WO2022010035A1 - Pharmaceutical composition for anticancer comprising herbal extract - Google Patents

Pharmaceutical composition for anticancer comprising herbal extract Download PDF

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WO2022010035A1
WO2022010035A1 PCT/KR2020/013083 KR2020013083W WO2022010035A1 WO 2022010035 A1 WO2022010035 A1 WO 2022010035A1 KR 2020013083 W KR2020013083 W KR 2020013083W WO 2022010035 A1 WO2022010035 A1 WO 2022010035A1
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weight
parts
cancer
pharmaceutical composition
anticancer
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PCT/KR2020/013083
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French (fr)
Korean (ko)
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손형섭
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주식회사 얼라셀바이오
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/35—Caprifoliaceae (Honeysuckle family)
    • A61K36/355—Lonicera (honeysuckle)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/02—Algae
    • A61K36/04—Rhodophycota or rhodophyta (red algae), e.g. Porphyra
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/13—Coniferophyta (gymnosperms)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/23—Apiaceae or Umbelliferae (Carrot family), e.g. dill, chervil, coriander or cumin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/28—Asteraceae or Compositae (Aster or Sunflower family), e.g. chamomile, feverfew, yarrow or echinacea
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/48—Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
    • A61K36/484—Glycyrrhiza (licorice)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/53—Lamiaceae or Labiatae (Mint family), e.g. thyme, rosemary or lavender
    • A61K36/535—Perilla (beefsteak plant)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/70—Polygonaceae (Buckwheat family), e.g. spineflower or dock
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/75—Rutaceae (Rue family)
    • A61K36/752—Citrus, e.g. lime, orange or lemon
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/78—Saururaceae (Lizard's-tail family)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/10—Preparation or pretreatment of starting material
    • A61K2236/19—Preparation or pretreatment of starting material involving fermentation using yeast, bacteria or both; enzymatic treatment
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2300/00—Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00

Definitions

  • the present invention relates to an anticancer pharmaceutical composition comprising a herbal extract.
  • Cancer is one of the incurable diseases that civilization must solve, and huge capital is being invested in development to cure it worldwide. diagnosed, and more than 60,000 people have died. Examples of carcinogens that cause cancer include smoking, ultraviolet rays, chemicals, food, and other environmental factors. Substances currently used as therapeutic agents have considerable toxicity and cannot selectively remove only cancer cells, so there is an urgent need to develop an effective anticancer agent with less toxicity to prevent cancer as well as treatment after the onset of cancer. .
  • an object of the present invention is to provide a pharmaceutical composition for anticancer that has not only fewer side effects when administered to the human body, including herbal extracts, but also excellent apoptosis, in particular, apoptosis effect on cancer cells.
  • the present invention provides a mixed medicinal agent comprising at least one selected from the group consisting of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yakmomil, yew leaf, sorijae, licorice root, and perilla leaf; Or it provides a pharmaceutical composition for anticancer comprising the herbal extract of the mixed drug as an active ingredient.
  • the mixed medicinal material may include gold and silver flower, silk grass, tangerine peel, sunflower stem, centella asiatica, yam wheat, yew leaf, sorijae, licorice root, and perilla leaf
  • the mixed medicine is based on 10 parts by weight of gold and silver coin, silk grass 8-12 parts by weight, tangerine peel 6-8 parts by weight, sunflower stem 6-8 parts by weight, Centella asiatica 6-6.5 parts by weight, 4 to 6.5 parts by weight of buckwheat flour, 4-5 parts by weight of yew leaves, 4-5 parts by weight of sagebrush It may contain parts by weight, licorice root 1-3 parts by weight, and perilla leaf 0.1-2 parts by weight.
  • the herbal extract may be fermented.
  • the anticancer pharmaceutical composition according to the present invention can be applied to one or more solid cancers selected from the group consisting of lung cancer, colorectal cancer, breast cancer, uterine cancer, liver cancer, stomach cancer, bile duct cancer and kidney cancer.
  • the present invention is from a mixed medicine comprising at least one selected from the group consisting of gold and silver flowers, silk grass, tangerine rinds, sunflower stems, centella asiatica, yammilk, yew leaves, sorijai, licorice root and perilla leaf. It provides a method for preparing a pharmaceutical composition for anticancer, comprising the step of obtaining a herbal extract using water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.
  • the mixed medicinal material includes all of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yammilk, yew leaves, sorijai, licorice root and perilla leaf, 8 to 12 parts by weight of silk grass, Tangerine peel 6-8 parts by weight, sunflower stalk 6-8 parts by weight, Centella asiatica 6-6.5 parts by weight, Yak wheat 4 to 6.5 parts by weight, yew leaves 4-5 parts by weight, Sorijae 4-5 parts by weight, Licorice root 1- It may contain 3 parts by weight and 0.1 to 2 parts by weight of perilla leaf.
  • the method for preparing the anticancer pharmaceutical composition according to the present invention may further include, after the step of obtaining the herbal extract, fermenting the obtained herbal extract.
  • the fermenting step is,
  • It may include fermenting the prepared culture with Lactobacillus.
  • the method for preparing the anticancer pharmaceutical composition according to the present invention may further include the step of aging the fermented extract for 2 weeks or more after the step of fermenting the herbal extract.
  • the pharmaceutical composition for anticancer according to the present invention is a herbal extract derived from a mixed drug comprising at least one of gold and silver lilies, silk grass, tangerine peel, sunflower stem, centella asiatica, yew millet, yew leaf, sorijai, licorice root and perilla leaf.
  • a mixed drug comprising at least one of gold and silver lilies, silk grass, tangerine peel, sunflower stem, centella asiatica, yew millet, yew leaf, sorijai, licorice root and perilla leaf.
  • Example 1 is a graph showing the cell viability of the normal cell line (CCD-986sk) according to the concentration of the composition prepared in Example 2.
  • Example 2 is a graph showing the cell viability of the normal cell line (WI-38) according to the concentration of the composition prepared in Example 2.
  • Example 3 is an image showing the apoptosis effect of the composition prepared in Example 2 on the gastric cancer cell line (AGS).
  • Example 4 is an image showing the cell death effect of the composition prepared in Example 2 on the breast cancer cell line (MCF7).
  • Example 5 is an image showing the apoptosis effect of the composition prepared in Example 2 on the lung cancer cell line (A549).
  • the term comprise, comprises, comprising is meant to include the stated object, step or group of objects, and steps, and any other object. It is not used in the sense of excluding a step or a group of objects or groups of steps.
  • the term “treat”, “treating” or “treatment” refers to alleviating, attenuating or ameliorating one or more signs of a disease or condition, preventing further signs, or inhibiting the disease or symptom. For example, preventing the development of a disease or condition, alleviating the disease or symptom, causing regression of the disease or symptom, alleviating the symptoms caused by the disease or symptom, or prophylactically and/or therapeutically stopping the symptoms of a disease or condition.
  • composition for anticancer is provided.
  • the present invention provides a mixed medicinal agent comprising at least one selected from the group consisting of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yakmomil, yew leaf, sorijae, licorice root, and perilla leaf; Or it provides a pharmaceutical composition for anticancer comprising the herbal extract of the mixed drug as an active ingredient.
  • the anticancer pharmaceutical composition according to the present invention is a mixed drug comprising one or more drugs;
  • a mixed drug comprising one or more drugs;
  • side effects were significantly low and exhibited a high apoptosis effect on cancer cells even at low concentrations that did not show toxicity to normal cells.
  • the mixed medicinal material includes at least one of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yammilk, yew leaf, sorijai, licorice root, and perilla leaf, in some cases, all of the above medicinal substances may be included. have.
  • the Lonicera Japonica (Honeysuckle) Flower is a dried bud of Lonicera japonica Thunb. belonging to the Caprifoliaceae family or a flower that has just begun to bloom, and is used in various traditional herbal medicine prescriptions, It is known to have excellent anti-inflammatory effects.
  • the Ceramium Kondoi has a plant body of 5 to 50 cm, and a slightly thick thread-like side branch emerges in a columnar shape and branches in turn and spreads in a fan shape. In the armpit area of the upper branch, small branches may come out alternately.
  • a branch consists of a group of cells arranged in a row. The body color is reddish-purple, the older the individual, the darker the red and the rougher the texture. It is a perennial plant that grows abundantly in spring and summer and declines in autumn and winter. It is collected and used as an auxiliary material for the manufacture of agar.
  • the tangerine peel (Citrus Unshiu Peel) is a dried tangerine peel, and contains bioflavonoids and carotenoids along with fructose and vitamins, thereby lowering the absorption of fat and cholesterol, which is effective in arteriosclerosis, hyperlipidemia, diabetes, etc. It is known to have anti-inflammatory and antiviral properties.
  • sunflower stem Helianthus Annuus (Sunflower) Stem
  • sunflower stem generally contains cellulose, hemicellulose and lignin as structural components, and in addition, it is known to have pectin, essential oil component and ash that can be extracted in some water, so it is used for animal feed
  • pectin, essential oil component and ash that can be extracted in some water, so it is used for animal feed
  • pectin, essential oil component and ash that can be extracted in some water, so it is used for animal feed
  • it has been used as a diuretic, antitussive, and hemostatic effect in the stem, and has been used for urinary incontinence, urolithiasis, bladder stones, renal calculi, hypoxia, pertussis, and traumatic bleeding.
  • centella asiatica is a perennial creeping herb belonging to the Umbrella family, and in oriental medicine, it has been used as a medicine for skin diseases, antipyretics, hemoptysis, diuretics, tonics, umbilical cords, hypothyroidism, arthritis, and the like.
  • the main known components of Centella asiatica are triterpenic acid sugar esters, and asiaticoside, adecassoside, brahmoside, brahminoside, etc. have been reported, and it is known that asiatic acid, madecassic acid, thankunic acid, isothankunic acid, etc. are produced during hydrolysis.
  • Centella asiatica As the main components of Centella asiatica, asiaticoside and madecassoside, which are pentacyclic triterpene glycosides belonging to the ⁇ -amyrin-ursolic acid group, have been used for the treatment of skin wounds and chronic ulcers for a long time. It has been reported to be effective in treating psychiatric diseases, tuberculosis, venous disease, and dementia. Due to these effects, Centella asiatica is used for various purposes such as food, skin treatment, wound treatment, memory enhancer, and tonic, etc.
  • the yakmomil (Houttuynia Cordata) is also called eoseongcho, while it looks similar to buckwheat and has excellent pharmacological effects, so it is called yakmomil.
  • the yakmomil is used in folklore as a diuretic and anthelmintic for plants before flowering, crushed leaves and applied when bitten by boils and poisonous insects, and is used for swelling, suppuration or hemorrhoids. In oriental medicine, it is used for gonorrhea, enteritis, urinary tract infection, pneumonia or bronchitis.
  • the yew leaf (Taxus Cuspidata Leaf) is a leaf of a yew tree that lives in a deep mountain, runs in a spiral shape in a row, but is arranged in two rows like a feather on a branch extending to the side, 1.5-2.5 mm in length, and 2 in width It is ⁇ 3 mm, the surface is dark green, and there is a yellow-green line on the back side. The veins protrude on both sides, and on the back side, there is a light yellow pore line between the edge and the midrib.
  • yew leaves are bitter in taste, sun-dried decoctions are effective in treating diabetes by lowering blood sugar and diuretic effects.
  • sorijae (Rumex Crispus Root) is a perennial grass of the family Candidiaceae that mainly inhabits Korea, Japan, China, North Africa, etc., and it is said that the sorijae is effective for acute hepatitis and chronic bronchitis, hemostasis, hematemesis, It is used for uterine bleeding, thrombocytopenic purpura, etc., and has an effect on constipation.
  • it is known to have herbicidal activity, so it is used as a herbicide.
  • the licorice root (Glycyrrhiza Uralensis (Licorice) Root) is the root of licorice, and licorice is a medicinal plant belonging to the legume family, and the roots and rhizomes are used as they are, or the bark is removed. Licorice has been used for the purpose of relief, pain relief, and detoxification, and the extract is known to have effects such as brain cell protection and anti-inflammatory properties.
  • perilla frutescens Leaf is the leaf and the tip branch of the leaf or wrinkled leaf belonging to the family Labiatae, and it is dried and used for medicinal purposes.
  • Perilla leaves have traditionally been used to treat chills, fever, cough, asthma, and chest tightness caused by colds, as well as food poisoning caused by fish and crabs. can indicate
  • the mixed medicine used in the present invention may include one or more of the above-mentioned medicines, and specifically, it may be used by mixing gold and silver coins and the above-mentioned other medicinal substances.
  • the mixed medicinal material may be used by mixing gold and silver flowers, sunflower stems and sagebrush, and as another example, all of the above-mentioned medicinal materials may be mixed and used.
  • each of the drugs included in the mixed medicine is based on 10 parts by weight of gold and silver coins, 8 to 12 parts by weight of silk grass, 6 to 8 parts by weight of tangerine peel, 6 to 8 parts by weight of sunflower stem, 6 to 6.5 parts by weight of centella asiatica, 4 parts by weight of ryegrass. -6.5 parts by weight, 4-5 parts by weight of yew leaves, 4-5 parts by weight of sagebrush, 1-3 parts by weight of licorice root, and 0.1-2 parts by weight of perilla leaf.
  • the present invention can further improve the apoptosis ability of the composition against cancer cells by controlling the components and content of the mixed drug within the above range.
