KR0155977B1 - Anticancer composition - Google Patents

Abstract

The present invention relates to a novel use of taurousodeoxycholine acid (T-UDCA) as a pharmaceutical ingredient for cancer treatment, and to a pharmaceutical composition comprising the same. A pharmaceutical composition is prepared by containing one or two or more additives.

As a carrier, one or two or more kinds of diluents, lubricants, binders, disintegrants, sweeteners, stabilizers, and preservatives are selected and used. The additive is one or two or more selected from fragrances and vitamins. It is formulated in the form of a capsule, a soft capsule, a liquid, a powder.

The daily dose based on T-UDCA is preferably 5 mg-1 g, particularly 10-600 mg.

Description

Anticancer composition

1 is a graph of the anticancer effect of T-UDCA by the nude mouse model.

The present invention relates to the anticancer use of tauroursodeoxycholic acid (T-UDCA) of formula (I) and a pharmaceutical composition comprising the same.

In general, anticancer drugs not only act selectively on cancer cells but also damage normal cells, especially tissue cells with active cell division, resulting in side effects such as bone marrow dysfunction, gastric mucosa, and hair loss. That is, since the general anticancer drugs have the disadvantage that the side effect increases as the effect is excellent, the development of anticancer drugs without side effects is urgently emerging.

Taurorusodeoxycholine acid of the above structural formula (I) is a known compound. Looking at the presently reported pharmacologic area and its use example, taurolusodeoxycholine acid has a gallstone dissolving action, and urosodeoxycholine acid Because of its active form, it has been reported to have faster drug expression than urusodeoxycholic acid, as well as high absorption rate in the body, high fat digestion capacity, stable at a wide pH, and no formation of precipitate in the intestine. Taurolite as a gallstone dissolving agent whose main component is leusodeoxycholine acid is Italy) is commercially available.

In addition, taurorusodeoxycholic acid has been reported to be effective in treating fatty liver. That is, when orally administered taurusodeoxycholine acid has been reported to improve liver function and decrease in liver fat mass (Japanese Patent Publication No. 平 4-235918).

However, there are no reports of T-UDCA's anticancer effects.

Accordingly, the present inventors have newly researched that T-UDCA has little side effects and excellent effects as an anticancer agent, and has completed the present invention.

An object of the present invention is to provide a novel use of T-UDCA as an anticancer agent and a novel pharmaceutical composition for treating cancer containing the same as a main ingredient.

Hereinafter, the present invention will be described in detail.

The pharmaceutical composition for treating cancer of the present invention contains T-UDCA as an active ingredient with little side effects and excellent effects.

The T-UDCA of the present invention can be prepared and used in conventional pharmaceutical formulations containing one or two or more pharmaceutically acceptable conventional carriers and additives.

The carrier may be used by selecting one or two or more from diluents, lubricants, binders, disintegrants, sweeteners, stabilizers and preservatives, and may be used by selecting one or two or more from fragrances and vitamins. It may be prepared orally by tablets, capsules, soft capsules, liquids, powders, and the like.

In the present invention, the carrier and the additive may be used in any pharmaceutically acceptable, but if it is listed in more detail, diluents such as lactose monohydrate, corn starch, soybean oil, microcrystalline cellulose ( microcrystalline cellulose) or mannitol (D-mannitol) is preferable, and magnesium stearate or talc is preferable as a lubricant, and polyvinylpyrrolidone (PVP: polyvinylpyrrolidone) or hydroxypropyl cellulose (PVP) is preferable. It is preferable to select from HPC: hydorxypropylcellulose. In addition, the disintegrant may be selected from among carboxymethylcellulose calcium (Ca-CMC: carboxymethylcellulose calcium), sodium starch glycolate, potassium acrylamide (polacrylin kalium) or cross-linked polyvinylpyrrolidone. The sweetener is selected from white sugar, fructose, sorbitol or aspartame, and the stabilizer is carboxymethylcellulose sodium (Na-CMC), beta-cyclodextrin, or white lead. (white bee's wax) or xanthan gum, and preservatives include methyl p-hydroxy benzoate (methylparaben), propyl p-hydroxybenzoate (propyl paraben) or potassium sorbate ( potassium sorbate).

As the additive of the present invention, the fragrance may be echivanilline, masking flavor, fruit or herb, and the vitamins may be selected from vitamin B groups such as vitamin B ₁ and B ₂ .

Formulation method of the present invention can be carried out by conventional methods known in the art, it is possible to formulate in the formulation of tablets, capsules, soft capsules, solutions, powders and the like.

T-UDCA is currently used as a gallstone dissolving agent and is recognized to have little side effects, and LD ₅₀ is a very safe drug of 3g / kg or more.

