JPH07252141A - Antitumor agent - Google Patents

Abstract

PURPOSE:To obtain an antitumor agent comprising a pentacyclic triterpenoid as an active ingredient. CONSTITUTION:This antitumor agent comprises a compound of the formula (R<1> is H, an acyl, a lower alkyl, an aralkyl or carbamoyl; R<2> is carboxyl, cyano, hydroxyiminomethyl, a lower alkoxycarbonyl, etc.) or its salt as an active ingredient. The compound is properly mixed with a well-known carrier for a medicine and can be pharmaceutically manufactured into an oral agent such as tablet, capsule, granule, fine granule or powder or a parenteral preparation such as injection or suppository. A dose is usually 1-5,000mg/kg per time in the case of oral administration, usually 0.5-300mg/kg per time in the case of intravenous, intramuscular and hypodermic administration. The number of administrations is 1-3 times daily in the case of oral administration and once or twice daily in the case of injection.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to an antitumor agent containing a pentacyclic triterpenoid as an active ingredient.

[0002]

2. Description of the Related Art The following formula (I):

[0003]

[Chemical 2]

As a compound having a pentacyclic triterpenoid skeleton represented by, bryonolic acid
(In the formula (I), R ¹ = H, R ² = COOH), 3-acetylbrionolic acid (in the formula (I), R ¹ = Ac, R ² =
COOH), 3-succinylbrionolic acid (formula (I)
Where R ¹ = succinyl, R ² = COOH), methyl brionolate (in formula (I), R ¹ = H, R ² = COOCH
Various compounds such as ₃ ) are known (JP-A-2-73).
012 publication, JP-A-4-290846 publication, Chem.
Pharm. Bull., 40 , 1199-1202 (1992), Acta Cryst., S
ect. C., 43 , 1779 (1987)).

It is known that the compound having the pentacyclic triterpenoid skeleton represented by the above formula (I) has an anti-inflammatory action and an anti-allergic action (JP-A-2-73).
No. 012 and Japanese Patent Laid-Open No. 4-290846), it is not known to have an antitumor effect.

[0006]

SUMMARY OF THE INVENTION It is an object of the present invention to provide an antitumor agent containing a pentacyclic triterpenoid as an active ingredient.

[0007]

The present invention provides the following formula (I):

[0008]

[Chemical 3]

(Wherein R ¹ represents a hydrogen atom, an acyl group, a substituted or unsubstituted lower alkyl group, a substituted or unsubstituted aralkyl group or a substituted or unsubstituted carbamoyl group, and R ² represents a carboxyl group, cyano group Group, a hydroxyiminomethyl group, a lower alkoxycarbonyl group, a substituted or unsubstituted aralkyloxycarbonyl group, a substituted or unsubstituted lower alkyl group, or a substituted or unsubstituted carbamoyl group). It is an antitumor agent containing the salt as an active ingredient.

The acyl group represented by R ¹ is, for example, a lower acyl group having 1 to 6 carbon atoms such as formyl group, acetyl group, propionyl group, butyryl group, valeryl group, oxalyl group, malonyl group and succinyl group. An aromatic acyl group such as a benzoyl group, a p-dimethylaminobenzoyl group and a toluoyl group. The lower alkyl group represented by R ¹ is an alkyl group having 1 to 6 carbon atoms, such as a methyl group, an ethyl group, a propyl group, an isopropyl group,
A butyl group, an isobutyl group, a sec-butyl group, a tert-butyl group, a pentyl group, a hexyl group can be mentioned, and these lower alkyl groups can be substituted with an amino group, a dimethylamino group, a carboxyl group, a hydroxyl group or the like. Good. The substituted lower alkyl group, for example a carboxymethyl group, the formula: - (CH ₂₎ (. Wherein, n represents an integer of 1 to 5) _n -NH ₂ or _{- (CH 2) n -N (} CH ₃ ) ₂ (in the formula, n represents an integer of 1 to 5).

The aralkyl group represented by R ¹ is, for example, a phenyl group represented by the following formula:-(CH ₂ ) _n --C ₆ H ₅ (wherein n represents an integer of 1 to 5). Examples thereof include lower alkyl groups such as benzyl group and phenethyl group, and these aralkyl groups may be substituted with amino group, dimethylamino group, carboxyl group, hydroxyl group and the like. The substituted aralkyl group, for example, the following formula: - (CH ₂₎ (wherein, n represents an integer of _{1~5.) N -C 6 H 4} -NH 2 aminophenyl represented by - lower alkyl group Can be mentioned.