  • the herbal extract may be fermented.
  • the herbal extract may be a fermented product obtained by mixing an extract obtained from a mixed drug with Bacillus bacteria, culturing the bacteria, adding sterilization and sugar to the obtained culture, and then fermenting the extract with Lactobacillus bacteria.
  • the anticancer pharmaceutical composition according to the present invention can effectively suppress the invasion or proliferation of various bacteria that cause decay or degeneration, including fermented herbal extracts, as well as improve the absorption rate of the human body to realize a higher anticancer effect. .
  • the anticancer pharmaceutical composition may be used for preventing and/or treating one or more solid cancer diseases selected from the group consisting of lung cancer, colon cancer, breast cancer, uterine cancer, liver cancer, stomach cancer, bile duct cancer and kidney cancer.
  • the anticancer pharmaceutical composition is not particularly limited in its formulation, but may preferably be in a liquid form.
  • suitable routes of administration may include oral, intravenous, rectal, aerosol, parenteral, ocular, pulmonary, transmucosal, intradermal, vaginal, otic, nasal and topical administration. Specifically, oral administration or topical administration may be performed.
  • the parenteral delivery may include intramuscular, subcutaneous, intravenous, intramedullary injections, as well as intrathecal, direct intraventricular, intraperitoneal, intraoral and intranasal injections.
  • administration may be in a topical rather than systemic manner, often as a depot preparation or sustained release formulation, for example, via injection of the compound directly into the organ.
  • long-acting formulations may be administered by implantation (eg, subcutaneously or intramuscularly) or by intramuscular injection.
  • the anticancer pharmaceutical composition may be delivered to a target drug delivery system, for example, a liposome coated with a period-specific antibody.
  • a target drug delivery system for example, a liposome coated with a period-specific antibody.
  • the liposome can optionally be targeted and selected by the organ.
  • the pharmaceutical composition according to the present invention may be provided in the form of a rapid release formulation, in the form of an extended release formulation, or in the form of an intermediate release formulation, and may be administered topically. .
  • the anticancer pharmaceutical composition of the present invention may be administered parenterally or intravenously, in which case it may be administered by injection.
  • the anticancer pharmaceutical composition may be administered orally.
  • the anticancer pharmaceutical composition of the present invention can be administered for a long period of time, including throughout the patient's life, to improve or otherwise control or limit the symptoms of the patient's disease or symptoms at the discretion of the doctor when the patient's symptoms are not improved. may be If the patient's condition improves, the administration of the composition may, at the discretion of the physician, be temporarily and continuously discontinued for a specified period of time (ie, a “drug resting period”).
  • the therapeutic efficacy and toxicity of such treatment regimens can be measured in cell culture or It can be measured by standard pharmaceutical procedures in laboratory animals.
  • the dose ratio between the toxic and therapeutic effects is the therapeutic index, which may be the ratio between LD50 and ED50.
  • Results obtained from cell culture assays and animal studies can be used to determine dosage ranges for use in humans.
  • the dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with minimal toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration employed.
  • the anticancer pharmaceutical composition described in the present invention does not have to be administered in the same pharmaceutical composition, and in some cases, different physical and Due to their chemical nature, they may be administered by different routes.
  • initial administration is carried out according to established protocols and then, depending on the effect observed, the dosage, mode of administration and time of administration can be adjusted by the skilled person.
  • the therapeutically-effective dosage may vary when the drugs are used in combination therapy.
  • Combination therapy may further include periodic treatments that are started and stopped at various time points to assist with the clinical management of the patient.
  • the dosage of the co-administered compound may vary depending on the type of co-drug used, the specific drug used, the disease, disease or condition being treated, and the like.
  • the dosing regimen for treating, preventing or ameliorating the symptom(s) to be alleviated will be modified depending on various factors. Such factors include the subject's disease as well as the subject's age, weight, sex, diet, and medical conditions. Accordingly, the dosing regimens actually used vary widely and may deviate from the dosing regimens described in the present invention.
  • the present invention is from a mixed medicine containing at least one selected from the group consisting of gold and silver chrysanthemum, silk grass, tangerine rind, sunflower stem, centella asiatica, yammilk, yew leaf, sorijai, licorice root, and perilla leaf. It provides a method for preparing a pharmaceutical composition for anticancer comprising the step of obtaining a herbal extract.
  • the method for preparing a pharmaceutical composition for anticancer according to the present invention includes the step of obtaining a herbal extract from a mixed drug.
  • the mixed medicinal material may include at least one of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yew wheat, yew leaf, sorijai, licorice root, and perilla leaf, in some cases including all of the above-mentioned medicinal substances can do.
  • each of the drugs included in the mixed medicine is based on 10 parts by weight of gold and silver coins, 8 to 12 parts by weight of silk grass, 6 to 8 parts by weight of tangerine peel, 6 to 8 parts by weight of sunflower stem, 6 to 6.5 parts by weight of centella asiatica, 4 parts by weight of ryegrass. -6.5 parts by weight, 4-5 parts by weight of yew leaves, 4-5 parts by weight of sagebrush, 1-3 parts by weight of licorice root, and 0.1-2 parts by weight of perilla leaf.
  • extracting the herbal extract may be extracted in a manner commonly used in the art.
  • the extract may be extracted through methods such as filtration, hot water extraction, immersion extraction, reflux cooling extraction, and ultrasonic extraction, and preferably may be prepared using hot water extraction.
  • the hot water extraction may be performed by grinding the washed mixed medicinal material to an appropriate size, putting it in an extraction container, injecting an extraction solvent into the extraction container, and then heating.
  • the extraction solvent water, alcohol, or a mixture thereof may be used, and as the alcohol, an alcohol having 1 to 4 carbon atoms is preferably used.
  • the extraction solvent may use ethanol or methanol, preferably 70 to 80 wt% ethanol, but is not limited thereto.
  • the extraction solvent may be added 1 to 10 times, preferably 1 to 3 times, based on the weight of the mixed drug.
  • the extraction temperature is 80 to 110 °C
  • the extraction time may be 24 to 48 hours
  • the number of times of extraction may be 1 to 5 times.
  • the herbal extract thus extracted may be used by filtration without separate concentration or drying, or may be used as a fermented product obtained by fermenting the herbal extract using a fermented strain.
  • the method for preparing the anticancer pharmaceutical composition according to the present invention may further include fermenting the herbal extract obtained after obtaining the herbal extract, and the fermenting step may be performed by the following process. :
  • the step of preparing the culture may be performed by uniformly mixing the herbal extract and the Bacillus strain, specifically, Bacillus subtilis , and culturing the same, wherein the culture temperature of the herbal extract and the Bacillus strain may be appropriately and selectively used within the range conventionally applied in the art, specifically, it may be carried out by culturing for 24 to 72 hours at a temperature of 30 to 60 °C, preferably 35 to 55 °C.
  • the herbal extract cultured with the Bacillus strain is freeze-dried, sterilized plant raw materials and a carbon source (0.025 to 0.06 wt% based on the total weight) are added to the freeze-dried product, and then fermented with Lactobacillus. It can be done by
  • the fermentation process is not particularly limited and may be carried out under conditions commonly applied in the art, specifically, it may be carried out for 24 to 72 hours under conditions of pH 5.0 to 7.5 and 20 to 70° C., more specifically As such, it may be carried out for 24 to 72 hours under conditions of pH 6.0 to 7.0 and 25 to 60 °C or 30 to 50 °C.
  • the fermentation temperature may be varied within the range of 20 ⁇ 70 °C.
  • the fermentation process is performed for 48 hours, the temperature of which is 20° C. to 60° C. for 24 hours; or from 30°C to 40°C.
  • the herbal extracts prepared by the method include various enzymes biosynthesized in the body of the strain, that is, intracellular enzymes in the microbial cells; Existing microorganisms can contain a significant amount of extracellular enzymes and enzymes released from dead cells, and amino acids, proteins, vitamins, minerals, unsaturated fatty acids, Carbohydrates, dietary fiber, organic acids and bio-available physiologically active ingredients, digestive enzymes amylase, protease, lipase, cellulase, or other enzymes are produced in large amounts, thereby exhibiting excellent bioavailability.
  • the method for preparing the anticancer pharmaceutical composition according to the present invention may further include the step of aging the fermented herbal extract, and the aging may be performed at 20-40° C. for 2 weeks or more.
  • each of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yew millet, yew leaf, sorijae, licorice root, and perilla leaf was washed, cut to an appropriate size, and weighed according to the content ratio shown in Table 1 below. It was placed in the extraction vessel so that the total weight was 7 kg. Thereafter, sterile purified water (21 L), which is three times the total weight of the mixed drug contained in the extraction container, was poured, and the mixture was heated to 100 ⁇ 2° C. to obtain a herbal extract. The obtained herbal extract was filtered to prepare an anticancer pharmaceutical composition.
  • Example 1 Each herbal extract prepared in Example 1 and Comparative Examples 1 to 9 was filtered to remove solids, and 0.2 kg of Bacillus subtilis was inoculated, mixed uniformly, and injected in an incubator, cultured at 30 ⁇ 1° C. and humidity. The strain was cultured at 55 ⁇ 1% for 24 hours.
  • MTT assay was performed on the compositions obtained in Examples 1 to 2 and Comparative Examples 1 to 18.
  • human fibroblasts 1 human skin normal cell line (CCD-986sk) and 2 WI-38 cells, were respectively cultured in DMEM (Dulbecco's Modefied Eagle's Medium, Gibco-BRL) medium with 10% FBS (Fetal Bovine Serum, Hyclone). )) and 1% penicillin/streptomycin (Sigma) was added to the cultured in an incubator (Forma) at 37° C. and 5% CO 2 condition.
  • the human fibroblasts were each plated in a 96-well plate at a density of 5 ⁇ 10 3 cells/well, and then cultured for 1 hour in DMEM medium from which serum was removed before sample treatment.
  • compositions of Examples and Comparative Examples had low cytotoxicity close to that of the control group, indicating that the absorbance change rate was significantly low.
  • the composition of Example 2 had little cytotoxicity at a concentration of 12 vol.% or less for the CCD-986sk cell line and 17 vol.% or less for the WI-38 cell line, as shown in FIGS. 1 and 2 . .
  • the anticancer pharmaceutical composition according to the present invention contains a herbal extract as an active ingredient, and thus has remarkably low cytotoxicity.
  • the MTT assay was performed on the anticancer pharmaceutical compositions obtained in Examples 1 to 2 and Comparative Examples 10 to 18.
  • lung cancer cell line A549), colon cancer cell line (HCT116), uterine cancer cell line (HeLa), cholangiocarcinoma cell line (HuCC-T1), gastric cancer cell line (AGS), liver cancer cell line (HepG2), breast cancer cell line (MCF7) and A renal cancer cell line (HEK239) was prepared, and each cancer cell line was cultured in a 100 mm dish at 37° C., 5% CO 2 in an incubator.
  • the culture medium for each cell line was cultured using DMEM (Dulbeco's Modified Eagle's Medium) medium supplemented with 2 mg/ml sodium bicarbonate, 10% Fetal Bovine Serum, and 1% penicillin-streptomycin. .
  • DMEM Dulbeco's Modified Eagle's Medium
  • Each cultured cancer cell line was cultured in a 96-well plate, and the compositions of Examples 1 to 2 and Comparative Examples 10 to 18 were added to the cultured cell line at a concentration of 1, 2, 5, 10, 15, 20 and 25 vol.%, respectively.
  • Treated with 37 °C, 5% CO 2 Incubated for 24 hours in an incubator. After incubation, 20 ⁇ l of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] reagent was treated and incubated for 4 hours. Absorbance was measured at 490 nm with an ELISA reader. , The cancer cell death rate was calculated as a percentage compared to the absorbance of the control group.
  • the cancer cell death rates according to the treatment concentration of the composition of Example 2 are shown in Table 2, and the cancer cell death rates of the compositions of Examples and Comparative Examples at a concentration of 15 vol.% are shown in Table 3 and FIGS. 3 to 5. indicated.
  • the anticancer pharmaceutical composition according to the present invention has an excellent killing effect on cancer cells.
  • Example 2 exhibited a cancer cell death rate of 60% or more even at concentrations of 10 vol.% and 15 vol.% that do not show toxicity to normal cells, particularly lung cancer and cholangiocarcinoma. , it was confirmed that the cancer cell death rate of 80% or more for gastric cancer and breast cancer.
  • Example 2 in which the herbal extract was fermented had a higher rate of cancer cell death compared to the composition of Example 1 in which fermentation was not performed, and the compositions of Comparative Examples 10 to 18 It was confirmed to have a significantly higher rate of cancer cell death even at a low concentration of 15 vol.% compared to .
  • the anticancer pharmaceutical composition according to the present invention is cytotoxic by including extracts of gold and silver lilies, silk grass, tangerine peels, sunflower stems, centella asiatica, yamba wheat, yew leaves, sorijai, licorice root and perilla leaf in a specific content. Since the apoptosis effect on cancer cells is very low as well as significantly excellent, it can be usefully used as a substance for preventing and/or treating solid cancers such as lung cancer, colon cancer, breast cancer, uterine cancer, liver cancer, stomach cancer, bile duct cancer, and kidney cancer.
  • the pharmaceutical composition for anticancer according to the present invention is a herbal extract derived from a mixed medicine containing at least one of gold and silver lilies, silk grass, tangerine peel, sunflower stem, centella asiatica, ryegrass, yew leaves, sorijai, licorice root and perilla leaf.
