있다.The alkylating agent is nitrogen mustard (e.g., cyclophosphamide, chlormethine, uramustine, melphalan, chlorambucil, ifosfamide, bendamustine, etc.), alkyl sulfonate (e.g., busulfan, procarbazine, etc.) ), nitrosoureas (e.g., carmustine, lomustine, streptozocin, etc.), platinum-based alkylating agents (e.g., cisplatin, carboplatin, dicycloplatin, eptaplatin, lobaplatin, myriplatin, daplatin, oxaliplatin, picoplatin, satraplatin, triplatin tetranitrate, etc.), but is not limited thereto. Alkylating agents can cause destruction of cancer cells by binding to the DNA within cancer cells and damaging the DNA structure.
상기 항대사제는 피리미딘 유도체 (예컨대, 5-플루오로우라실, 카페시타빈, 시스타라빈, 젬시타빈, 플루다라빈 등), 폴레이트 유도체 (예컨대, 메토트렉세이트, 페메트렉시드 등), 퓨린 유도체 (예컨대, 머캅토퓨린 등)를 들 수 있으나, 이에 제한되지 않는다. 항대사제는 DNA 복제 및 세포 생존에 필요한 대사를 억제함으로써 암 세포 사멸을 유도할 수 있다.The antimetabolites include pyrimidine derivatives (e.g., 5-fluorouracil, capecitabine, cystarabine, gemcitabine, fludarabine, etc.), folate derivatives (e.g., methotrexate, pemetrexed, etc.), and purine derivatives (e.g., , mercaptopurine, etc.), but is not limited thereto. Antimetabolites can induce cancer cell death by inhibiting metabolism required for DNA replication and cell survival.
상기 천연물 항암제로는 토포이소머라제 억제제 (예컨대, 캄프토테신, 에피포도필로톡신, 탁산 계열 약물 (도시탁셀, 파클리탁셀)), 항생제 (예컨대, 닥티노마이신, 독소루비신, 다우노루비신, 마이토마이신, 플레오마이신, 이다루비신, 미톡산트론 HCl 등)를 들 수 있으나, 이에 제한되지 않는다.The natural anticancer agents include topoisomerase inhibitors (e.g., camptothecin, epipodophyllotoxin, taxane-based drugs (docitaxel, paclitaxel)), antibiotics (e.g., dactinomycin, doxorubicin, daunorubicin, and mitomycin). , phleomycin, idarubicin, mitoxantrone HCl, etc.), but is not limited thereto.
본 발명에서 표적 항암제는 암의 성장에 관여하는 타겟 단백질 (수용체 또는 효소)을 억제함으로써 암 세포의 사멸을 유도하는 항암제로, 그 의미는 본 발명이 속한 기술 분야에서 통상적으로 사용되는 것과 같다. 상기 세포독성 항암제는 표적 단백질 (티로신 키나아제 등) 억제 저분자 화합물 및 단클론항체를 포함한다.In the present invention, the targeted anticancer agent is an anticancer agent that induces death of cancer cells by inhibiting target proteins (receptors or enzymes) involved in cancer growth, and its meaning is the same as that commonly used in the technical field to which the present invention pertains. The cytotoxic anticancer agents include small molecule compounds and monoclonal antibodies that inhibit target proteins (tyrosine kinases, etc.).
일례로, 상기 표적 항암제는 VEGF-A, HER2 및 EGFR로 구성된 군에서 선택된 하나 이상의 타겟을 표적하는 항체 또는 티로신 키나아제 억제제일 수 있다.For example, the targeted anticancer agent may be an antibody or tyrosine kinase inhibitor targeting one or more targets selected from the group consisting of VEGF-A, HER2, and EGFR.
일 실시양태에서, 말레이트 금속염과 병용 투여될 수 있는 표적 항암제는 VEGF-A 억제제이다. 본 발명에서, VEGF-A 억제제는 베바시주맙 (bevacizumab), 라니비주맙 (ranibizumab), 아플리베르셉트 (aflibercept), 라무시루맙 (ramucirumab)과 같은 단클론 항체, 수니티닙 (sunitinib), 파조파닙 (pazopanib), 소라페닙 (sorafenib), 악시티닙 (axitinib)과 같은 저분자 화합물을 들 수 있으나, 이에 제한되는 것은 아니다.In one embodiment, the targeted anti-cancer agent that can be administered in combination with maleate metal salt is a VEGF-A inhibitor. In the present invention, the VEGF-A inhibitor is monoclonal antibodies such as bevacizumab, ranibizumab, aflibercept, ramucirumab, sunitinib, and pazopar. Low molecular weight compounds such as pazopanib, sorafenib, and axitinib may be included, but are not limited thereto.
일 실시양태에서, 말레이트 금속염과 병용 투여될 수 있는 표적 항암제는 EGFR 억제제이다. 본 발명에서, EGFR 억제제는 오시메르티닙 (osimertinib), 제피티닙 (gefitinib), 엘로티닙 (erlotinib), 아파티닙 (afatinib), 브리가티닙 (brigatinib), 이코티닙 (icotinib), 반데타닙 (vandetanib)과 같은 저분자 화합물과 함께 세툭시맙 (cetuximab), 파니투무맙 (panitumumab), 잘루투무맙 (zalutumumab), 니모투주맙 (nimotuzumab), 마투주맙 (matuzumab)과 같은 단클론 항체를 들 수 있으나, 이에 제한되지 않는다. In one embodiment, the targeted anti-cancer agent that can be administered in combination with the maleate metal salt is an EGFR inhibitor. In the present invention, the EGFR inhibitor is osimertinib, gefitinib, erlotinib, afatinib, brigatinib, icotinib, vandetanib ( monoclonal antibodies such as cetuximab, panitumumab, zalutumumab, nimotuzumab, and matuzumab, along with small molecule compounds such as vandetanib, It is not limited to this.
일 실시양태에서, 말레이트 금속염과 병용 투여될 수 있는 표적 항암제는 HER2 억제제로서 트라스투주맙 (trastuzumab), 페르투주맙 (pertuzumab), 라파니팁 (lapatinib), 네라티닙 (neratinib), 또는 아파티닙 (afatinib)일 수 있으나, 이에 제한되지 않는다.In one embodiment, the targeted anti-cancer agent that can be administered in combination with the maleate metal salt is a HER2 inhibitor such as trastuzumab, pertuzumab, lapatinib, neratinib, or It may be, but is not limited to, afatinib.
