JPH07103020B2 - Anti-cancer drug - Google Patents

Description

TECHNICAL FIELD The present invention relates to an anticancer agent containing a 1-oxo-tetrahydroanthracene compound.

1-oxo-containing as a main component in the anticancer agent of the present invention
The following compounds are known among the tetrahydroanthracene compounds (hereinafter abbreviated as the compound of the present invention).

Germichrysone [Michio Takido et al., Phytochemistry, Volume 15, 1295
P. 1976], Torosachrysone Michio Takito et al., Lloydia, 40, 191, 1977.
Year], 7-methyltorosachon
rysone) [Michio Takito, etc., Kita Nakashin, etc., Chemical Pharmaceutical Bulletin (Chem.Pharm.Bull.), Volume 33,
971, p. 1985], Fleg Machine B ₂ (phlegmacin B ₂ ) Michio Takito, Susumu Kitachu, etc., Fight Chemistry, Volume 16, 999
Page, 1977], Trosanin-9,10-quinone (torosanin
-9,10-quinone), Torosanin (torosanin), anhydrosorbitan Hulagu Khan machine B ₂ (anhydrophlegmacin B ₂₎ [Takito Michio, proceeds like North and Central Fight Chemistry, Vol. 21, 2103
Page, 1982], Trosachrysone B
[Michio Takito, Susumu Kitachu, etc., Proceedings of the 28th Annual Meeting of the Japanese Society of Biopharmacy (Tokyo), page 24, 1981], Tolosakrison D
(Trosachrysone D) [Michio Takito, Susumu Kitanaka, etc., 32nd Annual Meeting of the Japanese Society of Biopharmacy (Okayama), 27, 1985],
Octidentalin A (occidentalin A), Octidentalin B (new name) (occidentalin B) [Michio Takito, Susumu Kitachu, Nihon University College of Pharmaceutical Sciences, 24, 81, 19
1984], Germitorosone, Methylgermitorosone, [Michio Takito, Susumu Kitachu, etc., Fight Chemistry, 21, 425, 1
982], singuesnol-I
[JP-A-56-55334] and the like.

The above compound is a legume (Cassia torosa Cav.)
, And a compound that can be isolated from Cassia occidentalis L., the Chinese name for these hubs and mosquitoes is called Wangjiangan, and the seeds are hypertensive headache, eye congestion and tumor pain, chronic constipation, and abdominal pain due to diarrhea. It is used for etc. In addition, Singeanor-I in the compound of the present invention was prepared by Endo et al.
Ingueana), the compound whose isolated structure was determined,
It is described that -55334 has an antibacterial action and a smooth muscle relaxing action.

However, the anticancer agent of the compound of the present invention is not known.

Problems to be Solved by the Invention Most conventional anticancer agents have the property of inhibiting nucleic acid synthesis of cells, and since inhibition occurs in both normal cells and tumor cells, anticancer agents that do not act on normal cells Development is desired.

Means for Solving the Problems The present inventors are studying isolated structure determination of various 1-oxo-tetrahydroanthracene compounds contained in Cassius torosa Cav. And Cassia occidentalis L. The inventors have found that the compound of the present invention has antitumor activity and completed the present invention.

That is, the present invention [Wherein, R ¹ represents a hydrogen atom or a hydroxyl group, R ^2, R ⁵ are the same or different and each represents a hydrogen atom or a methyl group, R ^3, R ⁷ are both hydrogen atoms or hydrogen either R ^3, R ⁷ is Atom and the other R ⁴ represents a methoxy group or a methyl group, and R ⁶ represents a hydrogen atom or an acetyl group. ] It is an anticancer agent containing the 1-oxo- tetrahydro anthracene compound shown by these.

Examples of the compounds of the present invention are shown in Table 1 below.

In the table, A is B is C is D is E is Indicates.

These compounds are extracted from roots, flowers, seeds, seedlings, etc. of Habuso halibut or Rhododendron with benzene, chloroform, ether, ethyl acetate, etc., and the extract is subjected to adsorption column chromatography, preparative thin layer chromatography (PLC) or Countercurrent partition chromatography (DC
It can be obtained by isolation and purification using C.) and the like, and then recrystallization using various solvents. Further, the compound thus obtained can be obtained by acetylating acetic anhydride-pyridine in a conventional manner to obtain a compound of R ⁶ = Ac.