  • a mixed medicine containing at least one of gold and silver lilies, silk grass, tangerine peel, sunflower stem, centella asiatica, ryegrass, yew leaves, sorijai, licorice root and perilla leaf.

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Abstract

The present invention relates to a pharmaceutical composition for anticancer comprising a herbal extract, wherein the pharmaceutical composition for anticancer includes an extract derived from a mixed medicine including Lonicera japonica, Ceramium kondoi, tangerine peels, sunflower stems, Centella asiatica, Houttuynia cordata thumb, leaves of Taxus cuspidata, Rumex Crispus L., Glycyrrhizae radix, and Perilla frutescens leaves, and thus, when administered to the human body, side effects are small and the killing effect on cancer cells is excellent, so that the pharmaceutical composition can be usefully used for solid cancer diseases such as lung cancer, colon cancer, breast cancer, uterine cancer, liver cancer, gastric cancer, cholangiocarcinoma, and kidney cancer.

Description

생약 추출물을 포함하는 항암용 약학적 조성물Pharmaceutical composition for anticancer comprising herbal extract
본 발명은 생약 추출물을 포함하는 항암용 약학적 조성물에 관한 것이다.The present invention relates to an anticancer pharmaceutical composition comprising a herbal extract.
암은 인류가 해결해야 할 난치병 중의 하나로, 전 세계적으로 이를 치유하기 위한 개발에 막대한 자본이 투자되고 있는 실정이며, 우리나라의 경우, 질병 사망원인 중 제 1위의 질병으로서 연간 약 10 만 명 이상이 진단되고, 약 6 만 명 이상이 사망하고 있다. 이러한 암의 유발 인자인 발암물질로는 흡연, 자외선, 화학물질, 음식물, 기타 환경인자들이 있으나, 그 유발 원인이 다양하여 치료제의 개발이 어려울 뿐만 아니라 발생하는 부위에 따라 치료제의 효과 또한 각기 다르다. 현재 치료제로 사용되는 물질들은 상당한 독성을 지니고 있으며, 암 세포만을 선택적으로 제거하지 못하므로, 암의 발생 후 이의 치료뿐 아니라, 암의 발생을 예방하기 위한 독성이 적고 효과적인 항암제의 개발이 절실히 필요하다.Cancer is one of the incurable diseases that mankind must solve, and huge capital is being invested in development to cure it worldwide. diagnosed, and more than 60,000 people have died. Examples of carcinogens that cause cancer include smoking, ultraviolet rays, chemicals, food, and other environmental factors. Substances currently used as therapeutic agents have considerable toxicity and cannot selectively remove only cancer cells, so there is an urgent need to develop an effective anticancer agent with less toxicity to prevent cancer as well as treatment after the onset of cancer. .
한편, 보다 안전하고, 치료 효과가 높은 대체적인 암 치료 방법이 요구되고 있으며, 이에 최근에는 안정성이 보장된 식물이나 미생물 유래 등 천연자원을 이용한 기능성 식품 및 의약품 연구에 관심이 집중되고 있다. 실제로 신약개발 기초자원으로 천연물은 주요비중을 차지하고 있으며, 아스피린(aspirin)이 버드나무 추출물에서 유래한 것과 같이 오늘날 사용 약물의 60% 정도가 천연물로부터 유래하고 있다.On the other hand, there is a need for a safer and more effective alternative cancer treatment method, and in recent years, interest in functional foods and pharmaceutical research using natural resources such as plants or microorganisms with guaranteed stability has been focused. In fact, natural products occupy a major proportion as a basic resource for new drug development, and just as aspirin is derived from willow extract, about 60% of drugs used today are derived from natural products.
특히, 생약 추출물의 인체 면역계 조절작용에 대한 연구가 활발히 이루어지고 있는데, 이는 면역계의 이상현상이 건강을 깨뜨리고 다양한 질환을 일으키기 때문이다. 일례로 상백피, 인삼의 사포닌, 표고버섯 등이 면역증강제로, 별불가사리, 영지ㆍ상황버섯, 인삼유래 진세노사이드가 항암 보조제로 연구가 이루어지고 있고, 감초, 감초성분 글리시진(glycyrrhizin), 해삼, 웅담 성분 UDCA 등이 세포분화물질로 연구가 진행되고 있다. 이러한 천연물 소재를 이용한 면역조절제의 개발은, 부작용 없이 암이나 만성적인 염증 질환 등을 안전하게 치료할 수 있기 때문에 많은 사람들의 관심과 흥미를 불러일으키고 있다.In particular, studies on the regulating effect of herbal extracts on the human immune system are being actively conducted, because abnormalities of the immune system damage health and cause various diseases. For example, research is being carried out on sangbaekpi, ginseng saponin, and shiitake mushrooms as immune boosters, starfish, reishi/shanghai mushroom, and ginseng-derived ginsenoside as anticancer adjuvants. Licorice, licorice component glycyrrhizin, sea cucumber , and UDCA, which is a component of succulents, is being studied as a cell differentiation material. The development of immunomodulators using these natural materials is drawing interest and interest from many people because it can safely treat cancer or chronic inflammatory diseases without side effects.
그러나, 이러한 생약 추출물은 인체에 투여 시 부작용이 적은데 반해, 세포 사멸 효과가 크지 않아 실제 사용할 수 있는 유효한 치료 효과를 구현하지 못하므로, 항암제의 활성을 촉진시키는 보조제로서 역할밖에 할 수 없는 한계가 있다.However, while these herbal extracts have few side effects when administered to the human body, they do not have a large apoptosis effect, so they do not realize effective therapeutic effects that can be used in practice, so there is a limit in that they can only serve as an adjuvant to promote the activity of anticancer drugs. .
이에, 본 발명은 생약 추출물을 포함하여 인체에 투여 시 부작용이 적을 뿐만 아니라 세포 사멸, 특히 암세포에 대한 사멸 효과가 우수한 항암용 약학적 조성물을 제공하는데 있다.Accordingly, an object of the present invention is to provide a pharmaceutical composition for anticancer that has not only fewer side effects when administered to the human body, including herbal extracts, but also excellent apoptosis, in particular, apoptosis effect on cancer cells.
상술된 문제를 해결하기 위하여,In order to solve the above-mentioned problem,
본 발명은 일실시예에서, 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽으로 이루어진 군으로부터 선택되는 1종 이상을 포함하는 혼합 약재; 또는 상기 혼합약재의 생약 추출물을 유효성분으로 포함하는 항암용 약학적 조성물을 제공한다.In one embodiment, the present invention provides a mixed medicinal agent comprising at least one selected from the group consisting of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yakmomil, yew leaf, sorijae, licorice root, and perilla leaf; Or it provides a pharmaceutical composition for anticancer comprising the herbal extract of the mixed drug as an active ingredient.
상기 혼합 약재는 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽을 포함할 수 있으며, 이 경우, 상기 혼합 약재는 금은화 10 중량부에 대하여, 비단풀 8~12 중량부, 귤 과피 6~8 중량부, 해바라기 줄기 6~8 중량부, 병풀 6~6.5 중량부, 약모밀 4~6.5 중량부, 주목 잎 4~5 중량부, 소리쟁이 4~5 중량부, 감초근 1~3 중량부 및 자소엽 0.1~2중량부를 포함할 수 있다.The mixed medicinal material may include gold and silver flower, silk grass, tangerine peel, sunflower stem, centella asiatica, yam wheat, yew leaf, sorijae, licorice root, and perilla leaf, in this case, the mixed medicine is based on 10 parts by weight of gold and silver coin, silk grass 8-12 parts by weight, tangerine peel 6-8 parts by weight, sunflower stem 6-8 parts by weight, Centella asiatica 6-6.5 parts by weight, 4 to 6.5 parts by weight of buckwheat flour, 4-5 parts by weight of yew leaves, 4-5 parts by weight of sagebrush It may contain parts by weight, licorice root 1-3 parts by weight, and perilla leaf 0.1-2 parts by weight.
또한, 상기 생약 추출물은 발효된 것일 수 있다.In addition, the herbal extract may be fermented.
아울러, 본 발명에 따른 항암용 약학적 조성물은 폐암, 대장암, 유방암, 자궁암, 간암, 위암, 담관암 및 신장암으로 이루어진 군으로부터 선택되는 1종 이상의 고형암에 적용될 수 있다.In addition, the anticancer pharmaceutical composition according to the present invention can be applied to one or more solid cancers selected from the group consisting of lung cancer, colorectal cancer, breast cancer, uterine cancer, liver cancer, stomach cancer, bile duct cancer and kidney cancer.
나아가, 본 발명은 일실시예에서, 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽으로 이루어진 군으로부터 선택되는 1종 이상을 포함하는 혼합 약재로부터 물, 탄소수 1~4의 알코올 또는 이들의 혼합 용매를 이용하여 생약 추출물을 얻는 단계를 포함하는 항암용 약학적 조성물의 제조방법을 제공한다.Furthermore, in one embodiment, the present invention is from a mixed medicine comprising at least one selected from the group consisting of gold and silver flowers, silk grass, tangerine rinds, sunflower stems, centella asiatica, yammilk, yew leaves, sorijai, licorice root and perilla leaf. It provides a method for preparing a pharmaceutical composition for anticancer, comprising the step of obtaining a herbal extract using water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.
여기서, 상기 혼합 약재는 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽을 모두 포함하는 경우 금은화 10 중량부에 대하여, 비단풀 8~12 중량부, 귤 과피 6~8 중량부, 해바라기 줄기 6~8 중량부, 병풀 6~6.5 중량부, 약모밀 4~6.5 중량부, 주목 잎 4~5 중량부, 소리쟁이 4~5 중량부, 감초근 1~3 중량부 및 자소엽 0.1~2중량부를 포함할 수 있다.Here, when the mixed medicinal material includes all of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yammilk, yew leaves, sorijai, licorice root and perilla leaf, 8 to 12 parts by weight of silk grass, Tangerine peel 6-8 parts by weight, sunflower stalk 6-8 parts by weight, Centella asiatica 6-6.5 parts by weight, Yak wheat 4 to 6.5 parts by weight, yew leaves 4-5 parts by weight, Sorijae 4-5 parts by weight, Licorice root 1- It may contain 3 parts by weight and 0.1 to 2 parts by weight of perilla leaf.
또한, 본 발명에 따른 항암용 약학적 조성물의 제조방법은 상기 생약 추출물을 얻는 단계 이후에, 얻어진 생약 추출물을 발효시키는 단계를 더 포함할 수 있다. In addition, the method for preparing the anticancer pharmaceutical composition according to the present invention may further include, after the step of obtaining the herbal extract, fermenting the obtained herbal extract.
이때, 상기 발효시키는 단계는,In this case, the fermenting step is,
생약 추출물과 바실러스 서브틸리스균을 혼합하여 배양된 배양물을 제조하는 단계; 및Preparing a cultured culture by mixing the herbal extract and Bacillus subtilis; and
제조된 배양물을 락토바실러스균으로 발효시키는 단계를 포함할 수 있다.It may include fermenting the prepared culture with Lactobacillus.
또한, 본 발명에 따른 항암용 약학적 조성물의 제조방법은 상기 생약 추출물을 발효시키는 단계 이후에, 발효된 추출물을 2주 이상 숙성시키는 단계를 더 포함할 수 있다.In addition, the method for preparing the anticancer pharmaceutical composition according to the present invention may further include the step of aging the fermented extract for 2 weeks or more after the step of fermenting the herbal extract.
본 발명에 따른 항암용 약학적 조성물은 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽 중 적어도 1종 이상을 포함하는 혼합 약재로부터 유래된 생약 추출물을 포함하여 인체에 투여 시 부작용이 적을 뿐만 아니라 암세포에 대한 사멸 효과가 우수하므로, 폐암, 대장암, 유방암, 자궁암, 간암, 위암, 담관암, 신장암 등의 고형암 질환에 유용하게 사용될 수 있다.The pharmaceutical composition for anticancer according to the present invention is a herbal extract derived from a mixed drug comprising at least one of gold and silver lilies, silk grass, tangerine peel, sunflower stem, centella asiatica, yew millet, yew leaf, sorijai, licorice root and perilla leaf. When administered to the human body including
도 1은 실시예 2에서 제조된 조성물의 농도에 따른 정상 세포주(CCD-986sk)에 대한 세포 생존률을 나타내는 그래프이다.1 is a graph showing the cell viability of the normal cell line (CCD-986sk) according to the concentration of the composition prepared in Example 2.
도 2는 실시예 2에서 제조된 조성물의 농도에 따른 정상 세포주(WI-38)에 대한 세포 생존률을 나타내는 그래프이다.2 is a graph showing the cell viability of the normal cell line (WI-38) according to the concentration of the composition prepared in Example 2.
도 3은 실시예 2에서 제조된 조성물의 위암 세포주(AGS)에 대한 세포 사멸 효과를 나타내는 이미지이다.3 is an image showing the apoptosis effect of the composition prepared in Example 2 on the gastric cancer cell line (AGS).
도 4는 실시예 2에서 제조된 조성물의 유방암 세포주(MCF7)에 대한 세포 사멸 효과를 나타내는 이미지이다.4 is an image showing the cell death effect of the composition prepared in Example 2 on the breast cancer cell line (MCF7).