이 외에도 본 발명의 표적 항암제는 또한, 이매티닙 (imatinib), 다사티닙 (dasatinib), 닐로티닙 (nilotinib)과 같은 Bcr-Abl 표적 항암제; 보수티닙 (bosutinib)과 같은 Src 표적 항암제; 레스타우르티닙 (lestaurtinib), 룩소리티닙 (ruxolitinib), 파크리티닙 (pacritinib)과 같은 JAK 표적 항암제; 코비메티닙 (cobimethinib), 셀루메티닙 (selumetinib), 트라메티닙 (trametinib), 비니메티닙 (binimetinib)과 같은 MAP2 키나아제 표적 항암제; 세리티빈 (ceritibin), 크리조티닙 (crizotinib)과 같은 MEL4-ALK 표적 항암제 등을 포함하며, 그 종류에 특별히 제한되지 않는다.In addition, the targeted anticancer agent of the present invention also includes Bcr-Abl targeting anticancer agents such as imatinib, dasatinib, and nilotinib; Src-targeting anticancer drugs such as bosutinib; JAK-targeting anticancer drugs such as lestaurtinib, ruxolitinib, and pacritinib; MAP2 kinase targeting anticancer drugs such as cobimethinib, selumetinib, trametinib, and binimetinib; It includes MEL4-ALK targeting anticancer drugs such as ceritibin and crizotinib, and is not particularly limited to the type.
본 발명의 제2항암제는 하나 이상의 표적 항암제 또는 면역 항암제의 조합일 수 있으며, 이들은 동시에 또는 이시에 투여될 수 있다. 본 발명의 구체적인 일 실시예에서는 말레이트 금속염, 또는 말레이트 금속염 및 DCA를 제2항암제인 소라페닙, 트라스투주맙 (허셉틴) 또는 오시메르티닙과 병용하여 여러 암종에 처리한 결과, 제2항암제 또는 말레이트 금속염 (또는 DCA 추가) 단독 처리에 비해 현저하게 암 세포를 사멸시키는 것을 확인하였다 (도 7). The second anticancer agent of the present invention may be a combination of one or more targeted anticancer agents or immune anticancer agents, and they may be administered simultaneously or at different times. In a specific embodiment of the present invention, as a result of treating several carcinomas by combining maleate metal salt, or maleate metal salt and DCA with the second anticancer drug, sorafenib, trastuzumab (Herceptin), or osimertinib, the second anticancer drug or It was confirmed that cancer cells were killed significantly compared to treatment with maleate metal salt (or addition of DCA) alone (FIG. 7).
일 실시 양태로서, 본 발명의 제2항암제는 CAR-T, CAR-NK, 면역체크포인트 억제제, 항암백신 등의 면역 항암제일 수 있으나, 이에 제한되지 않는다. 면역 세포는 종양 조직 내 저산소 환경에서 암 세포의 급격한 해당과정에 의해 에너지 고갈을 겪게 되는데, 이러한 환경은 항암 치료와 관련하여 면역 활성이 억제되는 중요한 원인 중 하나이다. 본 발명의 구체적 일 실시예에서는 T 세포, NK 세포에 IL-2 또는 IL-12와 함께 말레이트 금속염을 처리하였을 때 사이토카인 단독 처리에 비해 면역 세포의 활성이 크게 향상되는 것을 확인하였다 (도 9). 따라서 말레이트 금속염은 제2항암제로서 CAR-T, CAR-NK, 면역체크포인트 억제제, 항암백신 등의 면역 항암제를 사용할 경우에도 시너지 효과를 나타낼 수 있다.In one embodiment, the second anticancer agent of the present invention may be an immunological anticancer agent such as CAR-T, CAR-NK, immune checkpoint inhibitor, or anticancer vaccine, but is not limited thereto. Immune cells experience energy depletion due to the rapid glycolysis of cancer cells in the hypoxic environment within tumor tissue, and this environment is one of the important causes of suppressed immune activity in relation to anticancer treatment. In a specific example of the present invention, it was confirmed that when T cells and NK cells were treated with maleate metal salt together with IL-2 or IL-12, the activity of immune cells was significantly improved compared to treatment with cytokines alone (Figure 9 ). Therefore, maleate metal salt can have a synergistic effect even when used as a second anticancer drug with immunological anticancer drugs such as CAR-T, CAR-NK, immune checkpoint inhibitors, and anticancer vaccines.
또한, 본 발명의 제2항암제는 암의 종류와 진행 정도에 따라 암의 완치, 조절, 증상 완화를 위해 적절한 종류를 선택할 수 있다. 본 발명의 말레이트 금속염을 포함하는 조성물은 암 세포 내 NAD+의 고갈을 촉진시켜 제2항암제에 대한 반응성을 증진시키는 효과를 가져 항암 보조제로서 사용될 수 있으므로, 사용되는 제2항암제의 종류나 암의 종류가 특별히 제한되지 않는다. 일례로, 상기 암은 간암, 폐암, 대장암, 뇌암, 신장암, 췌장암, 유방암 및 암성 당뇨일 수 있으나, 이에 제한되는 것은 아니다.In addition, the second anti-cancer agent of the present invention can be selected according to the type and progression of cancer, appropriate for curing, controlling, and alleviating symptoms of cancer. The composition containing the maleate metal salt of the present invention has the effect of promoting depletion of NAD + in cancer cells and enhancing reactivity to the second anticancer agent, so it can be used as an anticancer adjuvant, so it can be used as an anticancer adjuvant, so it can be used as an anticancer adjuvant. The type is not particularly limited. For example, the cancer may be liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, breast cancer, and cancerous diabetes, but is not limited thereto.
본 발명의 말레이트 금속염을 포함한 조성물은 제2항암제와 독립적으로 존재하는 형태로 병용투여될 수 있을 뿐만 아니라, 목적에 따라 공지된 임의의 방법으로 제2항암제와 물리/화학적인 결합을 형성한 상태에서 투여될 수 있다. 예컨대, 말레이트 금속염을 포함한 조성물은 제2항암제와 직접 결합된 상태로 사용되거나, 또는 공지의 링커를 통해 제2항암제와 연결된 상태로 사용될 수 있으며, 본 발명의 말레이트 금속염을 포함하는 조성물이 제2항암제와 함께 작용하여 상승된 항암 효과를 나타내는 한 그 적용 방법에 특별히 제한되지 않는다.The composition containing the maleate metal salt of the present invention can be co-administered in a form that exists independently of the second anticancer agent, as well as forming a physical/chemical bond with the second anticancer agent by any known method depending on the purpose. It can be administered at. For example, the composition containing the maleate metal salt may be used in a state directly bound to the second anticancer agent, or may be used in a state connected to the second anticancer agent through a known linker, and the composition containing the maleate metal salt of the present invention may be used as the second anticancer agent. 2. There are no particular restrictions on the method of application as long as it works together with anticancer drugs to exhibit increased anticancer effects.