Examples of the substituted carbamoyl group represented by R ¹ include dimethylcarbamoyl group, diethylcarbamoyl group, methylcarbamoyl group, phenylcarbamoyl group and hydroxycarbamoyl group. The lower alkoxycarbonyl group represented by R ² has 1 to 6 carbon atoms.
Of alkoxycarbonyl groups, and examples thereof include a methoxycarbonyl group, an ethoxycarbonyl group, a propoxycarbonyl group, an isopropoxycarbonyl group, and a butoxycarbonyl group.

The aralkyloxycarbonyl group represented by R ² is, for example, the following formula: —COO— (CH ₂ ) _n —C ₆ H ₅ (wherein n represents an integer of 1 to 5). Examples of the phenyl-lower alkoxycarbonyl group shown include benzyloxycarbonyl group and phenethyloxycarbonyl group, and these aralkyloxycarbonyl groups are substituted with an amino group, a di-lower alkylamino group, a carboxyl group, a hydroxyl group or the like. May be. The substituted aralkyloxycarbonyl group, for example, the following _{formula: -COO- (CH 2) n -C} 6 H 4 -NR 2 ( wherein, n represents an integer of 1 to 5, R is a hydrogen atom or a carbon atoms The group represented by 1) to 6 is represented.

The lower alkyl group represented by R ² is an alkyl group having 1 to 6 carbon atoms, for example, methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group, sec-butyl group, tert-butyl group, pentyl group, hexyl group, and these lower alkyl groups are substituted with an amino group, di-lower alkylamino group, carboxyl group, hydroxyl group, carboxymethoxy group, lower alkoxycarbonylmethoxy group, etc. May be. Examples of the substituted lower alkyl group include a carboxymethyl group, the following formula: —CH ₂ NR ₂ (wherein R represents a hydrogen atom or an alkyl group having 1 to 6 carbon atoms), or —CH ₂ OCH ₂ COOR (formula). In the above, R represents a hydrogen atom or an alkyl group having 1 to 6 carbon atoms).

Examples of the substituted carbamoyl group represented by R ² include a dimethylcarbamoyl group, a diethylcarbamoyl group, a methylcarbamoyl group, a phenylcarbamoyl group, a hydroxycarbamoyl group, and the following formula: —CONHCH ₂ COOR (wherein R is hydrogen. represents an atom or an alkyl group having 1 to 6 carbon atoms.) or _{-CONH- (CH 2) n -NR 2} ( wherein, n represents an integer of 1 to 5, R is 1-6 hydrogen or carbon atoms Represents an alkyl group of).

Specific examples of the pentacyclic triterpenoid represented by the above formula (I) include, but are not limited to, those shown below. Brionolic acid (3β-hydroxy-D: C-freedoolean-8-ene-29-oic acid) (in formula (I),
R ¹ = H, R ² = COOH), 3-acetylbrionolic acid (3β-acetoxy-D: C-freedoolean-
8-en-29-oic acid) (in the formula (I), R
¹ = Ac, R ² = COOH), 3-succinylbrionolic acid (3β-succinyloxy-D: C-freedoolean-8-ene-29-oic acid) (in formula (I), R ¹ = Succinyl, R ² = COOH), methyl brionoleate (in formula (I), R ¹ = H, R ² = CO
OCH _3), D: C- Freed me Anne-8-en -3
β, 29-diol (in the formula (I), R ¹ = H, R ² = C
H ₂ OH).

The pentacyclic triterpenoid represented by the above formula (I) is a known compound and can be obtained by a known method. For example, bryonolic acid is Bryonia di
oica (Cucurbitaceae), Luffa cylindrica (Cucurbitace
ae), Trichosanthes kirilowii Maxim. (Cucurbitacea
e), Sandoricum indicum (Meliaceae) and other plants can be obtained by extraction and isolation (Gazz. Chim.
Ital., 99 , 830 (1969), Annu. Rev. Plant Physiol.,
26 , 209 (1975), Phytochemistry., 11 , 3341 (1972)
Etc.). It can also be obtained by extracting and isolating from a cultured tissue produced by organ culture of the plant (see Japanese Patent Laid-Open No. 2-107194, etc.).

By treating bryonolic acid with diazomethane, methyl brionolic acid can be obtained (see Production Example 1 of JP-A-4-290846). The compound in which R ² in the formula (I) is a hydroxymethyl group or a formyl group can be obtained by reducing brionolic acid or methyl brionolate with a suitable catalyst (Production Example of JP-A-4-290846). Refer to 1).