도 5는 실시예 2에서 제조된 조성물의 폐암 세포주(A549)에 대한 세포 사멸 효과를 나타내는 이미지이다.5 is an image showing the apoptosis effect of the composition prepared in Example 2 on the lung cancer cell line (A549).
이하에 본 발명을 상세하게 설명하기에 앞서, 본 명세서에 사용된 용어는 특정의 실시예를 기술하기 위한 것일 뿐 첨부하는 특허청구의 범위에 의해서만 한정되는 본 발명의 범위를 한정하려는 것은 아님을 이해하여야 한다. 본 명세서에 사용되는 모든 기술용어 및 과학용어는 다른 언급이 없는 한은 기술적으로 통상의 기술을 가진 자에게 일반적으로 이해되는 것과 동일한 의미를 가진다.Prior to describing the present invention in detail below, it is to be understood that the terminology used herein is for the purpose of describing specific embodiments and is not intended to limit the scope of the present invention, which is limited only by the appended claims. shall. All technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art, unless otherwise stated.
본 명세서 및 청구범위의 전반에 걸쳐, 다른 언급이 없는 한 포함(comprise, comprises, comprising)이라는 용어는 언급된 물건, 단계 또는 일군의 물건, 및 단계를 포함하는 것을 의미하고, 임의의 어떤 다른 물건, 단계 또는 일군의 물건 또는 일군의 단계를 배제하는 의미로 사용된 것은 아니다.Throughout this specification and claims, unless stated otherwise, the term comprise, comprises, comprising is meant to include the stated object, step or group of objects, and steps, and any other object. It is not used in the sense of excluding a step or a group of objects or groups of steps.
본 발명에서, "또는" 또는 "및"의 사용은 별도의 표시가 없으면 "및/또는"을 의미한다. 게다가, "동반하는"이라는 용어뿐만 아니라 다른 형태, 예를 들어 "동반하다" 및 "동반되는"의 사용은 제한되지 않는다. 본 발명에 사용되는 섹션의 제목은 단지 체계화하기 위한 것이며 기술된 대상 물질을 제한하는 것으로 해석되어서는 안 된다.In the present invention, the use of “or” or “and” means “and/or” unless otherwise indicated. Moreover, the use of the term “accompanying” as well as other forms such as “accompanying” and “accompanying” is not limiting. Section headings used in the present invention are for organizational purposes only and should not be construed as limiting the subject matter described.
본 발명에 사용되는 "치료하다", "치료하는" 또는 "치료"란 용어는, 질환 또는 증상의 하나 이상의 징후를 경감하거나 약화하거나 개선하는 것, 추가적인 징후를 예방하는 것, 질환 또는 증상을 억제하는 것, 예를 들어 질환 또는 증상의 발생(development)을 막는 것, 질환 또는 증상을 완화시키는 것, 질환 또는 증상의 퇴행을 야기하는 것, 질환 또는 증상에 의해 야기되는 증상을 완화시키는 것, 또는 예방적으로 및/또는 치료적으로 질환 또는 증상의 징후를 중단시키는 것을 포함한다.As used herein, the term "treat", "treating" or "treatment" refers to alleviating, attenuating or ameliorating one or more signs of a disease or condition, preventing further signs, or inhibiting the disease or symptom. For example, preventing the development of a disease or condition, alleviating the disease or symptom, causing regression of the disease or symptom, alleviating the symptoms caused by the disease or symptom, or prophylactically and/or therapeutically stopping the symptoms of a disease or condition.
이하에서 본 발명을 보다 상세히 설명한다.Hereinafter, the present invention will be described in more detail.
항암용 약학적 조성물Pharmaceutical composition for anticancer
본 발명은 일실시예에서, 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽으로 이루어진 군으로부터 선택되는 1종 이상을 포함하는 혼합 약재; 또는 상기 혼합 약재의 생약 추출물을 유효성분으로 포함하는 항암용 약학적 조성물을 제공한다.In one embodiment, the present invention provides a mixed medicinal agent comprising at least one selected from the group consisting of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yakmomil, yew leaf, sorijae, licorice root, and perilla leaf; Or it provides a pharmaceutical composition for anticancer comprising the herbal extract of the mixed drug as an active ingredient.
본 발명에 따른 항암용 약학적 조성물은 1종 이상의 약재를 포함하는 혼합 약재; 또는 상기 혼합 약재로부터 생약 추출물을 포함하여 이를 섭취하거나 인체에 투여하는 경우 부작용이 현저히 낮을 뿐만 아니라 정상세포에 대하여 독성을 나타내지 않는 낮은 농도에서도 암세포에 대한 높은 사멸 효과를 나타내는 것을 실험적으로 확인하였다.The anticancer pharmaceutical composition according to the present invention is a mixed drug comprising one or more drugs; Alternatively, it was experimentally confirmed that, when ingested or administered to the human body, including herbal extracts from the above mixed drugs, side effects were significantly low and exhibited a high apoptosis effect on cancer cells even at low concentrations that did not show toxicity to normal cells.
이때, 상기 혼합 약재는 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽 중 적어도 1종을 포함하며, 경우에 따라서는 상기 약재를 모두 포함할 수 있다.At this time, the mixed medicinal material includes at least one of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yammilk, yew leaf, sorijai, licorice root, and perilla leaf, in some cases, all of the above medicinal substances may be included. have.
상기 금은화(Lonicera Japonica (Honeysuckle) Flower)는 인동과 (Caprifoliaceae)에 속하는 Lonicera japonica Thunb.의 꽃봉오리 또는 막 피기 시작한 꽃을 건조한 약재로 각종 전통 한약 처방에 이용되고 있으며, 체열 조절, 해열, 해독 및 항염 효과가 뛰어난 것으로 알려져 있다.The Lonicera Japonica (Honeysuckle) Flower is a dried bud of Lonicera japonica Thunb. belonging to the Caprifoliaceae family or a flower that has just begun to bloom, and is used in various traditional herbal medicine prescriptions, It is known to have excellent anti-inflammatory effects.
또한, 상기 비단풀(Ceramium Kondoi)은 식물체는 5∼50cm이고 원기둥 모양으로 약간 두꺼운 실모양의 곁가지가 나와 차례로 분지하여 부채꼴로 퍼진다. 위쪽 가지의 겨드랑이 부분에는 작은 가지가 엇갈려 나오기도 한다. 가지는 한 줄로 늘어선 세포군으로 이루어져 있다. 체색은 홍자색으로 오래된 개체일수록 검붉게 변하며 결이 거칠어진다. 여러해살이로 봄·여름에 무성하며 가을·겨울에 쇠퇴한다. 채집하여 한천(寒天) 제조의 부원료로 이용한다. 한국·일본·사할린 등지의 조간대(潮間帶) 바위 위 또는 다른 해조류에 붙어살며 해안 오염에 따른 서식환경의 악화로 개체군이 점차 줄어들고 있다. 우리나라 전 해안에 분포하나 특히 서남해안에서 흔히 발견된다. 비단풀의 효능으로는 요로결석 제거, 해독작용, 항균작용, 진정작용 감기예방 및 치매예방 등 다양한 효과를 나타낸다.In addition, the Ceramium Kondoi has a plant body of 5 to 50 cm, and a slightly thick thread-like side branch emerges in a columnar shape and branches in turn and spreads in a fan shape. In the armpit area of the upper branch, small branches may come out alternately. A branch consists of a group of cells arranged in a row. The body color is reddish-purple, the older the individual, the darker the red and the rougher the texture. It is a perennial plant that grows abundantly in spring and summer and declines in autumn and winter. It is collected and used as an auxiliary material for the manufacture of agar. It lives on rocks or other seaweeds in the intertidal zone in Korea, Japan, and Sakhalin, and the population is gradually decreasing due to deterioration of the habitat caused by coastal pollution. It is distributed all over the coast of Korea, but is particularly common in the southwest coast. As for the efficacy of silk grass, it shows various effects such as removal of urinary stones, detoxification, antibacterial action, sedative action, cold prevention and dementia prevention.
아울러, 상기 귤 과피(Citrus Unshiu Peel)는 감귤의 껍질을 건조시킨 것으로서, 과당, 비타민과 함께 바이오플라보노이드, 카로티노이드 등을 함유하여 지방, 콜리스테롤의 흡수를 낮춰 동맥경화증, 고지혈증, 당뇨 등에 효과가 있으며, 항염증 및 항바이러스 작용을 하는 것으로 알려져 있다.In addition, the tangerine peel (Citrus Unshiu Peel) is a dried tangerine peel, and contains bioflavonoids and carotenoids along with fructose and vitamins, thereby lowering the absorption of fat and cholesterol, which is effective in arteriosclerosis, hyperlipidemia, diabetes, etc. It is known to have anti-inflammatory and antiviral properties.
또한, 해바라기 줄기(Helianthus Annuus (Sunflower) Stem)는 대체로 셀룰로오스, 헤미셀룰로오스 및 리그닌을 구조적 성분으로 포함하며, 그 외에 일부 물에 추출될 수 있는 펙틴, 정유 성분 및 회분을 가지고 있는 것으로 알려져 있어 동물의 사료용으로 사용되어 왔으며 최근에는 줄기속의 이뇨, 진해, 지혈 효능을 이용하여 소변불리, 요로결석, 방광결석, 신결선, 허소, 백일해, 외상출혈 등의 용도에 사용되고 있다.In addition, sunflower stem (Helianthus Annuus (Sunflower) Stem) generally contains cellulose, hemicellulose and lignin as structural components, and in addition, it is known to have pectin, essential oil component and ash that can be extracted in some water, so it is used for animal feed Recently, it has been used as a diuretic, antitussive, and hemostatic effect in the stem, and has been used for urinary incontinence, urolithiasis, bladder stones, renal calculi, hypoxia, pertussis, and traumatic bleeding.
이와 더불어, 상기 병풀(Centella Asiatica)은 산형과에 속하는 다년생 포복성 초본으로, 한방에서는 피부병, 해열, 각혈, 이뇨제, 강장제, 음위, 대하증, 관절염 등의 약재로 사용 되어 왔다. 병풀의 알려진 주요성분은 triterpenic acid sugar esters로 asiaticoside, adecassoside,brahmoside, brahminoside 등이 보고되어 있으며, 가수분해 시 asiatic acid, madecassic acid, thankunic acid, isothankunic acid 등이 생산되는 것으로 알려져 있다. 병풀의 주요성분으로서 α-amyrin-ursolic acid group에 속하는 pentacyclic triterpene glycoside인 asiaticoside와 madecassoside는 오래 전부터 피부 상처나 만성 궤양 등의 치료에 사용되어 왔으며, 나병 치료 및 항균 효능, 상처, 위궤양, 다양한 피부질병, 정신질환, 결핵, 정맥질환, 치매 등에 대해 치료효과가 있는 것으로 보고되어 있다. 이러한 효능으로 인해 병풀은 식용, 피부치료제, 상처치료제, 기억력 증강제 및 강장제 등 다양한 용도에 이용되고 있고, 국내에서도 의약품, 화장품, 기능성 식품 등의 분야에서 병풀을 사용하고 있다.In addition, the centella asiatica is a perennial creeping herb belonging to the Umbrella family, and in oriental medicine, it has been used as a medicine for skin diseases, antipyretics, hemoptysis, diuretics, tonics, umbilical cords, hypothyroidism, arthritis, and the like. The main known components of Centella asiatica are triterpenic acid sugar esters, and asiaticoside, adecassoside, brahmoside, brahminoside, etc. have been reported, and it is known that asiatic acid, madecassic acid, thankunic acid, isothankunic acid, etc. are produced during hydrolysis. As the main components of Centella asiatica, asiaticoside and madecassoside, which are pentacyclic triterpene glycosides belonging to the α-amyrin-ursolic acid group, have been used for the treatment of skin wounds and chronic ulcers for a long time. It has been reported to be effective in treating psychiatric diseases, tuberculosis, venous disease, and dementia. Due to these effects, Centella asiatica is used for various purposes such as food, skin treatment, wound treatment, memory enhancer, and tonic, etc.
또한, 상기 약모밀(Houttuynia Cordata)은 어성초라고도 불리는데, 메밀과 유사하게 생겼으면서도 약리효과가 우수하여 약모밀이라고 불린다. 상기 약모밀은 민간에서 꽃이 피기 전의 식물체를 이뇨제와 구충제로 사용되고, 잎을 짓찧어 종기와 독충에 물렸을 때 바르며, 부스럼, 화농 또는 치질에 사용되고 있다. 한방에서는 임질, 장염, 요로 감염증, 폐렴 또는 기관지염에 사용한다. In addition, the yakmomil (Houttuynia Cordata) is also called eoseongcho, while it looks similar to buckwheat and has excellent pharmacological effects, so it is called yakmomil. The yakmomil is used in folklore as a diuretic and anthelmintic for plants before flowering, crushed leaves and applied when bitten by boils and poisonous insects, and is used for swelling, suppuration or hemorrhoids. In oriental medicine, it is used for gonorrhea, enteritis, urinary tract infection, pneumonia or bronchitis.