본 발명의 또 다른 하나의 양태는, 상기 말레이트 금속염을 유효성분으로 포함하는, 방사선 항암 치료에 대한 반응성을 증진시키는 항암 보조용 약학적 조성물이다.Another aspect of the present invention is an adjuvant anti-cancer pharmaceutical composition that improves responsiveness to radiation anti-cancer treatment, comprising the maleate metal salt as an active ingredient.
말레이트 금속염에 대해서는 상술한 바와 같다. The maleate metal salt is as described above.
본 발명에서 용어, "방사선 항암 치료"는 암 세포를 사멸시키기 위한 목적으로 암 세포 또는 종양 조직에 방사선을 조사하는 치료 행위를 말한다. 일반적으로 절개 불가능하거나 수술 불가능한 종양, 또는 종양 전이를 제어하기 위한 표준 치료법으로서 표적 부위로 전달된 방사선이 재생성 세포 (reproductive cell)의 사멸을 초래한다는 원리에 기초한다. 본 발명에서 방사선 항암 치료는 이온화 방사선 요법 (ionizing radiation therapy), 전자기 방사선 요법 (electromagnetic radiation), 근접 방사선 요법 (brachytherapy) 또는 외부 방사선 요법 (external beam radiation therapy)일 수 있으나, 이에 제한되는 것은 아니다. In the present invention, the term “radiation anti-cancer treatment” refers to the treatment of irradiating cancer cells or tumor tissue with radiation for the purpose of killing cancer cells. In general, it is a standard treatment for controlling unresectable or inoperable tumors or tumor metastases, and is based on the principle that radiation delivered to the target site causes the death of regenerative cells. In the present invention, the radiation anti-cancer treatment may be ionizing radiation therapy, electromagnetic radiation therapy, brachytherapy, or external beam radiation therapy, but is not limited thereto.
본 발명에 따른 말레이트 금속염을 포함하는 조성물은 방사선 요법과 병용될 경우 상승적 항암 효과를 나타내므로 방사선 요법에 대한 항암 보조제 또는 방사선 민감도를 향상시키는 방사선 민감화제로 유용하게 활용될 수 있으며, 적용 가능한 암의 종류가 특별히 제한되지 않는다. 일례로, 상기 암은 간암, 폐암, 대장암, 뇌암, 신장암, 췌장암, 유방암 및 암성 당뇨일 수 있으나, 이에 제한되는 것은 아니다. The composition containing maleate metal salt according to the present invention exhibits a synergistic anti-cancer effect when used in combination with radiotherapy, so it can be usefully used as an anti-cancer adjuvant for radiotherapy or as a radiosensitizer to improve radiosensitivity, and is applicable to cancer. The type is not particularly limited. For example, the cancer may be liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, breast cancer, and cancerous diabetes, but is not limited thereto.
본 발명의 실시 형태는 여러가지 다른 형태로 변형될 수 있으며, 본 발명의 범위가 이하 설명하는 실시 형태로 한정되는 것은 아니다. 또한 본 발명의 실시 형태는 당해 기술분야에서 평균적인 지식을 가진 자에게 본 발명을 더욱 완전하게 설명하기 위해서 제공되는 것이다. 나아가, 명세서 전체에서 어떤 구성요소를 "포함"한다는 것은 특별히 반대되는 기재가 없는 한 다른 구성요소를 제외하는 것이 아니라 다른 구성요소를 더 포함할 수 있다는 것을 의미한다.The embodiments of the present invention may be modified into various other forms, and the scope of the present invention is not limited to the embodiments described below. Additionally, embodiments of the present invention are provided to more completely explain the present invention to those with average knowledge in the relevant technical field. Furthermore, “including” a certain element throughout the specification means that other elements may be further included, rather than excluding other elements, unless specifically stated to the contrary.
본 발명의 말레이트 금속염을 유효성분으로 포함하는 암의 예방 또는 치료용 조성물은 암 세포 내 에너지원인 NAD+를 고갈시키고 면역 활성을 증가시켜 우수한 항암 효과를 가지며, 말산 활성 증가로 인슐린 분비를 증가시켜 다양한 항암 치료에 의한 독성으로 유도되는 인슐린 분비 감소나 저항성 문제를 해결할 수 있다. 또한, 다른 항암제의 투여나 방사선 항암 치료 시 병용 투여될 경우 항암제 및 방사선 치료에 대한 반응성을 증진시켜 항암 보조제로서도 매우 우수한 효과를 가진다.The composition for preventing or treating cancer containing malate metal salt as an active ingredient of the present invention has excellent anticancer effects by depleting NAD + , an energy source in cancer cells, and increasing immune activity, and increases insulin secretion by increasing malic acid activity. It can solve the problem of decreased insulin secretion or resistance caused by toxicity from various anticancer treatments. In addition, when administered in combination with other anticancer drugs or during radiotherapy, it improves responsiveness to anticancer drugs and radiotherapy, making it very effective as an anticancer adjuvant.
도 1은 암 세포에 CaMal, CaMal2 또는 CaMal-DCA 처리 시 MDH2와 ME2 및 ME3의 수준을 비교한 그래프이다.Figure 1 is a graph comparing the levels of MDH2, ME2, and ME3 when cancer cells are treated with CaMal, CaMal 2 , or CaMal-DCA.
도 2는 암 세포에 CaMal, CaMal2 또는 CaMal-DCA 처리 시 NAD+ 수준을 비교한 그래프이다.Figure 2 is a graph comparing NAD + levels when cancer cells are treated with CaMal, CaMal 2 , or CaMal-DCA.
도 3은 췌장 베타 세포에 CaMal 또는 CaMal2 처리 시군의 글루코스 유무에 따른 인슐린 분비 수준을 비교한 그래프이다.Figure 3 is a graph comparing the insulin secretion level according to the presence or absence of glucose in the pancreatic beta cells in the CaMal or CaMal 2 treatment group.
도 4는 CaMal 처리 농도에 따른 암 세포의 생존율을 비교한 그래프이다.Figure 4 is a graph comparing the survival rate of cancer cells according to CaMal treatment concentration.
도 5는 CaMal2 처리 농도에 따른 암 세포의 생존율을 비교한 그래프이다.Figure 5 is a graph comparing the survival rate of cancer cells according to CaMal 2 treatment concentration.
도 6은 CaMal-DCA 처리 농도에 따른 암 세포의 생존율을 비교한 그래프이다.Figure 6 is a graph comparing the survival rate of cancer cells according to CaMal-DCA treatment concentration.
