KR20060060921A - Pharmaceutical composition comprising old platycodon extracts as an effective components for inhibiting 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone(nnk) inducing lung cancer - Google Patents

Abstract

The present invention prevents and inhibits Jangsaeng Bellflower Extract (CK) against lung cancer caused by 4- (methylnitrosamino) -1- (3-pyridyl) -1-butanone (NNK), a well known carcinogen in tobacco smoke. (Inhibition) and therapeutic use.

The present invention inhibits the expression of Proliferating cell nuclear antigen (PCNA), which is an indicator of tumor cell proliferation in lungs, increased by NNK, a lung cancer-causing substance contained in tobacco smoke, and NNK, a lung cancer-causing substance. It has been confirmed that it has an effect of suppressing the generation of lung tumors caused by, and the Jangsaeng Doraji Extract (CK) is used as an active ingredient to prevent, suppress and treat lung cancer caused by tobacco. The use of Jangsaeng Bellflower Extract

Jangsaeng Bellflower, 4- (methylnitrosamino) -1- (3-pyridyl) -1-butanone (NNK), lung cancer

Description

Pharmaceutical composition comprising old platycodon extracts as an effective components for inhibiting 4- (methylnitrosamino) -1- (3-pyridyl) -1-butanone (NNK) inducing lung cancer}             

1 is a diagram showing the amount of PCNA protein expression measured by Western blot method according to the administration of Jangsaeng Bellflower Extract (CK).

Figure 2 is a lung photograph of the experimental animal according to the administration of Jangsaeng Bellflower Extract (CK).

Figure 3 is a graph showing the results of measuring the number of tumors generated outside the alveolar according to the administration of Jangsaeng Bellflower Extract (CK).

Figure 4 is a photograph of lung tissue stained by Hematoxylin-Eosin (H.E) staining according to the administration of Jangsaeng Bellflower Extract (CK).

The present invention relates to the use of Jangsaeng Doraji Extract (CK), and more particularly 4- (methylnitrosamino) -1- (3-pyridyl) -1-butanone (NNK), which is well known as a carcinogen contained in tobacco smoke. It relates to the prevention, inhibition (suppression) and therapeutic use of Jangsaeng Bellflower Extract (CK) against lung cancer caused by.

Bellflower (Gilyeong Platycodon grandiflorum A. DC) is a plant of Campanulaceae and has a root length of about 10 to 15 cm, diameter of 1 to 3 cm, irregular stem spots on the upper part, gray to milky white, deep vertical wrinkles, and horizontally. There are sows and wrinkles. The vagina is hard but non-fibrous, so it is easy to fracture, smells a little, and tastes good. (Yook Chang-soo et al., Modern Herbal Medicine, Seoul, Hakchangsa, 460-461, 1993)

The main pharmacological component of these bellflower is triterpene-based saponins (platycodin A, C, D, D ₂ , D ₃ ), accounting for about 3% of the roots. Inhibitory effect (sedative, analgesic, antipyretic effect), anti-inflammatory action against acute and chronic inflammation, anti-ulcer and gastric juice secretion, anticholinergic action to lower blood pressure by expanding blood vessels, lowering blood sugar, improving cholesterol metabolism ( Chem. Pharm. Bull ., 20 , 1952 (1972), Chem. Pharm. Bull ., 23 , 2965 (1975), J. Chem. Soc. , Perkin trans I, 661 (1984), Sogoigaku , 3 , 1 (1951), J. Pharm. Soc.Kor., 19 , 164 (1975). In addition, steroidal compounds such as alpha-spinasterol, stigmas-7-enol, alpha-spinasterol glucoside, and polysaccharides such as inulin and betulin were also isolated as pharmacological components of bellflower.

Jangsaeng Bellflower (長 桔) refers to the bellflower for more than 20 years, and despite the various pharmacological actions of the bellflower, it has been difficult to cultivate it for a long time. (Lee Sung-ho, "Perennial Bellflower Cultivation Act" Korean Patent No. 045731) Mass production was possible. The Jangsaeng Bellflower has excellent pharmacological and physiological activity compared to the common bellflower of 2-4 years.