아울러, 상기 주목 잎(Taxus Cuspidata Leaf)은 깊은 산 속에 서식하는 주목 나무의 잎으로서, 줄 모양으로 나선상으로 달리지만 옆으로 뻗은 가지에서는 깃처럼 2줄로 배열하며, 길이 1.5~2.5mm, 너비는 2∼3mm로 표면은 짙은 녹색이고 뒷면에 황록색 줄이 있다. 잎맥은 양면으로 도드라지고 뒷면에는 가장자리와 중륵 사이에 연한 황색의 기공조선(氣孔條線 : 잎이 숨 쉬는 부분으로 보통 잎 뒤에 흰 선으로 나타남)이 있고, 잎은 2∼3년 만에 떨어진다. 주목 잎은 맛이 쓰지만, 햇볕에 말린 것을 달여서 복용하면 혈당치를 내리고 이뇨 작용을 하며 당뇨병 치료에 효능이 있다.In addition, the yew leaf (Taxus Cuspidata Leaf) is a leaf of a yew tree that lives in a deep mountain, runs in a spiral shape in a row, but is arranged in two rows like a feather on a branch extending to the side, 1.5-2.5 mm in length, and 2 in width It is ∼3 mm, the surface is dark green, and there is a yellow-green line on the back side. The veins protrude on both sides, and on the back side, there is a light yellow pore line between the edge and the midrib. Although yew leaves are bitter in taste, sun-dried decoctions are effective in treating diabetes by lowering blood sugar and diuretic effects.
또한, 상기 소리쟁이(Rumex Crispus Root)는 주로 한국, 일본, 중국, 북아프리카 등에 서식하는 마디풀목 마디풀과의 여러해살이 풀로서, 소리쟁이가 급성 간염, 만성 기관지염에 효과가 있다고 하였고, 지혈, 토혈, 자궁출혈, 혈소판 감소성 자반 등에 활용되며, 변비에도 효능이 있다. 또한, 제초활성이 있는 것으로 알려져 있어 제초제로 사용되고 있으며, 지방세포 분화 억제 효과, 항균 효과 및 항산화 효과가 있는 것으로 보고되어 건강기능성 식품에 활용되고 있다.In addition, the sorijae (Rumex Crispus Root) is a perennial grass of the family Candidiaceae that mainly inhabits Korea, Japan, China, North Africa, etc., and it is said that the sorijae is effective for acute hepatitis and chronic bronchitis, hemostasis, hematemesis, It is used for uterine bleeding, thrombocytopenic purpura, etc., and has an effect on constipation. In addition, it is known to have herbicidal activity, so it is used as a herbicide.
나아가, 상기 감초근(Glycyrrhiza Uralensis (Licorice) Root)은 감초의 뿌리로서, 감초는 콩과에 속하는 약용식물로서 뿌리와 뿌리줄기를 그대로 사용하거나 주피를 제거하여 사용한다. 감초는 완화, 진통, 해독 등을 목적으로 사용되어 왔으며, 추출물은 뇌세포보호, 항염 등의 효능이 있는 것으로 알려졌다.Furthermore, the licorice root (Glycyrrhiza Uralensis (Licorice) Root) is the root of licorice, and licorice is a medicinal plant belonging to the legume family, and the roots and rhizomes are used as they are, or the bark is removed. Licorice has been used for the purpose of relief, pain relief, and detoxification, and the extract is known to have effects such as brain cell protection and anti-inflammatory properties.
또한, 상기 자소엽(Perilla frutescens Leaf)은 꿀풀과(Labiatae)에 속하는 차즈기 또는 주름소엽의 잎 및 끝 가지이며, 이를 말려서 약용으로 사용한다. 자소엽은 전통적으로 감기로 인한 오한, 발열, 기침, 천식, 가슴이 그득한 증상과 물고기와 게 등으로 인한 식중독의 치료에 사용하였으며, 현대적으로 해열작용, 항균작용, 혈당수치의 상승 등의 약리작용을 나타낼 수 있다.In addition, the perilla frutescens Leaf is the leaf and the tip branch of the leaf or wrinkled leaf belonging to the family Labiatae, and it is dried and used for medicinal purposes. Perilla leaves have traditionally been used to treat chills, fever, cough, asthma, and chest tightness caused by colds, as well as food poisoning caused by fish and crabs. can indicate
한편, 본 발명에서 사용되는 상기 혼합 약재는 상술된 약재 중 1종 이상을 포함할 수 있으며, 구체적으로는 금은화와 상술된 다른 약재를 혼합하여 사용할 수 있다.On the other hand, the mixed medicine used in the present invention may include one or more of the above-mentioned medicines, and specifically, it may be used by mixing gold and silver coins and the above-mentioned other medicinal substances.
하나의 예로서, 상기 혼합 약재는 금은화, 해바라기 줄기 및 소리쟁이를 혼합하여 사용할 수 있고, 다른 예로서는 상술된 약재를 모두 혼합하여 사용할 수도 있다.As an example, the mixed medicinal material may be used by mixing gold and silver flowers, sunflower stems and sagebrush, and as another example, all of the above-mentioned medicinal materials may be mixed and used.
이 경우, 혼합 약재에 포함되는 각 약재들은 금은화 10 중량부에 대하여, 비단풀 8~12 중량부, 귤 과피 6~8 중량부, 해바라기 줄기 6~8 중량부, 병풀 6~6.5 중량부, 약모밀 4~6.5 중량부, 주목 잎 4~5 중량부, 소리쟁이 4~5 중량부, 감초근 1~3 중량부 및 자소엽 0.1~2중량부를 포함할 수 있다. 본 발명은 혼합 약재의 성분 및 함량을 상기 범위로 제어함으로써 조성물의 암 세포에 대한 사멸능을 보다 향상시킬 수 있다.In this case, each of the drugs included in the mixed medicine is based on 10 parts by weight of gold and silver coins, 8 to 12 parts by weight of silk grass, 6 to 8 parts by weight of tangerine peel, 6 to 8 parts by weight of sunflower stem, 6 to 6.5 parts by weight of centella asiatica, 4 parts by weight of ryegrass. -6.5 parts by weight, 4-5 parts by weight of yew leaves, 4-5 parts by weight of sagebrush, 1-3 parts by weight of licorice root, and 0.1-2 parts by weight of perilla leaf. The present invention can further improve the apoptosis ability of the composition against cancer cells by controlling the components and content of the mixed drug within the above range.
또한, 상기 생약 추출물은 발효된 것일 수 있다. 구체적으로, 상기 생약 추출물은 혼합 약재로부터 얻은 추출물을 바실러스계 균과 혼합하여 균을 배양하고, 이렇게 얻은 배양물에 멸균 및 당류를 첨가한 후 락토바실러스균으로 발효시킨 발효물일 수 있다.In addition, the herbal extract may be fermented. Specifically, the herbal extract may be a fermented product obtained by mixing an extract obtained from a mixed drug with Bacillus bacteria, culturing the bacteria, adding sterilization and sugar to the obtained culture, and then fermenting the extract with Lactobacillus bacteria.
본 발명에 따른 항암용 약학적 조성물은 발효된 생약 추출물을 포함하여 부패나 변성의 원인이 되는 잡균의 침범이나 증식을 효과적으로 억제시킬 수 있을 뿐만 아니라 인체 흡수율을 향상시켜 보다 높은 항암 효과를 구현할 수 있다.The anticancer pharmaceutical composition according to the present invention can effectively suppress the invasion or proliferation of various bacteria that cause decay or degeneration, including fermented herbal extracts, as well as improve the absorption rate of the human body to realize a higher anticancer effect. .
이와 더불어, 상기 항암용 약학적 조성물은 폐암, 대장암, 유방암, 자궁암, 간암, 위암, 담관암 및 신장암으로 이루어진 군으로부터 선택되는 1종 이상의 고형암 질환의 예방 및/또는 치료 용도로 사용될 수 있다.In addition, the anticancer pharmaceutical composition may be used for preventing and/or treating one or more solid cancer diseases selected from the group consisting of lung cancer, colon cancer, breast cancer, uterine cancer, liver cancer, stomach cancer, bile duct cancer and kidney cancer.
이때, 상기 항암용 약학적 조성물은 그 제형이 특별히 제한되는 것은 아니나, 바람직하게는 액상일 수 있다. 또한, 적합한 투여 경로는 경구, 정맥, 직장, 에어로졸, 비경구, 안구, 폐동맥, 점 막경유, 진피 내, 질, 귀, 비강 및 국소 투여를 포함할 수 있다. 구체적으로는 경구 투여 또는 국소 투여로 수행될 수 있다.In this case, the anticancer pharmaceutical composition is not particularly limited in its formulation, but may preferably be in a liquid form. In addition, suitable routes of administration may include oral, intravenous, rectal, aerosol, parenteral, ocular, pulmonary, transmucosal, intradermal, vaginal, otic, nasal and topical administration. Specifically, oral administration or topical administration may be performed.
하나의 예로서, 상기 비경구 전달 (delivery)은 근육 내, 피하, 정맥 내, 골수 내 주사뿐만 아니라 척수강 내, 직접 심실 내, 복강 내, 구강 내 및 비강 내 주사를 포함할 수 있다. 경우에 따라서, 전신계 보다는 국소 방식으로, 종종 데포제(depot preparation) 또는 서방성 배합물로, 예를 들어 기관으로의 직접적인 화합물의 주사를 통해 투여될 수 있다. 구체적으로, 길게 작용하는 배합물은 이식(예를 들어, 피하 또는 근육 내에 의해) 또는 근육 내 주사에 의해 투여될 수 있다.As an example, the parenteral delivery may include intramuscular, subcutaneous, intravenous, intramedullary injections, as well as intrathecal, direct intraventricular, intraperitoneal, intraoral and intranasal injections. Optionally, administration may be in a topical rather than systemic manner, often as a depot preparation or sustained release formulation, for example, via injection of the compound directly into the organ. Specifically, long-acting formulations may be administered by implantation (eg, subcutaneously or intramuscularly) or by intramuscular injection.
상기 항암용 약학적 조성물은 표적 약물 전달 시스템, 예를 들어 기간-특이적 항체로 코팅된 리포좀에 전달될 수 있다. 이 경우, 상기 리포좀은 선택적으로 기관에 의해 표적화되고 선택될 수 있다. 또한, 본 발명에 따른 약학적 조성물은 속방성(rapid release) 배합물의 형태로, 지방성(extended release) 배합물의 형태로, 또는 중간 방출 배합물의 형태로 제공될 수 있으며, 국소적으로 투여될 수도 있다.The anticancer pharmaceutical composition may be delivered to a target drug delivery system, for example, a liposome coated with a period-specific antibody. In this case, the liposome can optionally be targeted and selected by the organ. In addition, the pharmaceutical composition according to the present invention may be provided in the form of a rapid release formulation, in the form of an extended release formulation, or in the form of an intermediate release formulation, and may be administered topically. .
다른 예시로서, 본 발명의 항암용 약학적 조성물은 비경구적으로 또는 정맥으로 투여될 수 있으며, 이 경우 주사로 투여될 수 있다. 또 다른 예시로서, 상기 항암용 약학적 조성물은 경구로 투여될 수 있다.As another example, the anticancer pharmaceutical composition of the present invention may be administered parenterally or intravenously, in which case it may be administered by injection. As another example, the anticancer pharmaceutical composition may be administered orally.
한편, 본 발명의 항암용 약학적 조성물은 환자의 증상이 개선되지 않는 경우에 의사의 재량으로 환자의 질환 또는 증상의 징후를 개선하거나 다르게는 제어 또는 한정하도록 환자의 일생 동안을 비롯하여 장기간 동안 투여할 수도 있다. 환자의 상태가 개선되는 경우에는 의사의 재량으로 조성물의 투여를 특정한 기간(즉, "약물 휴지기") 동안 일시적으로 연속적으로 중지할 수 있다.On the other hand, the anticancer pharmaceutical composition of the present invention can be administered for a long period of time, including throughout the patient's life, to improve or otherwise control or limit the symptoms of the patient's disease or symptoms at the discretion of the doctor when the patient's symptoms are not improved. may be If the patient's condition improves, the administration of the composition may, at the discretion of the physician, be temporarily and continuously discontinued for a specified period of time (ie, a “drug resting period”).
개별적인 치료 요법과 관련된 변수들의 갯수가 많기 때문에, 전술한 범위는 단지 제안(suggestive)이며, 이러한 추천된 값으로부터의 상당한 이탈도 드문 것은 아니다. 이러한 투여량은 이로서 한정되는 것은 아니지만, 사용 된 화합물의 활성, 치료될 질환 또는 증상, 투여 모드, 개인의 요구사항들, 치료될 질환 또는 증상의 경중, 및 진료의의 판단을 비롯한 많은 변수에 따라 바뀔 수도 있다.Due to the large number of variables associated with individual treatment regimens, the ranges described above are only suggestive, and significant deviations from these recommended values are not uncommon. Such dosage will depend on many variables including, but not limited to, the activity of the compound used, the disease or condition to be treated, the mode of administration, individual requirements, the severity of the disease or condition to be treated, and the judgment of the practitioner. It may change.