도 7은 암 세포에 제2항암제와 CaMal, CaMal2 또는 CaMal-DCA를 단독 또는 병용으로 투여하여 세포의 생존율을 비교한 그래프이다.Figure 7 is a graph comparing the survival rate of cells when the second anticancer drug and CaMal, CaMal 2 , or CaMal-DCA were administered alone or in combination to cancer cells.
도 8은 암 세포에 CaMal, CaMal2 또는 CaMal-DCA와 방사선을 단독 또는 병용 처리하여 세포의 생존율을 비교한 그래프이다.Figure 8 is a graph comparing the survival rate of cancer cells treated with CaMal, CaMal 2 , or CaMal-DCA and radiation alone or in combination.
도 9는 IL-2 단독, 또는 IL-2 또는 IL-12와 CaMal2를 병용 처리하여 활성화된 T 세포 및 NK 세포의 수준을 비교한 그래프이다. Figure 9 is a graph comparing the levels of activated T cells and NK cells by treatment with IL-2 alone or in combination with IL-2 or IL-12 and CaMal 2 .
도 10은 췌장암 이식 마우스에서 CaMal의 피하 투여에 따른 종양 성장 억제 효과를 나타낸 그래프이다. Figure 10 is a graph showing the tumor growth inhibition effect of subcutaneous administration of CaMal in pancreatic cancer transplant mice.
도 11은 간암 이식 마우스에서 CaMal2의 경구 투여에 따른 종양 성장 억제 효과를 나타낸 그래프이다. Figure 11 is a graph showing the tumor growth inhibition effect of oral administration of CaMal 2 in liver cancer transplant mice.
도 12는 유방암 이식 마우스에서 CaMal-DCA의 정맥 내 투여에 따른 종양 성장 억제 효과를 나타낸 그래프이다. Figure 12 is a graph showing the tumor growth inhibition effect of intravenous administration of CaMal-DCA in breast cancer transplant mice.
이하, 실험예를 통하여 본 발명의 구성 및 효과를 더욱 상세히 설명하고자 한다. 이들 실험예는 오로지 본 발명을 예시하기 위한 것일 뿐 본 발명의 범위가 이들 실시예에 의해 제한되는 것은 아니다.Hereinafter, the configuration and effects of the present invention will be explained in more detail through experimental examples. These experimental examples are only for illustrating the present invention and the scope of the present invention is not limited by these examples.
실험예 1: 암 세포에서 말레이트 금속염의 대사 조절 효과 확인Experimental Example 1: Confirmation of metabolic regulation effect of maleate metal salt in cancer cells
1-1. MDH2 (malate dehydrogenase 2), ME2 (malic enzyme 2) 및 ME3 (malic enzyme 3) 수준의 변화1-1. Changes in malate dehydrogenase 2 (MDH2), malic enzyme 2 (ME2), and malic enzyme 3 (ME3) levels
1 x 106 개의 간암 (HepG2), 유방암 (MDA-MB-231), 또는 폐암 (H1975) 세포에 5 mM 칼슘 말레이트 (calcium malate, CaMal), 2.5 mM 칼슘 디말레이트 (calcium dimalate, CaMal2) 또는 1 mM 칼슘 말레이트-디클로로아세트산 (calcium malate-dichloroacetic acid, CaMal-DCA)을 각각 48 시간 동안 처리하였다. 그 후 RIPA 버퍼를 활용하여 단백질을 추출하였다. Biorbyt사의 MDH2 (Malate dehydrogenase 2), ME2 (Malic enzyme 2) 및 ME3 (Malic enzyme 3) ELISA 분석 키트를 활용하여 상기 추출된 단백질에서 각각의 인자를 정량 분석하였다.1 x 106 liver cancer (HepG2), breast cancer (MDA-MB-231), or lung cancer (H1975) cells with 5 mM calcium malate (CaMal), 2.5 mM calcium dimalate (CaMal 2 ) Alternatively, each was treated with 1 mM calcium malate-dichloroacetic acid (CaMal-DCA) for 48 hours. Afterwards, the protein was extracted using RIPA buffer. Each factor was quantitatively analyzed in the extracted protein using Biorbyt's MDH2 (Malate dehydrogenase 2), ME2 (Malic enzyme 2), and ME3 (Malic enzyme 3) ELISA assay kit.
간암 세포, 유방암 세포, 폐암 세포에 각각 CaMal, CaMal2 또는 CaMal-DCA를 처리한 결과, 미처리 대조군에 비해 MDH2, ME2, ME3의 양이 현저히 증가하는 것을 확인하였다 (도 1; **p < 0.001).When liver cancer cells, breast cancer cells, and lung cancer cells were treated with CaMal, CaMal 2 , or CaMal-DCA, respectively, the amounts of MDH2, ME2, and ME3 were confirmed to significantly increase compared to the untreated control group (Figure 1; ** p < 0.001 ).
1-2. NAD1-2. NAD ++ 수준의 변화change in level
1 x 106 개의 간암 (HepG2), 유방암 (MDA-MB-231), 또는 폐암 (H1975) 세포에 5 mM CaMal, 2.5 mM CaMal2 또는 1 mM CaMal-DCA를 각각 48 시간 동안 처리하였다. 그 후 Abcam사의 NAD+ 분석 키트의 방법에 따라 NAD+ 수준을 분석하였다.1 x 10 6 liver cancer (HepG2), breast cancer (MDA-MB-231), or lung cancer (H1975) cells were treated with 5mM CaMal, 2.5mM CaMal 2 , or 1mM CaMal-DCA, respectively, for 48 hours. Afterwards, the NAD + level was analyzed according to the method of Abcam's NAD + analysis kit.
간암 세포, 유방암 세포, 폐암 세포에 각각 CaMal, CaMal2 또는 CaMal-DCA를 처리한 결과, 미처리 대조군에 비해 모두 NAD+의 수준이 현저히 감소하는 것을 확인하였다 (도 2; **p < 0.001).When liver cancer cells, breast cancer cells, and lung cancer cells were treated with CaMal, CaMal 2 , or CaMal-DCA, respectively, the level of NAD + was confirmed to be significantly reduced compared to the untreated control group (Figure 2; ** p < 0.001).