To date, studies on Jangsaeng Doraji have been conducted on antidiabetic effects (J. Korean Soc. Food Sci. Nutr., 25, 986 (1996)), hyperlipidemia (J. Pharmaceutical Society of Japan 93 (1973) 1188 -1194), and studies on the prevention and protection of liver damage continue.

Lung cancer often occurs between the ages of 40 and 70 and rarely occurs before the age of 40. The incidence ratio of men and women has increased since the 1980s compared to 7: 1 in the 1960s. This is due to the increase in female smoking rate. Although the cause is unclear, many epidemiological and experimental studies indicate that occupational exposures such as smoking, asbestos, radon, chromium, nickel, aromatic hydrocarbons, arsenic compounds, and radiation are risk factors. Smoking is the biggest factor among them.

Among carcinogens found in tobacco smoke, 4- (methylnitrosamino) -1- (3-pyridyl) -1-butanone (NNK) is most specifically known to cause lung cancer in various experimental animals (MKK Shivji Cell., 69, 367-374 (1992). NNK is also the most potent carcinogenic nitrosamine found in cigarettes. These facts indicate that NNK is the leading cause of lung cancer in smokers.

Proliferating cell nuclear antigen (PCNA), whose expression is increased by NNK, is an indicator of the proliferation of tumor cells during carcinogenesis and is involved in the replication of DNA as a cofactor of DNA polymerase (H. Kohno et al. Cancer Letters 174., 141-150 (2001). Intracellular PCNAs are increased for DNA repair to repair damaged DNA and for proliferation of cancer cells at various stages of carcinogenesis.

The present inventors have repeatedly studied to find an agent for treating and preventing pulmonary disease that is not toxic while effectively preventing, suppressing and restoring lung cancer caused by NNK, a lung cancer-causing substance contained in tobacco smoke, Jangsaeng Doraji (長 桔) extract was found to prevent and inhibit the effects of NNK-induced lung cancer has completed the present invention.

It is an object of the present invention to provide a pharmaceutical composition comprising an extract of Jangsaeng Doraji which is a safe and nontoxic natural product having a prophylactic and inhibitory effect on lung cancer occurrence caused by smoking or inhalation of tobacco smoke.

The present invention is characterized in that Jangsaeng Doraji extract has a use for preventing, inhibiting and treating lung cancer caused by tobacco. That is, the present invention is characterized in that the Jangsaeng Doraji extract can be effectively used for protecting the lungs by effectively preventing, suppressing and restoring lung cancer caused by NNK, a lung cancer-causing substance contained in tobacco smoke.

Jangsaeng bellflower extract used in the present invention is prepared according to methods commonly used in the art. Jangsaeng Doraji extracts may include conventional hot water extracts or organic solvent extracts, and these extracts may be prepared alone or by adding an acceptable herbal medicine, and may be prepared including a pharmaceutically acceptable carrier.

Generally, Jangsaeng Bellflower powder is used to dry Jangsaeng Bellflower so as to have a moisture content of 5% or less, pulverize it, and then select a particle size of 0.6 mm or smaller. For hot water extract, add about 5 to 10 times of water to Jangsaeng Bellflower powder and use the filtrate obtained by extracting twice for 4 to 6 hours at a temperature of 90 to 100 ° C, or selecting an acceptable herbal or herbal extract. Add to and use. For organic solvent extract, about 3 times the organic solvent is added to Jangsaeng Bellflower Powder, and the filtrate obtained by extracting twice at room temperature is used after concentration under reduced pressure. Ethanol may be preferably used as the organic solvent.