이러한 치료 요법의 치료 효율 및 독성은 이로서 한정하는 것은 아니지만 LD50(개체수의 50%까지 죽음을 초래한 투여량) 및 ED50(개체수의 50%에서 치료 효과적인 투여량)를 측정하는 것을 비롯한, 세포 배양액 또는 실험 동물에서의 표준 약학 절차에 의해 측정될 수 있다. 독성 효과와 치료 효과 사이의 투여량 비율이 치료 지수 (therapeutic index)이며, 이는 LD50와 ED50 사이의 비율일 수 있다. 세포 배양 분석법 및 동물 연구로부터 수득된 결과가 인간에게 사용하기 위한 투여량의 범위를 구하는데 사용될 수 있다. 이러한 화합물의 투여량은 바람직하게는 최소 독성을 갖는 ED50를 포함하는 순환성 농도의 범위 내에 놓인다. 투여량은 사용된 투여 형태와 사용된 투여 경로에 따라 이 범위 내에서 변할 수 있다.The therapeutic efficacy and toxicity of such treatment regimens can be measured in cell culture or It can be measured by standard pharmaceutical procedures in laboratory animals. The dose ratio between the toxic and therapeutic effects is the therapeutic index, which may be the ratio between LD50 and ED50. Results obtained from cell culture assays and animal studies can be used to determine dosage ranges for use in humans. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with minimal toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration employed.
일반적으로, 본 발명에 기술된 작용 모드에 기초하여 병용 치료가 사용되는 예시에서, 본 발명에 기술된 항암용 약학적 조성물은 동일한 약학적 조성물로 투여되어야만 하는 것은 아니며, 경우에 따라서는 상이한 물리적 및 화학적 특성 때문에 상이한 경로로 투여될 수 있다. 또한, 초기 투여는 확립된 프로토콜을 따라 수행되며 그 다음 관찰되는 영향에 따라, 투여량, 투여 모드 및 투여 시간이 숙련자들에 의해 조절될 수 있다.In general, in the example in which combination therapy is used based on the mode of action described in the present invention, the anticancer pharmaceutical composition described in the present invention does not have to be administered in the same pharmaceutical composition, and in some cases, different physical and Due to their chemical nature, they may be administered by different routes. In addition, initial administration is carried out according to established protocols and then, depending on the effect observed, the dosage, mode of administration and time of administration can be adjusted by the skilled person.
아울러, 치료-효과 투여량은 약물이 병용 치료에 사용되는 경우에 변할 수 있다. 병용 치료는 추가로 환자의 임상 관리와 보조하도록 다양한 시점에서 시작하고 중지되는 주기적 치료를 포함할 수 있다. 본 발명에 기술된 병용 치료의 경우, 공동-투여되는 화합물의 투여량은 사용되는 공동-약물의 유형, 사용되는 구체적인 약물, 치료 중인 질환, 질병 또는 증상 등에 따라 변할 수 있다.In addition, the therapeutically-effective dosage may vary when the drugs are used in combination therapy. Combination therapy may further include periodic treatments that are started and stopped at various time points to assist with the clinical management of the patient. For the combination treatment described herein, the dosage of the co-administered compound may vary depending on the type of co-drug used, the specific drug used, the disease, disease or condition being treated, and the like.
이와 더불어, 경감될 증상(들)을 치료하거나 예방하거나 개선하는 투여 요법은, 다양한 인자들에 따라 개질되는 것으로 이해될 수 있다. 이러한 인자로는 대상이 앓는 질병뿐만 아니라 대상의 연령, 체중, 성별, 식이요법 및 의학적 증상 등을 포함한다. 따라서, 실제로 사용되는 투여 요법은 폭넓게 변하므로 본 발명에서 설명되는 투여 요법에서 벗어날 수 있다.In addition, it will be understood that the dosing regimen for treating, preventing or ameliorating the symptom(s) to be alleviated will be modified depending on various factors. Such factors include the subject's disease as well as the subject's age, weight, sex, diet, and medical conditions. Accordingly, the dosing regimens actually used vary widely and may deviate from the dosing regimens described in the present invention.
항암용 약학적 조성물의 제조방법Method for preparing a pharmaceutical composition for anticancer
또한, 본 발명은 일실시예에서, 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽으로 이루어진 군으로부터 선택되는 1종 이상을 포함하는 혼합 약재로부터 생약 추출물을 얻는 단계를 포함하는 항암용 약학적 조성물의 제조방법을 제공한다.In addition, in one embodiment, the present invention is from a mixed medicine containing at least one selected from the group consisting of gold and silver chrysanthemum, silk grass, tangerine rind, sunflower stem, centella asiatica, yammilk, yew leaf, sorijai, licorice root, and perilla leaf. It provides a method for preparing a pharmaceutical composition for anticancer comprising the step of obtaining a herbal extract.
본 발명에 따른 항암용 약학적 조성물의 제조방법은 혼합 약재로부터 생약 추출물을 얻는 단계를 포함한다. 이때, 상기 혼합 약재는 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽 중 적어도 1종을 포함할 수 있고, 경우에 따라서 상술된 약재를 모두 포함할 수 있다.The method for preparing a pharmaceutical composition for anticancer according to the present invention includes the step of obtaining a herbal extract from a mixed drug. In this case, the mixed medicinal material may include at least one of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yew wheat, yew leaf, sorijai, licorice root, and perilla leaf, in some cases including all of the above-mentioned medicinal substances can do.
또한, 상기 혼합 약재에 포함되는 각 약재들은 금은화 10 중량부에 대하여, 비단풀 8~12 중량부, 귤 과피 6~8 중량부, 해바라기 줄기 6~8 중량부, 병풀 6~6.5 중량부, 약모밀 4~6.5 중량부, 주목 잎 4~5 중량부, 소리쟁이 4~5 중량부, 감초근 1~3 중량부 및 자소엽 0.1~2중량부를 포함할 수 있다.In addition, each of the drugs included in the mixed medicine is based on 10 parts by weight of gold and silver coins, 8 to 12 parts by weight of silk grass, 6 to 8 parts by weight of tangerine peel, 6 to 8 parts by weight of sunflower stem, 6 to 6.5 parts by weight of centella asiatica, 4 parts by weight of ryegrass. -6.5 parts by weight, 4-5 parts by weight of yew leaves, 4-5 parts by weight of sagebrush, 1-3 parts by weight of licorice root, and 0.1-2 parts by weight of perilla leaf.
아울러, 생약 추출물을 추출하는 단계는 당업계에서 통상적으로 사용되는 방식으로 추출될 수 있다. 구체적으로, 상기 추출물은 여과법, 열수추출, 침지추출, 환류냉각추출 및 초음파추출 등의 방법을 통해 추출될 수 있으며, 바람직하게는 열수 추출을 이용하여 제조될 수 있다.In addition, extracting the herbal extract may be extracted in a manner commonly used in the art. Specifically, the extract may be extracted through methods such as filtration, hot water extraction, immersion extraction, reflux cooling extraction, and ultrasonic extraction, and preferably may be prepared using hot water extraction.
하나의 예로서, 상기 열수 추출은 세척된 혼합 약재를 적당한 크기로 분쇄하여 추출용기에 넣고, 추출용기에 추출 용매를 주입한 후 가열함으로써 수행될 수 있다.As an example, the hot water extraction may be performed by grinding the washed mixed medicinal material to an appropriate size, putting it in an extraction container, injecting an extraction solvent into the extraction container, and then heating.
여기서, 상기 추출용매는 물, 알코올 또는 이들의 혼합물을 사용할 수 있으며, 상기 알코올로는 탄소수 1 내지 4의 알코올을 이용하는 것이 바람직하다. 하나의 예로서, 상기 추출 용매는 에탄올 또는 메탄올을 이용할 수 있으며, 바람직하게는 70~80 wt% 에탄올을 이용할 수 있으나 이에 한정하지 않는다.Here, as the extraction solvent, water, alcohol, or a mixture thereof may be used, and as the alcohol, an alcohol having 1 to 4 carbon atoms is preferably used. As an example, the extraction solvent may use ethanol or methanol, preferably 70 to 80 wt% ethanol, but is not limited thereto.
또한, 상기 추출용매는 혼합 약재의 중량에 대하여 1 내지 10배 첨가할 수 있으며, 바람직하게는 1 내지 3배 첨가할 수 있다. 또한, 추출온도는 80 내지 110℃이고, 추출시간은 24 내지 48시간일 수 있으며, 추출횟수는 1회 내지 5회일 수 있다.In addition, the extraction solvent may be added 1 to 10 times, preferably 1 to 3 times, based on the weight of the mixed drug. In addition, the extraction temperature is 80 to 110 ℃, the extraction time may be 24 to 48 hours, the number of times of extraction may be 1 to 5 times.
이렇게 추출된 생약 추출물은 별도의 농축이나 건조 없이 여과하여 사용될 수 있으며, 발효 균주를 이용하여 상기 생약 추출물을 발효시킨 발효물로서 사용할 수도 있다.The herbal extract thus extracted may be used by filtration without separate concentration or drying, or may be used as a fermented product obtained by fermenting the herbal extract using a fermented strain.
구체적으로, 본 발명에 따른 항암용 약학적 조성물의 제조방법은 생약 추출물을 얻은 이후에 얻어진 생약 추출물을 발효시키는 단계를 더 포함할 수 있고, 상기 발효시키는 단계는 다음의 과정에 의해 수행될 수 있다:Specifically, the method for preparing the anticancer pharmaceutical composition according to the present invention may further include fermenting the herbal extract obtained after obtaining the herbal extract, and the fermenting step may be performed by the following process. :
생약 추출물과 바실러스 서브틸리스균을 혼합하고 배양하여 배양물을 제조하는 단계; 및Preparing a culture by mixing and culturing the herbal extract and Bacillus subtilis; and
제조된 배양물을 락토바실러스균으로 발효시키는 단계.Fermenting the prepared culture with Lactobacillus.
상기 배양물을 제조하는 단계는 생약 추출물과 바실러스계 균주, 구체적으로는 바실러스 서브틸리스 (Bacillus subtilis)를 균일하게 혼합하고, 이를 배양함으로써 수행될 수 있으며, 이때 생약 추출물과 바실러스계 균주의 배양 온도는 당업계에서 통상적으로 적용되는 범위 내에서 적절히 선택적으로 사용될 수 있으나, 구체적으로는 30~ 60℃, 바람직하게는 35~55℃의 온도에서 24~72시간 동안 배양함으로써 수행될 수 있다.The step of preparing the culture may be performed by uniformly mixing the herbal extract and the Bacillus strain, specifically, Bacillus subtilis , and culturing the same, wherein the culture temperature of the herbal extract and the Bacillus strain may be appropriately and selectively used within the range conventionally applied in the art, specifically, it may be carried out by culturing for 24 to 72 hours at a temperature of 30 to 60 ℃, preferably 35 to 55 ℃.
또한, 상기 발효시키는 단계는 바실러스계 균주와 함께 배양된 생약 추출물을 동결 건조시키고, 동결 건조물에 식물 원재료 멸균 및 탄소원 (전체 중량에 대햐여 0.025~0.06wt%)을 첨가한 후 락토바실러스균으로 발효시킴으로써 수행될 수 있다.In addition, in the fermenting step, the herbal extract cultured with the Bacillus strain is freeze-dried, sterilized plant raw materials and a carbon source (0.025 to 0.06 wt% based on the total weight) are added to the freeze-dried product, and then fermented with Lactobacillus. It can be done by
이때, 상기 발효 공정은 특별히 제한되지 않고 당업계에서 통상적으로 적용되는 조건 하에서 수행될 수 있으나, 구체적으로는 pH 5.0~7.5 및 20~70℃ 조건 하에서 24~72시간 동안 수행될 수 있고, 보다 구체적으로는 pH 6.0~7.0 및 25~60℃ 또는 30~50℃의 조건 하에서 24~72시간 동안 수행될 수 있다.At this time, the fermentation process is not particularly limited and may be carried out under conditions commonly applied in the art, specifically, it may be carried out for 24 to 72 hours under conditions of pH 5.0 to 7.5 and 20 to 70° C., more specifically As such, it may be carried out for 24 to 72 hours under conditions of pH 6.0 to 7.0 and 25 to 60 °C or 30 to 50 °C.
또한, 상기 발효 공정은 수행되는 동안 발효온도가 20~70℃ 범위 내에서 가변될 수 있다. 하나의 예로서, 상기 발효 공정은 48시간 동안 수행하되, 그 온도가 24시간 동안 20℃에서 60℃; 또는 30℃에서 40℃로 가변될 수 있다.In addition, during the fermentation process is carried out, the fermentation temperature may be varied within the range of 20 ~ 70 ℃. As an example, the fermentation process is performed for 48 hours, the temperature of which is 20° C. to 60° C. for 24 hours; or from 30°C to 40°C.
상기 방법에 의해 제조된 생약 추출물은 균주 체내에서 생합성되는 각종 효소, 즉, 미생물 균체 내에서는 세포 내 효소; 미생물 균체 외에서는 세포외 효소, 사멸된 세포에서 유리된 효소 등을 상당량 함유하게 할 수 있으며, 미생물에 의해 사용되고 남은 기존의 영양원과 미생물이 생육하면서 분비한 아미노산, 단백질, 비타민, 무기질, 불포화지방산, 당질, 식이섬유, 유기산 및 생체 유용한 생리활성 성분, 소화효소인 아밀라아제, 프로테아제, 리파아제 및 셀룰라아제 또는 이외의 효소 등이 다량 생성되어 생체 이용률이 우수한 특징을 나타낼 수 있다.The herbal extracts prepared by the method include various enzymes biosynthesized in the body of the strain, that is, intracellular enzymes in the microbial cells; Existing microorganisms can contain a significant amount of extracellular enzymes and enzymes released from dead cells, and amino acids, proteins, vitamins, minerals, unsaturated fatty acids, Carbohydrates, dietary fiber, organic acids and bio-available physiologically active ingredients, digestive enzymes amylase, protease, lipase, cellulase, or other enzymes are produced in large amounts, thereby exhibiting excellent bioavailability.