1-3. 인슐린 수준의 변화1-3. changes in insulin levels
1 x 106 개의 베타 세포에 글루코스가 없거나 5 mM 글루코스가 있는 조건에서 5 mM CaMal 또는 2.5 mM CaMal2를 각각 48 시간 동안 처리한 후 배양 배지를 회수하였다. 상기 배지를 스핀 농축기 (Corning, Inc.)에 넣고 7500 g로 10 분간 작동하여 인슐린 분석 샘플을 획득하였다. Enzo사의 분석 키트를 사용하여 상기 샘플을 정량 분석하였다.1 x 10 6 beta cells were treated with 5mM CaMal or 2.5mM CaMal 2 for 48 hours in the absence of glucose or in the presence of 5mM glucose, respectively, and then the culture medium was recovered. The medium was placed in a spin concentrator (Corning, Inc.) and operated at 7500 g for 10 minutes to obtain an insulin analysis sample. The sample was quantitatively analyzed using Enzo's analysis kit.
베타 세포에 CaMal 또는 CaMal2를 처리한 결과, 글루코스가 없거나 있는 조건에서 모두 미처리 대조군에 비해 인슐린의 수준이 유의성 있게 증가하는 것을 확인하였다 (도 3; **p < 0.001 vs Control (No-glucose); ## p < 0.001 vs Control (Glucose)).As a result of treating beta cells with CaMal or CaMal 2 , it was confirmed that the level of insulin significantly increased compared to the untreated control group in both the absence and presence of glucose (Figure 3; ** p < 0.001 vs Control (No-glucose) ; ## p < 0.001 vs Control (Glucose)).
실험예 2: Experimental Example 2: in vitroin vitro 에서 말레이트 금속염의 항암 효과Anticancer effect of maleate metal salt in
2-1. CaMal의 항암효과2-1. Anti-cancer effect of CaMal
3 x 103 개의 대장암 (HCT-116) 또는 뇌암 (U-87MG)세포를 24 시간 동안 배양한 후 0.5, 1, 2.5 또는 5 mM CaMal를 처리하여 다시 96 시간 배양하였다. 그 후 10 μl의 3-(4,5-디메틸티아졸 -2-일)-2,5-디페닐테트라졸리움 브로마이드 시약을 각 웰에 넣어 1 시간 동안 반응시켰다. 반응을 마친 시약을 제거한 후 200 μl의 DMSO를 각 웰에 넣은 뒤 흡광도를 측정하여 세포의 생존율을 확인하였다.3 x 103 colon cancer (HCT-116) or brain cancer (U-87MG) cells were cultured for 24 hours, then treated with 0.5, 1, 2.5, or 5 mM CaMal and cultured again for 96 hours. Afterwards, 10 μl of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 μl of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
대장암 (HCT-116) 또는 뇌암 (U-87MG) 세포에 CaMal를 처리한 결과, CaMal의 농도 의존적으로 암 세포의 생존율이 감소하였으며, 이러한 결과는 유의성이 있는 것으로 나타났다 (도 4; **p < 0.001 vs 0.5, 1, 2.5 mM groups).When colon cancer (HCT-116) or brain cancer (U-87MG) cells were treated with CaMal, the survival rate of cancer cells decreased in a CaMal concentration-dependent manner, and this result was found to be significant (Figure 4; ** p < 0.001 vs 0.5, 1, 2.5 mM groups).
2-2. CaMal2-2. CaMal 22 의 항암효과anticancer effect of
3 x 103 개의 신장암 (Caki-1) 또는 췌장암 (Aspc-1) 세포를 24 시간 동안 배양한 후 0.5, 1, 2.5 또는 5 mM CaMal2를 처리하여 다시 96 시간 동안 배양하였다. 그 후 10 μl의 3-(4,5-디메틸티아졸-2-일)-2,5-디페닐테트라졸리움 브로마이드 시약을 각 웰에 넣어 1 시간 동안 반응시켰다. 반응을 마친 시약을 제거한 후 200 μl의 DMSO를 각 웰에 넣은 뒤 흡광도를 측정하여 세포의 생존율을 확인하였다.3 x 10 3 kidney cancer (Caki-1) or pancreatic cancer (Aspc-1) cells were cultured for 24 hours, then treated with 0.5, 1, 2.5, or 5 mM CaMal 2 and cultured again for 96 hours. Afterwards, 10 μl of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 μl of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
신장암 (Caki-1) 또는 췌장암 (Aspc-1) 세포에 CaMal2를 처리한 결과, CaMal2의 농도 의존적으로 암 세포의 생존율이 감소하였고, 이러한 결과는 유의성이 있는 것으로 나타났다 (도 5; **p < 0.001 vs 0.5, 1 mM groups. ##p < 0.001 vs 0.5, 1, 2.5 mM groups).When kidney cancer (Caki-1) or pancreatic cancer (Aspc-1) cells were treated with CaMal 2 , the survival rate of cancer cells decreased in a CaMal 2 concentration-dependent manner, and this result was found to be significant (Figure 5; * * p < 0.001 vs 0.5, 1 mM groups. ## p < 0.001 vs 0.5, 1, 2.5 mM groups).
2-3. CaMal-DCA의 항암효과2-3. Anticancer effect of CaMal-DCA
3 x 103 개의 간암 (HepG2), 폐암 (H1975), 췌장암 (Aspc-1) 또는 유방암 (MDA-MB-231) 세포를 24 시간 동안 배양한 후 0.5, 1, 2.5 또는 5 mM CaMal-DCA를 처리하여 다시 96 시간 동안 배양하였다. 그 후 10 μl의 3-(4,5-디메틸티아졸-2-일)-2,5-디페닐테트라졸리움 브로마이드시약을 각 웰에 넣어 1 시간 동안 반응시켰다. 반응을 마친 시약을 제거한 후 200 μl의 DMSO를 각 웰에 넣은 뒤 흡광도를 측정하여 세포의 생존율을 확인하였다.3 x 10 liver (HepG2), lung (H1975), pancreatic (Aspc-1), or breast (MDA-MB-231) cells were cultured for 24 h and then incubated with 0.5, 1, 2.5, or 5 mM CaMal-DCA. The cells were treated and cultured again for 96 hours. Afterwards, 10 μl of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 μl of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
간암 (HepG2), 폐암 (H1975), 췌장암 (Aspc-1) 또는 유방암 (MDA-MB-231) 세포에 CaMal-DCA를 처리한 결과, CaMal-DCA의 농도 의존적으로 암 세포의 생존율이 감소하였고, 이러한 결과는 유의성이 있는 것으로 나타났다 (도 6; **p < 0.001 vs 0.5 group, ##p < 0.001 vs 0.5, 1 mM groups; $$p < 0.001 vs 0.5, 1, 2.5 mM groups).When liver cancer (HepG2), lung cancer (H1975), pancreatic cancer (Aspc-1), or breast cancer (MDA-MB-231) cells were treated with CaMal-DCA, the survival rate of cancer cells decreased in a concentration-dependent manner of CaMal-DCA. These results appeared to be significant (Figure 6; ** p < 0.001 vs 0.5 group, ## p < 0.001 vs 0.5, 1 mM groups; $$ p < 0.001 vs 0.5, 1, 2.5 mM groups).