In the above formula (I), R¹Is an acyl group
Compounds are R such as bryonolic acid ¹Is a hydrogen atom
The compound was treated with acetic anhydride in the presence of a base catalyst such as pyridine,
By reacting with carboxylic acid anhydride such as succinic anhydride
(Japanese Patent Laid-Open No. 4-290846)
Refer to Production Example 3). Carboxyl group such as brionolic acid
The compound having
It is converted to a pharmaceutically acceptable salt, such as
It can be used as a minute.

Further, a compound having an amino group, a dimethylamino group or the like can be converted into a pharmaceutically acceptable acid addition salt such as hydrochloride by a conventional method and used as an active ingredient. It is known that the compound represented by the formula (I) and a salt thereof have high safety (JP-A-2-730).
(See Experimental Example 4 of Japanese Patent No. 12 and Experimental Example 2 of Japanese Patent Laid-Open No. 4-290846). For example, it has been reported that the LD ₅₀ of bryonolic acid in mice is 3000 mg / kg or more by oral administration (Experimental Example 4 of JP-A-2-73012).

Next, the formulation and dose of the antitumor agent of the present invention will be described. The antitumor agent of the present invention can be formulated by combining the compound represented by the above formula (I) or a salt thereof with a known pharmaceutical carrier. The dosage form is not particularly limited and may be appropriately selected and used as needed, and examples thereof include oral agents such as tablets, capsules, granules, fine granules and powders, parenteral agents such as injections and suppositories. To be

The dose of the compound represented by the above formula (I) or a salt thereof, which is an active ingredient, depends on the patient's age, body weight,
Depending on the severity of the disease, usually 1 time per oral administration
˜5000 mg / kg body weight, and usually 0.5 to 300 mg / kg body weight once by intravenous, intramuscular or subcutaneous administration. Oral administration is usually 1-3 times daily.
For injections, 1 to 2 times a day.

Oral agents include, for example, starch, lactose, sucrose,
Mannitol, carboxymethyl cellulose, corn starch, inorganic salts and the like are used in a conventional manner. In addition to the above-mentioned excipients, a binder, a disintegrant, a surfactant, a lubricant, a fluidity promoter, a flavoring agent, a coloring agent, a fragrance and the like can be appropriately used in this type of formulation. Specific examples of each are as shown below.

[Binder] Crystalline cellulose, crystalline cellulose / carmellose sodium, methylcellulose, hydroxypropylcellulose, low-substituted hydroxypropylcellulose, hydroxypropylmethylcellulose 2
208, hydroxypropyl methylcellulose 290
6, hydroxypropylmethylcellulose 2910, hydroxypropylmethylcellulose phthalate 2007
31, hydroxypropyl methyl cellulose phthalate 220824, hydroxypropyl methyl cellulose acetate succinate, carmellose sodium, ethyl cellulose, carboxymethyl ethyl cellulose, hydroxyethyl cellulose, wheat starch, rice starch, corn starch, potato starch, dextrin, pregelatinized starch, partially pregelatinized starch , Hydroxypropyl starch, pullulan, polyvinylpyrrolidone K25, polyvinylpyrrolidone K3
0, aminoalkyl methacrylate copolymer E, aminoalkyl methacrylate copolymer RS, methacrylic acid copolymer L, methacrylic acid copolymer S,
Methacrylic acid copolymer LD, polyvinyl acetal diethylaminoacetate, polyvinyl alcohol, gum arabic, gum arabic powder, agar, gelatin, white shellac, tragacanth, purified white sugar, macrogol 200,
Macrogol 300, Macrogol 6000.

[Disintegrant] Crystalline cellulose, methyl cellulose, low-substituted hydroxypropyl cellulose, carmellose, carmellose calcium, carmellose sodium, croscarmellose sodium, wheat starch, rice starch, corn starch, potato starch, partially pregelatinized starch , Hydroxypropyl starch, sodium carboxymethyl starch,
Tragant.

[Surfactant] Soybean lecithin, sucrose fatty acid ester, polyoxyl 40 stearate, polyoxyethylene hydrogenated castor oil 100, polyoxyethylene hydrogenated castor oil 40, polyoxyethylene hydrogenated castor oil 5
0, polyoxyethylene hydrogenated castor oil 60, polyoxyethylene [42] polyoxypropylene [67] glycol, polyoxyethylene [54] polyoxypropylene [39] glycol, polyoxyethylene [105]
Polyoxypropylene [5] glycol, polyoxyethylene [160] polyoxypropylene [80] glycol, polyoxyethylene [196] polyoxypropylene [67] glycol, sorbitan sesquioleate, sorbitan trioleate, sorbitan monostearate. , Sorbitan monopalmitate, sorbitan monolaurate, polysorbate 40, polysorbate 6
0, polysorbate 65, polysorbate 80, glyceryl monostearate, sodium lauryl sulfate, lauromacrogol.