Jangsaeng Doraji extract of the present invention can be prepared in a pharmaceutical formulation according to a conventional method. In the preparation of the formulation, it is preferred to mix or dilute the extract of Jangsaeng Doraji as an active ingredient with the carrier, or to enclose it in a carrier in the form of a container. When the carrier is used as a diluent, it may be a solid, semisolid or liquid substance which acts as a vehicle, excipient or medium for the active ingredient. Thus, the formulations may be in the form of tablets, pills, powders, assays, elixirs, suspensions, emulsions, solutions, syrups, aerosols, soft or hard gelatin capsules, sterile injectables, sterile powders and the like. Examples of suitable carriers, excipients and diluents include lactose, dextrose, sucrose, sorbitol, mannitol, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, Propylhydroxybenzoate, talc, magnesium stearate and mineral oil. The formulation may further comprise fillers, anti-coagulants, lubricants, wetting agents, fragrances, emulsifiers, preservatives and the like. Pharmaceutical compositions according to the invention may be formulated using methods well known in the art to provide rapid, sustained or delayed release of the active ingredient after administration to a mammal.

The pharmaceutical compositions according to the invention can be administered via several routes including oral, transdermal, subcutaneous, intravenous or intramuscular. For humans, a typical daily dosage may range from 10 to 1,000 mg / kg body weight, preferably 10 to 100 mg / kg body weight, and may be administered once or in several doses. However, the actual dosage should be determined in light of several relevant factors such as the route of administration, the age, sex and weight of the patient, the state of health and the severity of the disease.

Hereinafter, the present invention will be described in more detail with reference to Examples. However, these Examples are only for illustrating the present invention, and the scope of the present invention is not limited thereto.

Example 1 Preparation of Jangsaeng Bellflower Hot Water Extract

Jangsaeng bellflower was dried and pulverized to 5% by weight or less of water content, and then sorted to have a particle size of 0.6 mm or less to prepare a powder. 2 L of distilled water was added to 0.2 kg of Jangsaeng Bellflower powder, extracted twice at 90-95 ° C. for 5 hours, filtered and lyophilized to obtain 100g of Jangsaeng Bellflower Extract (CK).

Experimental Example 1 Experimental Animal and Experimental Method

1. Reagent

The following reagent was purchased and used.

Antibodies against PCNA were purchased from Cell signaling, Tris-base and Bovine serum albumin (BSA) were purchased from USB, and Tween-20 and N, N, N ', N'-Tetramethyl ethylenediamine (TEMED) were obtained from Sigma. PVDF membrane was purchased from Pall corporation, Xylene was purchased from Duksan, Methyl alcohol and Ethyl alcohol were purchased from Hayman, and NNK was purchased from Toronto research chemicals.

2. Animal

The experimental animals purchased 20 ~ 25g of A / J mice 6 weeks old from Samtaco and used them freely for feed (purina korea) and water. The temperature of the kennel was 21 ~ 24 ℃, relative Humidity was maintained at 40-80%. In addition, the light in the kennel was adjusted to repeat day and night every 12 hours.

3. LNK-induced Lung Cancer and Jangsaeng Doraji Extract (CK) Treatment

Normal control group treated with 15 animals per animal (control), group injected with NNK (NNK 100 mg / kg), group administered with Jangsaeng Doraji extract (CK) after the injection of NNK (NNK 3 mg / kg + CK 10 mg / kg, NNK 3mg / kg + CK 100 mg / kg). NNK was intraperitoneally injected at 100 mg / kg at the start of the experiment and intraperitoneally injected at 100 mg / kg after 1 week. Jangsaeng Bellflower Extract (CK) was orally administered 10 mg / kg / day and 100 mg / kg / day three times a week for 23 weeks.

4. Statistical Analysis

For statistical significance test, one-way ANOVA with t-test method was used, and it was judged that the case where P value was less than 0.05 was significant.

[Example 2] Changes in body weight and lung weight by lung cancer-causing substances NNK and Jangsaeng Bellflower Extract (CK)

To investigate the changes in body weight and lung weight of experimental animals caused by lung cancer-causing substances NNK and Jangsaeng Doraji Extract (CK), the weights of NNK and Jangsaeng Doraji Extract (CK) were measured before The body weight was measured again 23 weeks after the treatment. In addition, the weight of the lung was measured to investigate the change in lung weight caused by NNK and Jangsaeng Doraji Extract (CK).