In the present invention, T-UDCA is a tablet, capsule, soft so that the total amount of T-UDCA is 5mg ~ 1g, preferably 10-600mg per day in adults 60kg body weight in order to obtain a medical anti-cancer effect It is preferable to administer a capsule, a liquid, or a powder arbitrarily several times.

Hereinafter, the present invention will be described in detail through experimental examples and examples, and the present invention is not limited to the following experimental examples and examples.

Experimental Example 1

In-vitro anticancer effect of T-UDCA

Experiment method

In general, MTT (3- [4,5-Dimethylthiazol-2-yl] -2,5-diphenyltetrazolium bromide: Thiazolyl blue), the most commonly used in-vitro anticancer drug screening method, inhibits the cancer cell proliferation of T-UDCA. For the eight human cancer cell lines, 10,000 μg / ml, 1,000 μg / ml, 100 μg / ml, 10 μg / ml, 1 μg / ml and 0.1 μg / ml on 96-well culture plate (CORNIG) were identified. T-UDCA was added at the concentration. Experimental cells were incubated for 50 days in a 50 μl cell suspension at 37 ℃, 5% CO ₂ incubator at the indicated concentration. 2.5 µg / ml was prepared with PBS (-), and 10 µl of MTT solution was added to each well, followed by incubation for 3 hours at 37 ° C and 5% CO ₂ . Thereafter, 100 μl of 0.01 N HCl solution containing 10% SDS was added to each well, and the absorbance was measured at 580 nm after overnight culture at 37 ° C. and 5% CO ₂ .

Evaluation method of growth inhibition rate

Proliferation inhibition rate was evaluated as follows.

Growth inhibition = = absorbance of 1-drug addition group / absorbance of solvent-added group) × 100

In addition, the IC ₅₀ value of each study cell was 100% of the cell proliferation of the drug addition group (T-UDCA or 5-FU addition group), and when taurourusodeoxycholine acid at a concentration of 0.1 to 10,000 µg / ml was added. The growth inhibition rate of was regressed.

Experiment result

T-UDCA showed a proliferation inhibitory effect of 50% at a concentration of about 60-500 µg / ml in all cell lines (Table 1). In other words, T-UDCA was found to have a proliferation inhibitory effect on cancer cells.

Experimental Example 2

In-vivo anticancer effect of T-UDCA

Laboratory animals

For the WiDr and LoVo transplantation of human tumor cells, 6-week-old male BALB / Cr SIc mice purchased from Japan Clear kk were used. In addition, 6-week-old male nude mice, such as WiDr, were used for transplantation of LoVo cell lines. Feed and water were sterilized and fed freely during breeding.

Transplant method

LoVo cell lines were cultured at 5% CO and 37 ° C by adding Ham'S F-2 medium (containing 10% FBS) in a 100 ml plastic culture flask containing 150 cm 2. The cells were washed with PBS (-) solution, detached from the flask with EDTA solution containing 0.05% trypsin, and prepared by floating the cells by mixing Hanks' solution and PBS (-) solution in a 1: 1 ratio. Cell suspension is 2 × 10 per head under 5 nude mice Cells (0.2 ml) were transplanted. Tumors were removed from the tumor-forming mice to remove the coating and necrotic sections, and then streptomycin and penicillin were added, washed in Han's solution, and evenly cut to 2-3 cm in size. Six nude mice per group are implanted using a trocar under the back of the abdomen. In the same way, WiDr cell lines were also cultured with Eagle MEM medium and transplanted into nude mice.

Medication Administration

Taururusodeoxycholine acid was diluted in physiological saline for injection to prepare 10, 40 µg / ml and orally administered 0.1 ml per 10 g of mouse. Floracil (5-FU) was diluted in physiological saline for injection, prepared at 4 µg / ml, and administered intraperitoneally at 0.1 ml per 10 g of mouse body weight. In the control group, physiological saline was administered intraperitoneally with 0.1 ml per 10 g of mouse body weight. T-UDCA was orally administered 3-4 weeks once a week after separating the groups by the volume of the mouse tumor. Floracil (5-FU) was administered intraperitoneally twice a week for 3-4 weeks.