Hereinafter, the isolation and purification processes of the compound of the present invention obtained from the psyllium and the compound of the present invention will be described in detail with reference to Production Examples.

Production Example All melting points are uncorrected values measured by Yanagimoto's trace melting point measuring machine. Ultraviolet absorption spectrum (UV) is Hitachi 200-10spr
Infrared absorption spectrum (IR) is JASC using a ctorometer
An OIR A-2 spectrometer was used. JEOL FX-100 was used for the nuclear magnetic resonance spectrum (NMR), and TMS was used as the internal standard. Mass spectrum (MS) is Hitachi RMU-7Mspectrometer
Was used.

Production Example 1 Separation from fresh root of P. perniciosus 3.2 kg of fresh root of P. persicae was chopped into benzene 7
After extraction with benzene for 8 hours at room temperature, the mixture is filtered, and the extraction residue is further extracted with benzene 7 for 8 hours at room temperature. 2 above
The extracts were combined twice and the solvent was distilled off under reduced pressure to obtain 18.2 g of a benzene extract. 18.0 g of this extract was subjected to column chromatography (diameter 8 cm, length 24 cm) using 500 g of Marincklot (registered trademark, Mallinckrodt) and eluted with benzene to obtain chrysophanol 16
mg, followed by elution with benzene-acetic acid ethyl ester (9: 1) to give 28 mg of emodin and 58 mg of compound 1 (germicrison). Next, 2.5 g of each fraction eluted with benzene-acetic acid ethyl ester (4: 1) was subjected to fractionation again by using Marincklot column chromatography. This n-hexane-acetic acid ethyl ester (4: 1) elution fraction was further treated with 0.5N oxalic acid and subjected to preparative thin-layer chromatography (PLC) to duplicate twice with benzene-acetic acid ethyl ester (9: 1). After development, the yellow band was scraped off, extracted with acetic acid ethyl ester, the solution was washed 3 times with water, and then concentrated under reduced pressure to give crude crystals which were recrystallized from benzene to give compound 10 20 mg of (octydentaline A) and 10 mg of compound 11 (octidentaline B) were obtained.

Compound 10 (octydentaline A) Yellowish brown needles mp 280 ° C (decomposition) High-resolution MS (calculated as m / z C ₃₃ H ₃₂ O ₁₀ ); 588.1993 observed; 588.1994 MS m / z: 558,570,552,284,283,270,269 Compound 11 (octidentalline B) Yellowish brown prism mp 270 ° C (decomposition) High-resolution MS (calculated as m / z C ₃₂ H ₃₀ O ₁₀ ); 574.1836 measured value; 574.1834 MSm / z: 574.556,538,270,269,255.241,267 The mother liquor from which compound 1 (germicricon) was obtained was treated in the same manner as above using the same PLC to obtain 65 mg of compound 12 (Singeanol-I).

Production Example 2 Separation from Hazelnut Flower 15.3 g of a benzene extract of dried flowers of Hawthorne (765 g) was subjected to the same Malinck column column chromatography as in Production Example 1, and eluted from the benzene-acetic acid ethyl ester (1: 1) eluate. The obtained crude product was recrystallized from benzene to give compound 8
580 mg of (trosacrison D) was obtained.

Compound 8 (Tolosacrysone D) Yellow needles mp205-207 ° C High-resolution MS (calculated as m / z C ₃₂ H ₃₀ O ₁₀ ); 574.1836 observed; 574.1840 MSm / z: 574,556,538,508,269 Production Example 3 Separation of fresh root of Habu-so (3.5 kg) Fresh root of Habu-so (3.5 kg) was minced and extracted with benzene 7 for 8 hours at room temperature and then filtered. Extract at room temperature for 8 hours. The two extracts described above were combined and the solvent was distilled off under reduced pressure to obtain 7.4 g of a benzene extract. 7.0 g of this extract
Column chromatography using 0 g (diameter 8 cm, length 24 c
m) and elute with benzene, then chrysophanol (c
20 mg of hrysohpanol) was obtained, followed by elution with benzene-acetic acid ethyl ester (9: 1) to obtain 30 mg of emodin and 102 mg of compound 1 (germicrison). Next, 2.5 g of each fraction eluted with benzene-acetic acid ethyl ester (4: 1) was subjected to fractionation again by using Marincklot column chromatography. This n-hexane-acetic acid ethyl ester (4:
1) The elution fraction was further treated with 0.5N oxalic acid and preparative thin-layer chromatography (PLC) was used to duplicate development twice with benzene-acetic acid ethyl ester (9: 1) to scrape the yellow band. The product was extracted with ethyl acetate, washed with water three times, and then concentrated under reduced pressure to recrystallize the obtained crude crystals to obtain 24 mg of Compound 13 (trosacrison B).