실험예 3: 항암제와 말레이트 금속염의 병용에 따른 항암효과Experimental Example 3: Anticancer effect due to combined use of anticancer drug and maleate metal salt
3 x 103 개의 간암 (HepG2), 유방암 (MDA-MB-231) 또는 폐암 (H1975) 세포를 24 시간 동안 배양하였다. 그 후 간암 세포는 1 μM 소라페닙 (sorafenib)과 5 mM CaMal을 단독 또는 병용 처리하였고, 유방암 세포는 2.5 μM 허셉틴 (Herceptin)과 2.5 mM CaMal2를 단독 또는 병용 처리하였으며, 폐암 세포는 1 nM 오시머티닙 (Osimertinib)과 1 mM CaMal-DCA를 단독 또는 병용 처리하여 다시 96 시간을 배양하였다. 그 후 10 μl의 3-(4,5-디메틸티아졸-2-일)-2,5-디페닐테트라졸리움 브로마이드 시약을 각 웰에 넣어 1 시간 동안 반응시켰다. 반응을 마친 시약을 제거한 후 200 μl의 DMSO를 각 웰에 넣은 뒤 흡광도를 측정하여 세포의 생존율을 확인하였다.3 x 103 liver cancer (HepG2), breast cancer (MDA-MB-231), or lung cancer (H1975) cells were cultured for 24 hours. Afterwards, liver cancer cells were treated with 1 μM sorafenib and 5 mM CaMal alone or in combination, breast cancer cells were treated with 2.5 μM Herceptin and 2.5 mM CaMal 2 alone or in combination, and lung cancer cells were treated with 1 nM Oshi. Osimertinib and 1mM CaMal-DCA were treated alone or in combination and cultured for another 96 hours. Afterwards, 10 μl of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 μl of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
간암 (HepG2), 유방암 (MDA-MB-231) 또는 폐암 (H1975) 세포에 말레이트 금속염과 항암제를 단독 또는 병용 처리한 결과, 말레이트 금속염은 단독 투여로도 항암 효과가 우수한 것으로 공지된 기존 항암제들과 동등하거나 보다 우수한 효과를 나타냈으며, 특히 각 항암제와 말레이트 금속염을 병용으로 처리한 경우 시너지 효과를 나타내 거의 모든 암 세포가 사멸된 것을 확인하였다. 이러한 결과는, 말레이트 금속염을 항암제와 병용투여함으로써 항암제 내성을 극복하고 매우 우수한 항암 효과를 나타낼 수 있음을 시사한다 (도 7; *p < 0.001 vs Control, **p < 0.001 vs Control, single treatment groups).As a result of treating liver cancer (HepG2), breast cancer (MDA-MB-231), or lung cancer (H1975) cells with maleate metal salt and anticancer drugs alone or in combination, maleate metal salt is an existing anticancer drug known to have excellent anticancer effects even when administered alone. It showed an effect equal to or better than that of other anticancer drugs, and in particular, when each anticancer drug and maleate metal salt were treated in combination, a synergistic effect was observed and almost all cancer cells were confirmed to be killed. These results suggest that co-administration of maleate metal salt with anticancer drugs can overcome anticancer drug resistance and exhibit excellent anticancer effects (Figure 7; * p < 0.001 vs Control, ** p < 0.001 vs Control, single treatment groups).
실험예 4: 방사선과 말레이트 금속염의 병용에 따른 항암효과Experimental Example 4: Anticancer effect due to combined use of radiation and maleate metal salt
3 x 103 개의 간암 (HepG2), 유방암 (MDA-MB-231) 또는 폐암 (H1975) 세포를 24 시간 동안 배양하였다. 그 후 각각의 세포에 2 Gy 선량의 방사선과 5 mM CaMal, 2.5 Mm CaMal2 또는 1 mM CaMal-DCA를 단독 또는 병용 처리하여 48 시간 동안 배양하였다. 그 후 10 μl의 3-(4,5-디메틸티아졸-2-일)-2,5-디페닐테트라졸리움 브로마이드 시약을 각 웰에 넣어 1 시간 동안 반응시켰다. 반응을 마친 시약을 제거한 후 200 μl의 DMSO를 각 웰에 넣은 뒤 흡광도를 측정하여 세포의 생존율을 확인하였다.3 x 103 liver cancer (HepG2), breast cancer (MDA-MB-231), or lung cancer (H1975) cells were cultured for 24 hours. Afterwards, each cell was treated alone or in combination with a dose of 2 Gy of radiation and 5 mM CaMal, 2.5 Mm CaMal 2 or 1 mM CaMal-DCA and cultured for 48 hours. Afterwards, 10 μl of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reagent was added to each well and reacted for 1 hour. After removing the reagent after completing the reaction, 200 μl of DMSO was added to each well and the absorbance was measured to confirm the survival rate of the cells.
간암 (HepG2), 유방암 (MDA-MB-231) 또는 폐암 (H1975) 세포에 말레이트 금속염과 방사선을 단독 또는 병용 처리한 결과, 말레이트 금속염은 2 Gy의 방사선 조사 보다도 효과적으로 암 세포를 사멸시켰고, 특히 방사선과 병용으로 말레이트 금속염을 처리한 경우 상승된 항암 효능을 나타냈다. 이러한 결과는, 말레이트 금속염을 방사선 조사와 병용함으로써 방사선 조사에 대한 내성을 극복할 수 있음을 시사한다 (도 8; *p < 0.001 vs Control, **p < 0.001 vs Control, single treatment groups).When liver cancer (HepG2), breast cancer (MDA-MB-231), or lung cancer (H1975) cells were treated with maleate metal salt and radiation alone or in combination, maleate metal salt killed cancer cells more effectively than 2 Gy of radiation. In particular, treatment with maleate metal salt in combination with radiation showed increased anticancer efficacy. These results suggest that resistance to irradiation can be overcome by combining maleate metal salt with irradiation (Figure 8; * p < 0.001 vs Control, ** p < 0.001 vs Control, single treatment groups).
실험예 5: 말레이트 금속염의 면역 세포 활성화 효과Experimental Example 5: Immune cell activation effect of maleate metal salt
마우스 CD8 분리 키트 (ThermoFisher)를 활용하여 C57bl/6 마우스의 비장 및 림프 조직으로부터 T 세포를 분리하였다. 상기 T 세포에 20 unit의 IL-2를 단독 또는 2.5 mM CaMal2와 병용 처리한 다음 CD25+, CD69+ 세포를 FACS로 분리하여 활성화된 T 세포를 정량하였다. T cells were isolated from the spleen and lymphoid tissue of C57bl/6 mice using a mouse CD8 isolation kit (ThermoFisher). The T cells were treated with 20 units of IL-2 alone or in combination with 2.5mM CaMal 2 , and then CD25 + and CD69 + cells were separated by FACS to quantify activated T cells.