How to measure

Using a vernier caliper, measure the maximum diameter (L) and the thickness (W) at right angles to it, where V = 1/2 × L × W Experiments were performed by dividing the group into six groups of 1 group at the time when the estimated tumor volume reached about 100 mm 3 by substituting the equation. At the end of the experiment, tumor diameter was measured twice a week until 3-4 weeks after the start of administration.

result

Taurorusodeoxycholine acid showed superior efficacy in both the LoVo and WiDr cell lines compared to the control of the Flora LASIK (5-FU) ® control (see FIG. 1). In the LoVo cell line, fluorouracil (5-FU) showed 40.8% inhibition rate compared to the control group, but tauurosodeoxycholine acid showed 69.1% and 41.1% inhibition rates in the 100 mg / kg and 400 mg / kg groups, respectively. In the WiDr cell line, fluorouracil (5-FU) showed an inhibition rate of 15.6% compared to the control group, but tauurosodeoxycholine acid showed an inhibition rate of 32.8% in the 100 mg / kg administration group and a 37.9% inhibition in the 400 mg / kg administration group. Indicated. From these results, it can be seen that tauurorusodeoxycholic acid in human cancer cell lines, LoVo and WiDr cell lines, has superior efficacy compared to the control drug fluorouracil (5-FU).

Cancer drug medicines based on taurorusodeoxycholic acid according to the present invention may be formulated according to the following examples.

Example 1

Example 2

[Product made in capsule]

After mixing the main ingredient and the excipient of the above formulation, the weight of one capsule is prepared by a conventional capsule production method by 207 mg each.

Example 3

[Soft capsule]

After mixing the main ingredient and the excipient of the above formulation, the weight of one capsule is prepared by a conventional soft capsule preparation method by 314.3 mg.

Example 4

[Liquid]

Beta-cyclonedextrin of the above-mentioned formulation is dissolved in a predetermined amount of purified water, followed by dissolving by adding a main ingredient and an excipient, and then preparing the solution by a liquid preparation method in which the final amount is 10L by adding purified water.

Example 5

[Powder]

After dissolving the beta-cyclonextrine of the formulation in a predetermined amount of purified water and adding the main ingredient and excipients, it is dried and prepared by the conventional powder preparation method by weight of 1,000 mg each.

Experimental Example 3

[Safety test]

1) Gum 쳬: Five formulations of tablets, capsules, soft capsules, solutions and powders prepared according to Examples 1 to 5 above.

2) Storage condition: Safety test for temperature and humidity

Packing Type

① Tablet, capsule: PTP packaging and cartoon box packaging

② Liquid: Put in brown glass container and sealed with aluminum lid

③ Soft capsule: PTP packaging

Storage temperature: 40 ± 1 ℃, 75% RH (± 5%)

[However, soft capsule is 35 ± 1 ℃, 70% RH (± 5%)]

Storage period: 6 months

3) Test period: 6 months

4) Measurement time: start of test and every 2 months

5) Test item: Content test for main ingredient

6) Test Method: Taururusodeoxycholic acid was quantified by HPLC as follows.

※ Determination of Taurorusodeoxycholic Acid

a) Column: Nova-Pak C ₁₈

b) Mobile phase: 0.005MK ₂ HPO ₄ buffer (pH 5): MeOH = 25: 75 (v / v%)

c) Folw rate: 1.0 ml / min

d) Detection: UV 210nm

e) Range: 0.2AUFS

f) Injection volume: 10µl

7) Test Results: Table 2 below.

8) Discussion: It is useful as anticancer agent because it is stable for 6 months in accelerated safety test with storage temperature of 40 ℃.

Experimental Example 4

[Acute Toxicity Test]

1) Test substance: Taurusodeoxycholine acid

Test animals: Male rats 6 weeks old and male mice 5 weeks old (10 animals / group)

3) Preparation of test substance: The drug was completely dissolved in sterile distilled water, and prepared before administration on the test day.

4) Test Method: After fasting the test animals for 6 hours, the drug was administered orally into the stomach of the test animals using a mouse and rat bulge. The dose was administered based on the weight of the body immediately before administration, and the death of the agent was observed for 14 days.

5) Test Results: As a result of acute toxicity test in rats and mice, the LD of the pharmaceutical composition was higher than 3g / kg in rats and mice.

As described above, the pharmaceutical composition for treating cancer according to the present invention can be seen that the anticancer action is not only superior to fluorouracil, but also very stable in terms of formulation, and thus the present invention has proved to be an industrially useful invention.

Claims (6)

A pharmaceutical composition for the treatment of cancer, characterized by containing taurorusodeoxycholine acid (T-UDCA) and one or two or more pharmaceutically acceptable carriers and additives. The pharmaceutical composition according to claim 2, wherein the carrier is selected from diluents, lubricants, binders, disintegrants, sweeteners, stabilizers and preservatives. The pharmaceutical composition according to claim 2, wherein the additive is selected from fragrances or vitamin B groups. The pharmaceutical composition according to claim 2, wherein the daily oral dose of taurousodeoxycholine acid (T-UDCA) is 5 mg / g. The pharmaceutical composition according to claim 6, wherein the daily oral dose of taurousodeoxycholine acid (T-UDCA) is 10-600 mg. The pharmaceutical composition according to claim 2, wherein the formulation is selected from tablets, capsules, soft capsules, solutions, and powders.