Compound 13 (Tolosacrysone B) Yellow powder mp 300 ° C (decomposition) High-resolution MS (m / z) C ₃₄ H ₃₄ O ₁₁ Calculated value; 618.2099 Measured value; 618.2110 MSm / z: 618.600.582 Production Example 4 Compound 4 (8-acetyltrosacrysone) 10 mg of trosacrysone was dissolved in acetic anhydride and pyridine, allowed to stand at room temperature overnight, poured into 12 ml of ice water, and extracted with chloroform.
The extract was washed with diluted hydrochloric acid and water in this order, dried over sodium sulfate, the solution medium was evaporated under reduced pressure, and the obtained crude product was recrystallized with water-methanol to obtain 8 mg of the desired product.

Yellow needle mp 155 ℃ MSm / z (%): 330 (M +, 36.1), 288 (M + -C 2 H 2 O, 100) 270
(M ⁺ -C ₂ H ₂ O-H ₂ O, 17.7) ¹ H-NMR (CDCl ₃ ) δ: 1.43 (3H, s, C ₃ -Me), 2.40 (3H, s,
OAc), 2.82 (2H, br s, C 2 -H), 3.08 (2H, brs, C 4 -
H), 3.91 (3H, C ₆ −OMe), 6.72 (1H, d, J = 2.4Hz, C ₇ −
H), 6.87 (1H, d , J = 2.4Hz, C 5 -H), 6.971H, br s, C 10
-H), 14.70 (1H, s , C 9 -OH) Next, the present invention compounds exhibit anti-tumor activity, and will be described by way of test examples for their safety.

Test Example 1 [In vitro cell growth inhibitory effect test by KB cells] (1) Tumor cells KB cells (Human epidermoid carcinoma of the mouth) subcultured in the MEM medium for medium described below are used as live cells 1 × It was used as a cell suspension at 10 ⁴ cells / ml.

(2) Medium 10 ml of inactivated Fetal calf serum (Flow) in 90 ml of MEM medium
1 was added, and 80 g / ml of gentamicin was further added to use as a culture MEM medium.

(3) Culture method 1 x 10 ⁴ cells of live cells in a 6-well plate (Falcon, diameter 35 mm)
2 ml of s / ml cell suspension was added (day 0), and the cells were cultured in a 5% carbon dioxide incubator at 37 ° C for 24 hours. 20 μl of the compound solution of the present invention at each concentration was added, and the cells were further cultured until day 4. After removing the medium on day 4, 0.5 ml of 0.25% trypsin-EDTA solution was added.

Leave it for about 5 minutes and confirm that the cells have peeled off, then 2% FCS
-PBS (1 ml) was added, the mixture was lightly pipetted, and the number of cells in the solution was determined by a Bueker-Tuerk hemocutometer.

(4) Judgment method Litchifield &
Wilcoxon) method was used to calculate the 50% inhibitory concentration, and the 50% inhibitory concentration was 4 μg according to the standards of the National Cancer Institute (NCI).
Compounds below / ml were judged to be effective.

(5) Test results The test results are shown in Table-2.

Test Example 2 [Lymphocytic leukemia P388 test] (1) Experimental animal 1 × 10 ⁶ of each P388 leukemia cells passaged in DNB / 2 female mice
CDF ₁ female mice (5-6 weeks old, body weight)
17-20g) was used. The day of transplantation was day 0.

(2) Administration method Compound 5 was suspended in 0.5% gum arabic / physiological saline,
It was intraperitoneally administered once a day, twice a total of the first day and the fifth day. The control group was similarly administered with 0.5% gum arabic / physiological saline only.