다음으로, 마우스 NK 세포 분리 키트 (ThermoFisher)를 활용하여 c57bl/6 마우스의 비장으로부터 NK 세포를 분리하였다. 상기 NK 세포에 2 ng/ml의 IL-12를 단독 또는 2.5 mM CaMal2와 병용 처리하여 CD16-, CD56+ 세포를 FACS로 분류하여 활성화된 NK 세포를 정량 하였다.Next, NK cells were isolated from the spleen of c57bl/6 mice using a mouse NK cell isolation kit (ThermoFisher). The NK cells were treated with 2 ng/ml IL-12 alone or in combination with 2.5 mM CaMal 2 , and CD16 - and CD56 + cells were sorted by FACS to quantify activated NK cells.
그 결과, IL-2와 말레이트 금속염을 동시에 처리한 그룹은 IL-2를 단독으로 처리한 그룹에 비해 활성화된 T 세포의 숫자가 유의성 있게 증가하였고 (도 9의 좌측), IL-12와 말레이트 금속염을 동시에 처리한 그룹은 IL-12를 단독으로 처리한 그룹에 비해 활성화된 NK 세포의 숫자가 유의성 있게 증가하는 것을 확인하였다 (도 9의 우측; **p < 0.001 vs low dose IL-2 or IL-12 group). As a result, the number of activated T cells in the group treated simultaneously with IL-2 and malate metal salt significantly increased compared to the group treated with IL-2 alone (left side of Figure 9), and the number of activated T cells was significantly increased compared to the group treated with IL-2 alone (left side of Figure 9). The group treated simultaneously with late metal salts was confirmed to have a significant increase in the number of activated NK cells compared to the group treated with IL-12 alone (right side of Figure 9; ** p < 0.001 vs low dose IL-2 or IL-12 group).
실험예 6: Experimental Example 6: In vivoIn vivo 에서 말레이트 금속염의 항암효과Anticancer effect of maleate metal salt in
6-1. CaMal의 항암효과6-1. Anti-cancer effect of CaMal
Athymic Balb/c 누드 마우스에 1 x 107 개의 췌장암 (Aspc-1) 세포를 이식한 후 약 200 mm3으로 성장하였을 때 20 mg/kg의 CaMal를 주 2 회씩 피하로 투여하여 30 일 동안 종양의 성장을 관찰하였다. After transplanting 1 Growth was observed.
췌장암 (Aspc-1) 세포 이식 마우스에 CaMal를 투여한 결과, 미처리 대조군에 비해 종양 크기가 감소한 것이 육안으로도 명확히 확인되었으며 (도 10의 좌측), 이러한 결과는 통계적으로 유의적인 수준으로 확인되었다 (도 10의 우측; **p < 0.001 vs Control). When CaMal was administered to mice transplanted with pancreatic cancer (Aspc-1) cells, a decrease in tumor size was clearly observed with the naked eye (left side of Figure 10) compared to the untreated control group, and this result was confirmed at a statistically significant level ( Right side of Figure 10; ** p < 0.001 vs Control).
6-2. CaMal6-2. CaMal 22 의 항암효과anticancer effect of
Athymic Balb/c 누드 마우스에 1 x 107 개의 간암 (HepG2) 세포를 이식한 후 약 200 mm3으로 성장하였을 때 80 mg/kg의 CaMal2를 주 5 회씩 구강으로 투여하여 30 일 동안 종양의 성장을 관찰하였다. After transplanting 1 was observed.
간암 (HepG2) 세포 이식 마우스에 CaMal2를 처리한 결과, 미처리 대조군에 비해 종양 크기가 감소한 것이 육안으로도 명확히 확인되었으며 (도 11의 좌측), 이러한 결과는 통계적으로 유의적인 수준으로 확인되었다 (도 11의 우측; **p < 0.001 vs Control).As a result of treating liver cancer (HepG2) cell-transplanted mice with CaMal 2 , a decrease in tumor size compared to the untreated control group was clearly confirmed with the naked eye (left side of Figure 11), and this result was confirmed at a statistically significant level (Figure Right side of 11; ** p < 0.001 vs Control).
6-3. CaMal-DCA의 항암효과6-3. Anticancer effect of CaMal-DCA
Athymic Balb/c 누드 마우스에 1 x 107 개의 유방암 (MDA-MB-231) 세포를 이식한 후 약 200 mm3으로 성장하였을 때 10 mg/kg의 CaMal-DCA를 주 1 회씩 정맥 투여하여 30 일 동안 종양의 성장을 관찰하였다. After transplanting 1 Tumor growth was observed during the period.
유방암 (MDA-MB-231) 세포 이식 마우스에 CaMal-DCA를 처리한 결과, 미처리 대조군에 비해 종양 크기가 감소한 것이 육안으로도 명확히 확인되었으며 (도 12의 좌측), 이러한 결과는 통계적으로 유의적인 수준으로 확인되었다 (도 12의 우측; **p < 0.001 vs Control).When mice transplanted with breast cancer (MDA-MB-231) cells were treated with CaMal-DCA, a decrease in tumor size was clearly observed with the naked eye compared to the untreated control group (left side of Figure 12), and this result was statistically significant. (right side of Figure 12; ** p < 0.001 vs Control).
이상의 설명으로부터, 본 발명의 속하는 기술분야의 당업자는 본 발명이 그 기술적 사상이나 필수적 특징을 변경하지 않고서 다른 구체적인 형태로 실시될 수 있다는 것을 이해할 수 있을 것이다. 이와 관련하여, 이상에서 기술한 실시예들은 모든 면에서 예시 적인 것이며 한정적인 것이 아닌 것으로 이해해야만 한다. 본 발명의 범위는 상기 상세한 설명보다는 후술하는 특허 청구범위의 의미 및 범위 그리고 그 등가 개념으로부터 도출되는 모든 변경 또는 변형된 형태가 본 발명의 범위에 포함되는 것으로 해석되어야 한다.From the above description, those skilled in the art will understand that the present invention can be implemented in other specific forms without changing its technical idea or essential features. In this regard, the embodiments described above should be understood in all respects as illustrative and not restrictive. The scope of the present invention should be construed as including the meaning and scope of the patent claims described below rather than the detailed description above, and all changes or modified forms derived from the equivalent concept thereof are included in the scope of the present invention.