[Lubricant] Wheat starch, rice starch, corn starch, stearic acid, calcium stearate, magnesium stearate, hydrous silicon dioxide, light anhydrous silicic acid, synthetic aluminum silicate, dried aluminum hydroxide gel, talc, metasilicic acid. Acid magnesium aluminate, calcium hydrogen phosphate, anhydrous calcium hydrogen phosphate, sucrose fatty acid ester, waxes, hydrogenated vegetable oil, polyethylene glycol.

[Flowability Accelerator] Hydrous silicon dioxide, light anhydrous silicic acid, dried aluminum hydroxide gel, synthetic aluminum silicate, magnesium silicate. The antitumor agent of the present invention can also be administered as a suspension, emulsion, syrup, or elixir, and these various dosage forms may contain a flavoring agent and a coloring agent.

On the other hand, parenteral preparations are manufactured by a conventional method,
Generally, distilled water for injection, physiological saline, aqueous glucose solution, vegetable oil for injection, sesame oil, peanut oil, soybean oil, corn oil, propylene glycol, polyethylene glycol and the like can be used as the diluent. Further, if necessary, a bactericidal agent, a preservative, and a stabilizer may be added. Further, from the viewpoint of stability, this parenteral preparation may be filled in a vial or the like and then frozen, the water content may be removed by a usual freeze-drying technique, and a liquid preparation may be re-prepared from the freeze-dried product immediately before use. Further, if necessary, a tonicity agent, a stabilizer, a preservative, a soothing agent and the like may be added appropriately.

[0030]

EXAMPLES The present invention will be described in more detail with reference to production examples and examples, but the scope of the present invention is not limited thereto. (Production Example 1) Trichosanthes kirilowii M
axim. var. japonica Kitamura) 11.9 g of hairy roots were pulverized and subjected to reflux extraction with chloroform at 60 to 70 ° C. The obtained extract was filtered and the solvent was distilled off to obtain 372 mg of an extract. This was separated and purified by silica gel column chromatography, and 160 g of bryonolic acid was added.
mg was obtained (yield: 1.34% with respect to the starting material).

The obtained bryonolic acid is colorless needle crystals (melting point 299-302 ° C.), ¹ H-NMR and ¹³ C-NM.
It was found from R that it has 30 carbon atoms, 7 tertiary methyl groups, 4 substituted double bonds, 1 secondary hydroxyl group and 1 carboxyl group, and various physicochemical properties are shown in literature values of bryonolic acid ( Pharmaceutical journal, 109 , 250 (1989)). (Example 1) (1) MTT method (J. Immunol. Methods, 65 , 55 (1983))
The antitumor activity was measured by.

B16 cells (mouse melanoma) were cultured in a medium (10% fetal bovine serum, 50 U / ml penicillin, 50).
1.8 × 10 ⁴ cells / in non-essential amino acid-containing Earl's solution (tissue culture medium) containing μg / ml streptomycin
It was diluted to a concentration of ml and seeded in a 96-well microplate at 0.1 ml / well. 24 hours culture (37
° C., after 5% CO ₂ containing air), the medium was removed, each drug final concentration 1μg / ml, 5μg / ml or 25 [mu] g /
The solution was suspended in 1% Tween 20 aqueous solution to give a volume of 0.1 ml, and 0.1 ml / well was added thereto. After culturing for 48 hours, MTT (3- (4,5-dimethylthiazole-2-
Yl) -2,5-diphenyltetrazolium bromide)
(SIGMA) was dissolved in a phosphate buffer to a concentration of 5 mg / ml, and 10 μl / well was added. After culturing for 4 hours, 0.1 ml of 10% sodium dodecyl sulfate aqueous solution
/ Well was added to dissolve the crystals of formazan produced in the living cells due to intracellular dehydrogenase, and after further culturing for 24 hours, the absorbance was measured at 577 nm and 655 nm. For the determination, the ratio of color development in the drug-added group to the drug-unadded group was produced, and the viable cell rate was determined.