The group to which the compound was administered was 6 in each group, and the control group was 12 in each group.

(3) Evaluation method The effect was determined according to the effect determination standard of National Cancer Institute (NCI).

The surviving animals in each group were recorded for 10 days, and T / C x 100 (%) was calculated from the median survival time (T) of the treatment group and the control group.

A T / C x 100 value of 130 or more is considered valid.

(4) Test results The test results are shown in Table-3.

Test Example 3 [Acute toxicity test] 7-week-old male Wistar rats (body weight 149 to 160 g) 7
A group of animals was subjected to the test.

A solution prepared by suspending Compound 5 in a 5% gum arabic solution was intraperitoneally administered to the rat, and 7 days after the administration, the progress was observed and
The LD ₅₀ value was 100 mg / kg or more.

From the above results, it is considered appropriate that the dose of the active ingredient in the anticancer agent of the present invention is 0.1 mg to 100 mg daily, although it varies depending on the age, body weight and degree of disease of the patient.

The 1-oxo-tetrahydroanthracene compound, which is the main component of the anticancer agent of the present invention, is a suitable solvent used in the formulation,
Using excipients, auxiliaries, etc., formulations such as liquids, powders, granules, tablets, capsules and the like can be prepared according to a conventional method for producing formulations.

In the prescription, it can be used as a compounding agent with other pharmaceutically active ingredients.

The anticancer agent of the present invention can be administered by an appropriate method such as intravenous injection in addition to oral administration.

For oral administration, tablets, pills, capsules, powders, using at least one excipient such as starch, lactose, sucrose, mannitol, carboxymethylcellulose, etc.
It can be formulated into granules and the like. For this type of formulation,
In addition to the above-mentioned excipients, for example, magnesium stearate, sodium lauryl sulfate, lubricants such as talc, dextrin, crystalline cellulose, polyvinylpyrrolidone, gum arabic, corn starch, binders such as gelatin, potato starch, carboxymethyl cellulose. Disintegrating agents such as can be used. The 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.

In the case of producing an injection, distilled water for injection, physiological saline, dextrose aqueous solution, vegetable oil for injection, propylene glycol, polyethylene glycol and the like can be generally used as a diluent. Further, if necessary, a tonicity agent, a stabilizer, an antiseptic agent, a soothing agent and the like may be added appropriately. In addition, in the case of this type of dosage form, it is desirable to dissolve in a sterilized injection medium.

EFFECTS OF THE INVENTION The compound of the present invention shows an effective concentration in an in vitro test using KB cells, exhibits a significant life-prolonging effect in a test using P388 leukemia cancer-bearing mice, and is useful as an anticancer agent.

EXAMPLES Next, the present invention will be described more specifically by showing examples, but the present invention is not limited thereto.

Example 1 6 g of compound 5 was made into fine powder, which was mixed with 30 g of lactose and 20 g of magnesium stearate, and this mixture was tabletted by a single-shot slug tablet machine to give a slug tablet having a diameter of 20 mm and a weight of about 2.3 g. Was prepared, crushed with an oscillator, sized and sieved to obtain particles of 20 to 50 mesh.

This granule contains 200 mg of compound 5 in 1 g.

Example 2 A hard capsule was filled with 100 mg of the above-mentioned compound 5 to obtain a capsule.

Example 3 0.2 g of the above-mentioned compound 5 was added according to a conventional method for producing an injection.
It was dissolved in distilled water for injection 1 heated to 60 ° C., made isotonic, and then enclosed in an ampoule. 1 ml of this injection contains 0.2 of compound 5
Contains mg.

Claims (1)

[Claims] 1. A formula [Wherein, R ¹ represents a hydrogen atom or a hydroxyl group, R ^2, R ⁵ are the same or different and each represents a hydrogen atom or a methyl group, R ^3, R ⁷ are both hydrogen atoms or hydrogen either R ^3, R ⁷ is Atom and the other R ⁴ represents a methoxy group or a methyl group, and R ⁶ represents a hydrogen atom or an acetyl group. ] The anticancer agent containing the 1-oxo- tetrahydro anthracene compound represented by these.