Claims (13)

  1. 말레이트 금속염을 유효성분으로 포함하는 암의 예방 또는 치료용 약학적 조성물.A pharmaceutical composition for preventing or treating cancer containing maleate metal salt as an active ingredient.
  2. 제1항에 있어서, According to paragraph 1,
    상기 말레이트 금속염은 칼슘 말레이트 또는 칼슘 디말레이트인 것인, 암의 예방 또는 치료용 약학적 조성물.A pharmaceutical composition for preventing or treating cancer, wherein the malate metal salt is calcium malate or calcium dimaleate.
  3. 제1항에 있어서, According to paragraph 1,
    상기 암은 간암, 폐암, 대장암, 뇌암, 신장암, 췌장암, 유방암 및 암성 당뇨로 구성된 군으로부터 선택된 1 종 이상인, 암의 예방 또는 치료용 약학적 조성물.A pharmaceutical composition for preventing or treating cancer, wherein the cancer is one or more types selected from the group consisting of liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, breast cancer, and cancerous diabetes.
  4. 제1항에 있어서, According to paragraph 1,
    상기 조성물은 DCA (Dichloroacetic acid)를 추가로 포함하는 것인, 암의 예방 또는 치료용 약학적 조성물.The composition is a pharmaceutical composition for preventing or treating cancer, which further includes DCA (Dichloroacetic acid).
  5. 제1항에 있어서,According to paragraph 1,
    상기 조성물은 피하, 경구 또는 정맥 내로 투여되는 것인, 암의 예방 또는 치료용 약학적 조성물.The composition is a pharmaceutical composition for preventing or treating cancer, which is administered subcutaneously, orally, or intravenously.
  6. 말레이트 금속염을 유효성분으로 포함하는, 제2항암제에 대한 반응성을 증진시키는 항암 보조용 약학적 조성물.An auxiliary anti-cancer pharmaceutical composition that improves reactivity to a second anti-cancer agent, comprising maleate metal salt as an active ingredient.
  7. 제6항에 있어서, According to clause 6,
    상기 말레이트 금속염은 칼슘 말레이트 또는 칼슘 디말레이트인 것인, 항암 보조용 약학적 조성물.A pharmaceutical composition for adjuvant anticancer use, wherein the malate metal salt is calcium malate or calcium dimaleate.
  8. 제6항에 있어서, 상기 제2항암제는 표적 항암제 또는 면역 항암제인 것인, 항암 보조용 약학적 조성물.The pharmaceutical composition for adjuvant anticancer use according to claim 6, wherein the second anticancer agent is a targeted anticancer agent or an immunological anticancer agent.
  9. 제6항에 있어서,According to clause 6,
    상기 제2항암제가 표적으로 하는 암은 간암, 폐암, 대장암, 뇌암, 신장암, 췌장암, 유방암 및 암성 당뇨로 구성된 군으로부터 선택된 1 종 이상인 것인, 항암 보조용 약학적 조성물.The cancer targeted by the second anticancer agent is at least one selected from the group consisting of liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, breast cancer, and cancerous diabetes.
  10. 말레이트 금속염 및 제2항암제를 유효성분으로 포함하는, 암의 예방 또는 치료용 약학적 조성물.A pharmaceutical composition for preventing or treating cancer, comprising a maleate metal salt and a second anticancer agent as active ingredients.
  11. 말레이트 금속염을 유효성분으로 포함하는, 방사선 항암 치료에 대한 반응성을 증진시키는 항암 보조용 약학적 조성물.An adjuvant anticancer pharmaceutical composition that improves responsiveness to radiation anticancer treatment, comprising maleate metal salt as an active ingredient.
  12. 제11항에 있어서, According to clause 11,
    상기 말레이트 금속염은 칼슘 말레이트 또는 칼슘 디말레이트인 것인, 항암 보조용 약학적 조성물.A pharmaceutical composition for adjuvant anticancer use, wherein the malate metal salt is calcium malate or calcium dimaleate.
  13. 제11항에 있어서,According to clause 11,
    상기 방사선 항암 치료가 표적으로 하는 암은 암은 간암, 폐암, 대장암, 뇌암, 신장암, 췌장암, 유방암 및 암성 당뇨로 구성된 군으로부터 선택된 1종 이상인, 항암 보조용 약학적 조성물.The cancer targeted by the radiation anticancer treatment is one or more types selected from the group consisting of liver cancer, lung cancer, colon cancer, brain cancer, kidney cancer, pancreatic cancer, breast cancer, and cancerous diabetes, and a pharmaceutical composition for adjuvant anticancer use.
PCT/KR2022/014491 2022-09-27 2022-09-27 Anticancer composition comprising maleate metal salt WO2024071464A1 (en)

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KR20040085196A (en) * 2002-02-22 2004-10-07 쉐링 코포레이션 Pharmaceutical formulations of antineoplastic agents, in particular temozolomide, processes of making and using the same
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KR20180103735A (en) * 2017-03-09 2018-09-19 동국대학교 산학협력단 Novel compound having malate dehydrogenases (MDH)-inhibitory activity and pharmaceutical composition for preventing or treating cancer comprising the same as an active ingredient
KR101998246B1 (en) * 2018-08-22 2019-07-10 주식회사 메타파인즈 A pharmaceutical composition for treating cancer comprising an ionic compound having metal ions binding thereto
KR20220034505A (en) * 2020-09-11 2022-03-18 연세대학교 산학협력단 Pharmaceutical composition for the death of cancer origin cell

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030003168A1 (en) * 1996-03-13 2003-01-02 Mark Empie Method of preparing and using compositions extracted from vegetable matter for the treatment of cancer
KR20040085196A (en) * 2002-02-22 2004-10-07 쉐링 코포레이션 Pharmaceutical formulations of antineoplastic agents, in particular temozolomide, processes of making and using the same
US20040241107A1 (en) * 2003-05-28 2004-12-02 Burzynski Stanislaw R. Toothpaste containing anticancer agents
KR20180103735A (en) * 2017-03-09 2018-09-19 동국대학교 산학협력단 Novel compound having malate dehydrogenases (MDH)-inhibitory activity and pharmaceutical composition for preventing or treating cancer comprising the same as an active ingredient
KR101998246B1 (en) * 2018-08-22 2019-07-10 주식회사 메타파인즈 A pharmaceutical composition for treating cancer comprising an ionic compound having metal ions binding thereto
KR20220034505A (en) * 2020-09-11 2022-03-18 연세대학교 산학협력단 Pharmaceutical composition for the death of cancer origin cell

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