The results are shown below. Drug-free group 100 Brionolic acid 1 μg / ml addition group 87.8 Brionolic acid 5 μg / ml addition group 62.1
(P <0.01) Brionolic acid 25 μg / ml addition group 57.4
(P <0.01) Brionolic acid 5 μg / ml addition group and bryonolic acid
A remarkable growth inhibitory effect was observed in the 25 μg / ml addition group.

(2) The cell killing effect of bryonolic acid was measured by B16
When examined by the same MTT test as that for cells, a strong cell killing effect was observed on mouse leukemia cells L1210 cells (IC ₅₀ : 0.024 μg / ml). Example 2 30 g of brionolic acid was mixed with 155 g of microcrystalline cellulose and 5 g of magnesium stearate,
This mixture was tableted with a single-shot tableting machine to produce tablets having a diameter of 7 mm and a weight of 190 mg. One tablet of this tablet contains 30 mg of bryonolic acid.

(Example 3) 10 g of bryonolic acid was mixed with 490 g of corn starch, water was added, and the mixture was kneaded.
Granules were dried on a screen having a mesh of 1 mm x 1 mm and dried to obtain granules. 20 mg of bryonolic acid is contained in 1 g of this granule. (Example 4) Brionolic acid (20 g) was mixed with lactose (70 g) and magnesium stearate (10 g) and filled into hard capsules (500 mg each). One capsule of this capsule contains 100 mg of bryonolic acid.

(Example 5) 10 g of brionolic acid was added to 8 parts of water.
It was dissolved in 0 ml and 2 ml of orange essence and a single syrup were added to obtain a total amount of 1000 ml of a syrup. 1 mg of this syrup contains 10 mg of bryonolic acid. (Example 6) According to a conventional method for producing an injectable solution, 5 g of bryonolic acid was dissolved in 1 liter of distilled water for injection, made isotonic by adding sodium chloride, and dispensed in 10 ml aliquots into vials. 1 mg of this injection contains 5 mg of bryonolic acid.

Example 7 30 g of 3-acetylbrionolic acid was mixed with 155 g of microcrystalline cellulose and 5 g of magnesium stearate, and the mixture was tabletted with a single-shot tableting machine to give tablets having a diameter of 7 mm and a weight of 190 mg. Was manufactured. One tablet of this tablet contains 30 mg of bryonolic acid.

Example 8 30 g of 3-succinylbrionolic acid was mixed with 155 g of microcrystalline cellulose and 5 g of magnesium stearate, and this mixture was tabletted with a single-shot tableting machine to give tablets having a diameter of 7 mm and a weight of 190 mg. Was manufactured. One tablet of this tablet contains 30 mg of bryonolic acid.

Example 9 30 g of methyl brionolate
Was mixed with 155 g of microcrystalline cellulose and 5 g of magnesium stearate, and this mixture was tabletted with a single-shot tableting machine to produce tablets having a diameter of 7 mm and a weight of 190 mg. One tablet of this tablet contains 30 mg of bryonolic acid.

(Example 10) D: C-freed olean-
30 g of 8-ene-3β, 29-diol was mixed with 155 g of microcrystalline cellulose and 5 g of magnesium stearate, and this mixture was tabletted with a single-shot tableting machine to obtain a diameter of 7 m.
m, weight 190 mg tablets were produced. One tablet of this tablet contains 30 mg of bryonolic acid.

[0041]

INDUSTRIAL APPLICABILITY According to the present invention, an antitumor agent containing a pentacyclic triterpenoid as an active ingredient can be provided.

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI Technical indication location A61K 31/245 9454-4C 31/27 9454-4C C07C 35/44 9155-4H 43/18 C 7419 −4H 69/013 A 9279-4H E 9279-4H 271/32 9451-4H (72) Inventor Chiba Bando 3586 Yoshihara, Ami-cho, Inashiki-gun, Ibaraki Prefecture Tsumura Co., Ltd.

Claims (1)

[Claims] 1. The following formula (I): (In the formula, R ¹ represents a hydrogen atom, an acyl group, a substituted or unsubstituted lower alkyl group, a substituted or unsubstituted aralkyl group or a substituted or unsubstituted carbamoyl group, and R 1
² represents a carboxyl group, a cyano group, a hydroxyiminomethyl group, a lower alkoxycarbonyl group, a substituted or unsubstituted aralkyloxycarbonyl group, a substituted or unsubstituted lower alkyl group, or a substituted or unsubstituted carbamoyl group. ) An antitumor agent containing a pentacyclic triterpenoid or a salt thereof as an active ingredient.