나아가, 본 발명에 따른 항암용 약학적 조성물의 제조방법은 발효된 생약 추출물을 숙성시키는 단계를 더 포함할 수 있으며, 상기 숙성은 20~40℃에 2주 이상 동안 수행될 수 있다.Furthermore, the method for preparing the anticancer pharmaceutical composition according to the present invention may further include the step of aging the fermented herbal extract, and the aging may be performed at 20-40° C. for 2 weeks or more.
이하, 본 발명을 제조예 및 실시예에 의해 상세히 설명한다.Hereinafter, the present invention will be described in detail by way of preparation examples and examples.
단, 하기 제조예 및 실시예는 본 발명을 예시하는 것일 뿐, 본 발명의 내용이 하기 제조예 및 실시예에 한정되는 것은 아니다.However, the following Preparation Examples and Examples only illustrate the present invention, the content of the present invention is not limited to the following Preparation Examples and Examples.
실시예 1 및 비교예 1~9.Example 1 and Comparative Examples 1 to 9.
혼합 약재로서, 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽을 각각 세척하고, 적당한 크기로 자른 다음 하기 표 1에 나타낸 함량 비율과 같이 칭량하여 총 중량이 7 kg이 되도록 추출 용기에 넣었다. 그 후, 추출 용기에 담긴 혼합 약재 전체 중량의 3배가 되는 멸균 정제수(21 L)를 붓고, 100±2℃로 가열하여 생약 추출물을 얻었다. 얻은 생약 추출물을 여과하여 항암용 약학적 조성물을 제조하였다.As a mixed medicine, each of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yew millet, yew leaf, sorijae, licorice root, and perilla leaf was washed, cut to an appropriate size, and weighed according to the content ratio shown in Table 1 below. It was placed in the extraction vessel so that the total weight was 7 kg. Thereafter, sterile purified water (21 L), which is three times the total weight of the mixed drug contained in the extraction container, was poured, and the mixture was heated to 100±2° C. to obtain a herbal extract. The obtained herbal extract was filtered to prepare an anticancer pharmaceutical composition.
단위:중량부Unit: parts by weight 실시예 1Example 1 비교예 1Comparative Example 1 비교예 2Comparative Example 2 비교예 3Comparative Example 3 비교예 4Comparative Example 4 비교예 5Comparative Example 5 비교예 6Comparative Example 6 비교예 7Comparative Example 7 비교예 8Comparative Example 8 비교예 9Comparative Example 9
금은화gold coin 1010 -- 1010 1010 88 1212 1010 1010 1010 1010
비단풀silk grass 99 99 99 99 99 99 99 99 99 99
귤 과피tangerine peel 77 77 77 77 77 77 77 77 77 77
해바라기 줄기sunflower stem 77 88 -- 88 77 88 55 1010 7.57.5 7.57.5
병풀Centella asiatica 6.56.5 6.56.5 6.56.5 6.56.5 6.56.5 6.56.5 6.56.5 6.56.5 6.56.5 6.56.5
약모밀weak wheat 5.55.5 5.55.5 5.55.5 5.55.5 5.55.5 5.55.5 5.55.5 5.55.5 5.55.5 5.55.5
주목 잎yew leaves 4.44.4 4.44.4 4.44.4 4.44.4 4.44.4 4.44.4 4.44.4 4.44.4 4.44.4 4.44.4
소리쟁이screamer 4.54.5 4.54.5 4.54.5 -- 4.54.5 4.54.5 4.54.5 4.54.5 22 77
감초근 licorice root 22 22 22 22 22 22 22 22 22 22
자소엽perilla leaf 1One 1One 1One 1One 1One 1One 1One 1One 1One 1One
실시예 2 및 비교예 10~18.Example 2 and Comparative Examples 10-18.
실시예 1과 비교예 1~9에서 제조된 각 생약 추출물을 여과하여 고형물을 제거하고, 바실러스 서브틸리스균을 0.2kg 접종한 다음 균일하게 혼합하여 배양기에서 주입하고, 배양온도 30±1℃ 및 습도 55±1%에서 24시간 동안 균주를 배양하였다.Each herbal extract prepared in Example 1 and Comparative Examples 1 to 9 was filtered to remove solids, and 0.2 kg of Bacillus subtilis was inoculated, mixed uniformly, and injected in an incubator, cultured at 30±1° C. and humidity. The strain was cultured at 55±1% for 24 hours.
배양이 종료되면 배양액에 식물 원재료 멸균과 탄소원 (전체 중량에 대햐여 0.04±0.05wt%)을 첨가하고, 락토바실러스균으로 24시간 동안 발효를 수행하였다. 이때, 발효 온도는 30±1℃에서 50±1℃로 가변시켰다. 발효가 완료되면 발효된 생약 추출물을 여과하여 항암용 약학적 조성물을 제조하였다.Upon completion of the culture, sterilization of plant raw materials and a carbon source (0.04±0.05 wt% based on the total weight) were added to the culture medium, and fermentation was performed with Lactobacillus bacteria for 24 hours. At this time, the fermentation temperature was varied from 30 ± 1 °C to 50 ± 1 °C. When the fermentation was completed, the fermented herbal extract was filtered to prepare an anticancer pharmaceutical composition.
실험예 1. 세포 독성 평가Experimental Example 1. Cytotoxicity evaluation
본 발명에 따른 항암용 약학적 조성물의 세포 독성을 평가하기 위하여, 실시예 1~2 및 비교예 1~18에서 얻은 조성물을 대상으로 MTT assay를 수행하였다.In order to evaluate the cytotoxicity of the anticancer pharmaceutical composition according to the present invention, MTT assay was performed on the compositions obtained in Examples 1 to 2 and Comparative Examples 1 to 18.
구체적으로, 2종의 인간 섬유 아세포인 ① 인체 피부 정상 세포주(CCD-986sk) 및 ② WI-38 세포를 각각 DMEM(Dulbecco's Modefied Eagle's Medium, Gibco-BRL) 배지에 10% FBS(Fetal Bovine Serum, Hyclone))와 1% 페니실린/스트렙토마이신(sigma 사)이 첨가된 배지를 사용하여 37℃, 5 % CO2 조건의 배양기(Forma)에서 배양하였다. 상기 인간 섬유 아세포들을 각각 96 웰 플레이트에 5×103 세포/웰의 밀도로 평판 배양한 후 시료를 처리하기 전 혈청이 제거된 DMEM 배지에서 1시간 동안 배양하였다. 앞서 준비된 실시예 1~2와 비교예 1~18의 조성물을 각각 0 vol.%, 5 vol.%, 10 vol.%, 15 vol.%, 20 vol.% 및 25 vol.% 농도 별로 첨가하여 18시간 동안 배양하였다. 18시간 동안 배양한 후 5mg/ml MTT(3-(4,5-다이메틸-2-티아졸릴)-2,5-다이페닐-2H-테트라졸륨 브로마이드) 용액을 웰 당 15 μl씩 넣어 4시간 배양 후 용해화 완충액 100% 다이메틸설폭사이드(DMSO)를 150 μl씩 첨가하였다. 마이크로플레이트 리더(sunrise, TECAN)를 이용하여 490 nm에서 흡광도를 측정하였다. 측정된 흡광도를 대조군의 흡광도와 비교하여 세포의 독성여부를 백분율로 산출하였으며, 그 결과는 도 1 및 도 2에 나타내었다.Specifically, two types of human fibroblasts, ① human skin normal cell line (CCD-986sk) and ② WI-38 cells, were respectively cultured in DMEM (Dulbecco's Modefied Eagle's Medium, Gibco-BRL) medium with 10% FBS (Fetal Bovine Serum, Hyclone). )) and 1% penicillin/streptomycin (Sigma) was added to the cultured in an incubator (Forma) at 37° C. and 5% CO 2 condition. The human fibroblasts were each plated in a 96-well plate at a density of 5×10 3 cells/well, and then cultured for 1 hour in DMEM medium from which serum was removed before sample treatment. By adding the compositions of Examples 1 to 2 and Comparative Examples 1 to 18 prepared above for each concentration of 0 vol.%, 5 vol.%, 10 vol.%, 15 vol.%, 20 vol.%, and 25 vol.%, respectively, Incubated for 18 hours. After incubation for 18 hours, 15 μl of a 5 mg/ml MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) solution was added per well for 4 hours. After incubation, 150 μl of a lysis buffer 100% dimethyl sulfoxide (DMSO) was added. Absorbance was measured at 490 nm using a microplate reader (sunrise, TECAN). By comparing the measured absorbance with the absorbance of the control group, the toxicity of the cells was calculated as a percentage, and the results are shown in FIGS. 1 and 2 .
실험 결과, 실시예 및 비교예의 조성물들은 대조군에 가까운 낮은 세포 독성을 가져 흡광도 변화율이 현저히 낮은 것으로 나타났다. 특히, 실시예 2의 조성물은 도 1 및 2에 나타낸 바와 같이 CCD-986sk 세포주에 대하여 12 vol.% 이하, WI-38 세포주에 대하여 17 vol.% 이하의 농도에서 세포독성이 거의 없는 것으로 확인되었다.As a result of the experiment, the compositions of Examples and Comparative Examples had low cytotoxicity close to that of the control group, indicating that the absorbance change rate was significantly low. In particular, it was confirmed that the composition of Example 2 had little cytotoxicity at a concentration of 12 vol.% or less for the CCD-986sk cell line and 17 vol.% or less for the WI-38 cell line, as shown in FIGS. 1 and 2 . .
이러한 결과로부터 본 발명에 따른 항암용 약학적 조성물은 생약 추출물을 유효성분으로 함유하여 세포 독성이 현저히 낮음을 알 수 있다.From these results, it can be seen that the anticancer pharmaceutical composition according to the present invention contains a herbal extract as an active ingredient, and thus has remarkably low cytotoxicity.
실험예 2. 암세포 사멸 평가Experimental Example 2. Evaluation of cancer cell death
또한, 본 발명에 따른 암세포 사멸 효과를 평가하기 위하여, 실시예 1~2 및 비교예 10~18에서 얻은 항암용 약학적 조성물을 대상으로 MTT assay를 수행하였다.In addition, in order to evaluate the cancer cell killing effect according to the present invention, the MTT assay was performed on the anticancer pharmaceutical compositions obtained in Examples 1 to 2 and Comparative Examples 10 to 18.
구체적으로, 먼저, 폐암 세포주(A549), 대장암 세포주(HCT116), 자궁암 세포주(HeLa), 담관암 세포주(HuCC-T1), 위암 세포주(AGS), 간암 세포주(HepG2), 유방암 세포주(MCF7) 및 신장암 세포주(HEK239)를 준비하고, 각 암 세포주를 100mm 디쉬로 37℃, 5% CO2 배양기에서 배양하였다. 각 세포주 배양을 위한 배지는 2㎎/㎖ 중탄산나트륨(sodium bicarbonte), 10% 우태혈청(Fetal Bovine Serum) 및 1% 페니실린-스트렙토마이신을 첨가한 DMEM(Dulbeco's Modified Eagle's Medium) 배지를 사용하여 배양하였다.Specifically, first, lung cancer cell line (A549), colon cancer cell line (HCT116), uterine cancer cell line (HeLa), cholangiocarcinoma cell line (HuCC-T1), gastric cancer cell line (AGS), liver cancer cell line (HepG2), breast cancer cell line (MCF7) and A renal cancer cell line (HEK239) was prepared, and each cancer cell line was cultured in a 100 mm dish at 37° C., 5% CO 2 in an incubator. The culture medium for each cell line was cultured using DMEM (Dulbeco's Modified Eagle's Medium) medium supplemented with 2 mg/ml sodium bicarbonate, 10% Fetal Bovine Serum, and 1% penicillin-streptomycin. .
배양된 각 암 세포주를 96웰 플레이트에 배양하고, 배양된 세포주에 실시예 1~2 및 비교예 10~18의 조성물들을 각각 1, 2, 5, 10, 15, 20 및 25 vol.%의 농도로 처리하여 37℃, 5%의 CO2의 배양기에서 24시간 동안 배양하였다. 배양 후 MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] 시약 20㎕를 처리하여 4시간 동안 배양한 후 효소면역측정기(ELISA reader) 490nm에서 흡광도를 측정하였으며, 대조군의 흡광도와 비교하여 암세포 사멸률을 백분율로 산출하였다. 산출된 결과 중 실시예 2 조성물의 처리 농도에 따른 암세포 사멸률은 표 2에 나타내었으며, 15 vol.% 농도에서의 실시예 및 비교예 조성물의 암세포 사멸률은 표 3 및 도 3 내지 도 5에 나타내었다.Each cultured cancer cell line was cultured in a 96-well plate, and the compositions of Examples 1 to 2 and Comparative Examples 10 to 18 were added to the cultured cell line at a concentration of 1, 2, 5, 10, 15, 20 and 25 vol.%, respectively. Treated with 37 ℃, 5% CO 2 Incubated for 24 hours in an incubator. After incubation, 20 μl of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] reagent was treated and incubated for 4 hours. Absorbance was measured at 490 nm with an ELISA reader. , The cancer cell death rate was calculated as a percentage compared to the absorbance of the control group. Among the calculated results, the cancer cell death rates according to the treatment concentration of the composition of Example 2 are shown in Table 2, and the cancer cell death rates of the compositions of Examples and Comparative Examples at a concentration of 15 vol.% are shown in Table 3 and FIGS. 3 to 5. indicated.