Table 1 shows the comparison between the initial weight before NNK treatment and the weight after 23 weeks of NNK treatment in the group injected with lung cancer-causing substance NNK and the group treated with Jangsaeng Doraji extract (CK) after NNK injection. It is a vote.

Table 1 Changes in weight of mice used for experiment

Group Initial body weight (g) Final bady weight (g) Weight gain (g) Control 22.3 ± 2.5 38.5 ± 1.3 14.3 ± 2.3 NNK 3mg / kg 21.2 ± 3.1 31.8 ± 2.5 10.4 ± 2.2 NNK + CKT 10mg / kg 20.3 ± 1.2 35.2 ± 1.3 14.3 ± 1.3 NNK + CKT 100mg / kg 22.1 ± 1.1 37.3 ± 2.5 15.3 ± 2.5

As shown in Table 1, when compared with the weight before treatment with NNK and Jangsaeng Doraji extract (CK) and the weight after 23 weeks of treatment, the weight of the group injected with lung cancer-causing substance NNK only decreased slightly. After the NNK injection, the group fed the Jangsaeng Doraji Extract (CK) recovered the reduced weight of the NNK-only group and showed no change compared to the normal group.

Example 3 Inhibitory Effect of Jangsaeng Doraji Extract (CK) on PCNA Expression, an Indicative of Increased Cancer Cell Proliferation by NNK

Used as an experimental animal to investigate the effect of CK extract on the expression of proliferating cell nuclear antigen (PCNA) involved in DNA damage caused by NNK, a known carcinogen, and DNA replication for cancer cell proliferation. 30 days after injection of NNK into the mice and administration of Jangsaeng Doraji Extract (CK), a portion of the lung was extracted, buffered, homogenized with a Homogeniger, and the supernatant was collected using a centrifuge. The expression level of PCNA protein was measured by the method of blot.

1 is a diagram showing the amount of expression of PCNA protein, from which the protein expression amount of PCNA, which is an increased cell proliferation index increased by NNK, a well known carcinogen, is significantly reduced by treatment of Jangsaeng Doraji Extract (CK). You can see that.

Example 4 Inhibitory Effects of Jang Saeng Doraji Extracts (CK) on Lung Tissue by LNK-causing Substance NNK

In the lung tissues of the experimental animals, the number of tumors formed in the lungs of the lung cancer-causing substance NNK injection group and the group treated with NNK injection and then treated with Jang Saeng Doraji extract (CK) was measured, and HE (Hematoxylin-) in lung tissue was compared. Inhibition of lung tumors by CK was compared by comparing the lungs of normal group treated with Eosin), NNK-abdominal group, and group treated with Jangsaeng Bellflower Extract (CK). .

2 is a lung photograph of a laboratory animal according to the administration of Jangsaeng Bellflower Extract (CK), from which it can be seen that lung tumors caused by NNK are inhibited by Jangsaeng Bellflower Extract (CK). 3 is a graph showing the results of measuring the number of lung tumors, from which it can be seen that Jangsaeng Doraji extract (CK) has an effect of inhibiting the production of lung tumors. Figure 4 is a photograph of the result of staining the lung tissue with Hematoxylin-Eosin (H.E), it can be seen that the lung tumor induced by NNK is inhibited by the administration of Jangsaeng Doraji extract (CK).

As described above, the composition according to the present invention comprising Jangsaeng Doraji extract exhibits a prophylactic and inhibitory effect on lung tumor formation caused by NNK, a potent lung cancer-causing substance contained in tobacco smoke. It will be useful for the prevention and treatment of lung cancer.

Claims (1)

Composition for preventing, inhibiting and treating lung cancer due to tobacco comprising Jangsaeng Bellflower Extract as an active ingredient.

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