농도: vol.%Concentration: vol.% 폐암lung cancer 대장암colorectal cancer 자궁암uterine cancer 담관암cholangiocarcinoma 위암stomach cancer 간암liver cancer 유방암breast cancer 신장암kidney cancer
00 00 00 00 00 00 00 00 00
1One 13.9013.90 8.848.84 23.8223.82 -4.39-4.39 10.9810.98 -4.36-4.36 -17.69-17.69 -29.52-29.52
22 23.2923.29 11.4011.40 24.4624.46 1.721.72 4.434.43 -3.89-3.89 40.8840.88 -30.66-30.66
55 49.9949.99 40.3540.35 24.1124.11 59.9359.93 41.8541.85 75.2875.28 76.1476.14 0.800.80
1010 82.0082.00 78.2678.26 31.1131.11 88.6388.63 90.0490.04 75.3175.31 80.6980.69 62.7762.77
1515 78.1178.11 82.7682.76 73.5473.54 84.4884.48 90.7790.77 72.0172.01 80.3180.31 75.0875.08
2020 76.7576.75 82.8182.81 86.6986.69 82.9082.90 91.0591.05 72.0372.03 80.2980.29 68.5768.57
2525 79.0879.08 82.7882.78 84.0584.05 85.0385.03 90.3290.32 72.7172.71 79.9479.94 65.7365.73
농도: vol.%Concentration: vol.% 폐암lung cancer 대장암colorectal cancer 자궁암uterine cancer 담관암cholangiocarcinoma 위암stomach cancer 간암liver cancer 유방암breast cancer 신장암kidney cancer
실시예 1Example 1 76.69%76.69% 80.51%80.51% 72.00%72.00% 82.58%82.58% 88.11%88.11% 70.15%70.15% 78.63%78.63% 70.89%70.89%
실시예 2Example 2 78.11%78.11% 82.75%82.75% 73.54%73.54% 84.48%84.48% 90.77%90.77% 72.01%72.01% 80.31%80.31% 75.08%75.08%
비교예 10Comparative Example 10 72.44%72.44% 73.43%73.43% 68.25%68.25% 77.98%77.98% 80.52%80.52% 66.37%66.37% 73.55%73.55% 65.95%65.95%
비교예 11Comparative Example 11 75.92%75.92% 75.16%75.16% 70.84%70.84% 76.12%76.12% 81.86%81.86% 67.49%67.49% 72.96%72.96% 66.57%66.57%
비교예 12Comparative Example 12 73.51%73.51% 74.21%74.21% 69.37%69.37% 76.54%76.54% 81.08%81.08% 68.32%68.32% 74.21%74.21% 66.26%66.26%
비교예 13Comparative Example 13 74.19%74.19% 77.59%77.59% 68.62%68.62% 79.65%79.65% 84.40%84.40% 68.94%68.94% 77.21%77.21% 68.28%68.28%
비교예 14Comparative Example 14 75.46%75.46% 78.45%78.45% 71.11%71.11% 80.13%80.13% 85.16%85.16% 69.23%69.23% 76.84%76.84% 69.83%69.83%
비교예 15Comparative Example 15 73.81%73.81% 76.98%76.98% 70.26%70.26% 81.62%81.62% 84.72%84.72% 68.74%68.74% 75.76%75.76% 68.65%68.65%
비교예 16Comparative Example 16 75.21%75.21% 79.01%79.01% 69.33%69.33% 81.74%81.74% 86.89%86.89% 67.22%67.22% 76.32%76.32% 67.94%67.94%
비교예 17Comparative Example 17 74.87%74.87% 78.47%78.47% 71.76%71.76% 80.64%80.64% 87.31%87.31% 70.10%70.10% 74.99%74.99% 68.75%68.75%
비교예 18Comparative Example 18 74.54%74.54% 80.22%80.22% 69.79%69.79% 79.25%79.25% 85.84%85.84% 69.85%69.85% 76.83%76.83% 69.34%69.34%
상기 표 2 및 표 3과 도 3 내지 도 5에 나타낸 바와 같이 본 발명에 따른 항암용 약학적 조성물은 암 세포에 대한 사멸효과가 우수한 것을 알 수 있다.As shown in Tables 2 and 3 and FIGS. 3 to 5, it can be seen that the anticancer pharmaceutical composition according to the present invention has an excellent killing effect on cancer cells.
구체적으로, 상기 표 2를 참고하면, 실시예 2의 조성물은 정상세포에 대하여 독성을 나타내지 않는 10 vol.% 및 15 vol.%의 농도에서도 60% 이상의 암세포 사멸률을 나타내었으며, 특히 폐암, 담관암, 위암 및 유방암에 대해서는 80% 이상의 암세포 사멸률을 나타내는 것으로 확인되었다.Specifically, referring to Table 2, the composition of Example 2 exhibited a cancer cell death rate of 60% or more even at concentrations of 10 vol.% and 15 vol.% that do not show toxicity to normal cells, particularly lung cancer and cholangiocarcinoma. , it was confirmed that the cancer cell death rate of 80% or more for gastric cancer and breast cancer.
또한, 상기 표 3을 참고하면, 생약 추출물을 발효시킨 실시예 2의 조성물이 발효가 수행되지 않은 실시예 1의 조성물과 비교하여 암세포 사멸률이 높은 것으로 나타났으며, 비교예 10~18의 조성물과 비교하여도 15 vol.%의 낮은 농도에서 현저히 높은 암세포 사멸률을 갖는 것으로 확인되었다.In addition, referring to Table 3, it was found that the composition of Example 2 in which the herbal extract was fermented had a higher rate of cancer cell death compared to the composition of Example 1 in which fermentation was not performed, and the compositions of Comparative Examples 10 to 18 It was confirmed to have a significantly higher rate of cancer cell death even at a low concentration of 15 vol.% compared to .
따라서, 본 발명에 따른 항암용 약학적 조성물은 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽을 특정 함량으로 조합하여 추출물을 포함함으로써 세포 독성이 매우 낮을 뿐만 아니라 암 세포에 대한 사멸 효과가 현저히 우수하므로, 폐암, 대장암, 유방암, 자궁암, 간암, 위암, 담관암, 신장암 등의 고형암 예방 및/또는 치료용 물질로서 유용하게 사용될 수 있다.Therefore, the anticancer pharmaceutical composition according to the present invention is cytotoxic by including extracts of gold and silver lilies, silk grass, tangerine peels, sunflower stems, centella asiatica, yamba wheat, yew leaves, sorijai, licorice root and perilla leaf in a specific content. Since the apoptosis effect on cancer cells is very low as well as significantly excellent, it can be usefully used as a substance for preventing and/or treating solid cancers such as lung cancer, colon cancer, breast cancer, uterine cancer, liver cancer, stomach cancer, bile duct cancer, and kidney cancer.
본 발명에 따른 항암용 약학적 조성물은 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽 중 적어도 1종 이상을 포함하는 혼합 약재로부터 유래된 생약 추출물을 포함하여 인체에 투여 시 부작용이 적을 뿐만 아니라 암세포에 대한 사멸 효과가 우수하므로, 폐암, 대장암, 유방암, 자궁암, 간암, 위암, 담관암, 신장암 등의 고형암 질환의 예방 및/또는 치료에 유용하게 사용될 수 있다.The pharmaceutical composition for anticancer according to the present invention is a herbal extract derived from a mixed medicine containing at least one of gold and silver lilies, silk grass, tangerine peel, sunflower stem, centella asiatica, ryegrass, yew leaves, sorijai, licorice root and perilla leaf. When administered to the human body, including can be used

Claims (10)

  1. 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽으로 이루어진 군으로부터 선택되는 1종 이상의 약재를 포함하는 혼합 약재; 또는 상기 혼합약재의 생약 추출물을 유효성분으로 포함하는 항암용 약학적 조성물.A mixed drug comprising at least one medicinal material selected from the group consisting of gold and silver flowers, silk grass, tangerine peel, sunflower stem, centella asiatica, yammilk, yew leaves, sorijae, licorice root, and perilla leaf; Or a pharmaceutical composition for anticancer comprising the herbal extract of the mixed drug as an active ingredient.
  2. 제1항에 있어서,The method of claim 1,
    상기 혼합 약재는 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽을 포함하는 항암용 약학적 조성물.The mixed medicine is a pharmaceutical composition for anticancer comprising gold and silver flower, silk grass, tangerine peel, sunflower stem, centella asiatica, yew millet, yew leaf, sorijae, licorice root, and perilla leaf.
  3. 제1항에 있어서,The method of claim 1,
    상기 혼합 약재는 금은화 10 중량부에 대하여, 비단풀 8~12 중량부, 귤 과피 6~8 중량부, 해바라기 줄기 6~8 중량부, 병풀 6~6.5 중량부, 약모밀 4~6.5 중량부, 주목 잎 4~5 중량부, 소리쟁이 4~5 중량부, 감초근 1~3 중량부 및 자소엽 0.1~2중량부를 포함하는 항암용 약학적 조성물.The mixed medicine is based on 10 parts by weight of gold and silver coins, 8 to 12 parts by weight of silk grass, 6 to 8 parts by weight of tangerine peel, 6 to 8 parts by weight of sunflower stem, 6 to 6.5 parts by weight of Centella asiatica, 4 to 6.5 parts by weight of ryegrass, yew leaves A pharmaceutical composition for anticancer comprising 4-5 parts by weight, 4-5 parts by weight of sorijae, 1-3 parts by weight of licorice root, and 0.1-2 parts by weight of perilla leaf.
  4. 제1항에 있어서,The method of claim 1,
    상기 생약 추출물은 발효된 것을 특징으로 하는 항암용 약학적 조성물.The herbal extract is an anticancer pharmaceutical composition, characterized in that it is fermented.
  5. 제1항에 있어서,The method of claim 1,
    상기 조성물은 폐암, 대장암, 유방암, 자궁암, 간암, 위암, 담관암 및 신장암으로 이루어진 군으로부터 선택되는 1종 이상의 고형암에 적용되는 항암용 약학적 조성물.The composition is an anticancer pharmaceutical composition applied to one or more solid cancers selected from the group consisting of lung cancer, colon cancer, breast cancer, uterine cancer, liver cancer, stomach cancer, bile duct cancer and kidney cancer.
  6. 금은화, 비단풀, 귤 과피, 해바라기 줄기, 병풀, 약모밀, 주목 잎, 소리쟁이, 감초근 및 자소엽으로 이루어진 군으로부터 선택되는 1종 이상을 포함하는 혼합 약재로부터 물, 탄소수 1~4의 알코올 또는 이들의 혼합 용매를 이용하여 추출물을 얻는 단계를 포함하는 항암용 약학적 조성물의 제조방법.Water, alcohol having 1 to 4 carbon atoms or these A method for preparing an anticancer pharmaceutical composition comprising the step of obtaining an extract using a mixed solvent of
  7. 제6항에 있어서,7. The method of claim 6,
    상기 혼합 약재는 금은화 10 중량부에 대하여, 비단풀 8~12 중량부, 귤 과피 6~8 중량부, 해바라기 줄기 6~8 중량부, 병풀 6~6.5 중량부, 약모밀 4~6.5 중량부, 주목 잎 4~5 중량부, 소리쟁이 4~5 중량부, 감초근 1~3 중량부 및 자소엽 0.1~2중량부를 포함하는 항암용 약학적 조성물의 제조방법.The mixed medicine is based on 10 parts by weight of gold and silver coins, 8 to 12 parts by weight of silk grass, 6 to 8 parts by weight of tangerine peel, 6 to 8 parts by weight of sunflower stem, 6 to 6.5 parts by weight of Centella asiatica, 4 to 6.5 parts by weight of ryegrass, yew leaves 4 to 5 parts by weight, 4 to 5 parts by weight of sorijae, 1-3 parts by weight of licorice root, and 0.1-2 parts by weight of perilla leaf for the preparation of an anticancer pharmaceutical composition.
  8. 제6항에 있어서,7. The method of claim 6,
    상기 추출물을 얻는 단계 이후에, 얻어진 추출물을 발효시키는 단계를 더 포함하는 항암용 약학적 조성물의 제조방법.After obtaining the extract, the method for producing a pharmaceutical composition for anticancer further comprising the step of fermenting the obtained extract.
  9. 제7항에 있어서,8. The method of claim 7,
    상기 발효시키는 단계는,The fermenting step is
    추출물과 바실러스 서브틸리스균을 혼합하여 배양된 배양물을 제조하는 단계; 및Preparing a cultured culture by mixing the extract and Bacillus subtilis; and
    제조된 배양물을 락토바실러스균으로 발효시키는 단계를 포함하는 항암용 약학적 조성물의 제조방법.A method for producing a pharmaceutical composition for anticancer comprising the step of fermenting the prepared culture with Lactobacillus.
  10. 제7항에 있어서,8. The method of claim 7,
    상기 추출물을 발효시키는 단계 이후에, 발효된 추출물을 2주 이상 숙성시키는 단계를 더 포함하는 항암용 약학적 조성물의 제조방법.After the step of fermenting the extract, the method for producing a pharmaceutical composition for anticancer further comprising the step of aging the fermented extract for 2 weeks or more.
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