JPH06256244A - Indenestrol derivative - Google Patents

Abstract

PURPOSE:To provide a new indenestrol derivative giving excellent proliferation- suppressive activity on oncocytes, thus useful as an antineoplastic agent. CONSTITUTION:The indenestrol derivative of formula I or II (R<1> and R are each H or 1-5C alkyl, however, excepting the case where they are H or alkyl at the same time), e.g. (-)-(S)-6-o-methylindenestrol A of formula III. The compound of the formula I or II can be obtained by alkylating one of the hydroxyl groups in dienestrol to form a dienestrol monoalkyl ether, which is then put to ring opening reaction. The compound of the formula I or II can be safely administered orally or parenterally as a medicine, its effective dose a day per adult being pref. 0.01-80mg per kg bw for injection, while 0.5-120mg in one to several portions for oral administration.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to an indenstrol derivative having antitumor activity or a salt thereof.

[0002]

BACKGROUND OF THE INVENTION Indenstrols A and B are known to have microtubule formation-inhibiting activity (Y. Sato et al.,
Chem.Pharm.Bull., 40, No. 3, 588-592 (1992
Year) ).

[0003]

DISCLOSURE OF THE INVENTION The present inventors have found that the hydroxy groups of indenstrols A and B are monoalkylated to have a stronger antitumor effect than indenstrols A and B. Was completed.

[0004]

The present invention has the general formula

[0005]

[Chemical 2]

(In the formula, R ¹ and R ² represent a hydrogen atom or an alkyl group having 1 to 5 carbon atoms.
Except when both ¹ and R ² are a hydrogen atom or an alkyl group. ) Indestrol derivative represented by or a salt thereof.

Where R ¹ or R ² has 1 to 5 carbon atoms
When showing an alkyl group having a number, for example, methyl, ethyl, propyl, 1-methylethyl, butyl, 1-methylpropyl, 2-methylpropyl, 1,1-dimethylethyl, pentyl, 1-methylbutyl, 2-methylbutyl,
Mention may be made of linear or branched alkyl groups such as 3-methylbutyl, 1,1-dimethylpropyl, 2,2-dimethylpropyl, 1,2-dimethylpropyl and 1-ethylpropyl. It is preferably a linear or branched alkyl group having 1 to 3 carbon atoms, and most preferably a methyl group.

The above general formulas (I) and (II) of the present invention
The compound represented by can be converted into a salt according to a conventional method.

When R ¹ or R ² is a hydrogen atom, such salts include salts of alkali metals such as lithium, sodium and potassium; magnesium, calcium,
Examples thereof include salts of alkaline earth metals such as barium; and the like. Preferred are alkali metal salts such as sodium and potassium.

Of these salts, the most suitable one is a pharmacologically acceptable salt.

In the compounds having the general formulas (I) and (II), R ¹ is preferably a hydrogen atom,
^A compound or salt thereof, wherein 2 represents an alkyl group having 1 to 3 carbon atoms.

Most preferably, it is a compound or salt thereof in which R ¹ represents a hydrogen atom and R ² represents a methyl group.

The above-mentioned general formulas (I) and (II) of the present invention
As specific examples of the compound having, there can be mentioned, for example, the compounds listed in the following table.

[0014]

[Table 1] ─────────────────┬───────────────── Compound number R ¹ R ² │ Compound number R ¹ R ² ─────────────────┼───────────────── 1 H Me │ 12 Me H 2 H Et │ 13 Et H 3 H Pr │ 14 Pr H 4 H iPr │ 15 iPr H 5 H Bu │ 16 Bu H 6 H iBu │ 17 iBu H 7 H sBu │ 18 sBu H 8 H tBu │ 19 tBu H 9 H 10 H iPn │ 21 iPn H 11 H tPn │ 22 tPn H ─────────────────┴───────────────── However, , H = hydrogen atom in the table, Me = methyl,
Et = ethyl, Pr = propyl, iP
r = isopropyl, Bu = butyl, iBu = isobutyl, sBu = secondary butyl, tBu = tertiary butyl, Pn = pentyl, iPn = isopentyl, tPn = tertiary pentyl.

In the above compound table, Exemplified Compound Nos. 1, 2, 3 and salts thereof are preferable.

Most preferably, it is Exemplified Compound No. 1 or a salt thereof.

The compounds having the general formulas (I) and (II) have various isomers. In the compounds having the general formulas (I) and (II), these isomers are all represented by a single formula. Therefore, the present invention includes all of these isomers and a mixture of these isomers.

Furthermore, in the compounds of the above-mentioned general formulas (I) and (II) of the present invention, so-called "prodrug compounds" are derived in vivo from the compounds of the above-mentioned general formulas (I) and (II). Is also included.

The above general formulas (I) and (II) of the present invention
The compound having: can be produced as follows.

(1) First Step The first step is achieved by alkylating one of the hydroxy groups of dienestrol.

Examples of the alkylating agent used in this reaction include methyl iodide, ethyl iodide, propyl iodide, butyl iodide, pentyl iodide, methyl chloride, ethyl chloride, propyl bromide and butyl bromide. Halogenated hydrocarbons such as pentyl chloride; Dialkyl sulphates such as dimethyl sulphate, diethyl sulphate; Diazo hydrocarbons such as diazomethane, diazoethane, diazopropane, diazobutane;
Can be given.

When using halogenated hydrocarbons as the alkylating agent, the reaction is suitably carried out in a solvent in the presence of a base. Examples of the base used include alkali metal hydroxides such as sodium hydroxide and potassium hydroxide,
Alkali metal carbonates such as sodium carbonate, potassium carbonate, sodium amides, alkali metal amides such as potassium amide, sodium methoxide, sodium ethoxide, alkali metal alcoholates such as potassium t-butoxide, sodium hydride, hydrogen Alkali metal hydrogen compounds such as potassium hydride and lithium hydride,
It is carried out in the presence of an organic base such as triethylamine, pyridine, dimethylaniline. The ratio of dientstrol to base is usually 1 mol of dientstrol.
It is 1 to 2 mol, preferably 1 to 1.2 mol.
Further, the ratio of the diene strol and the halogenated hydrocarbons is usually 0.8 to 1 mol with respect to 1 mol of the diene strol.
It is 1.2 mol, preferably 0.9 to 1.1 mol. The reaction is usually performed in the presence of a solvent. The solvent used is not particularly limited as long as it does not affect the reaction, and it varies depending on the type of base, but for example, hydrocarbons such as benzene, toluene, xylene, cyclohexane, diethyl ether, tetrahydrofuran, dioxane, etc. Examples thereof include ethers, amides such as dimethylformamide and dimethylacetamide, and sulfoxides such as dimethylsulfoxide. The reaction conditions such as reaction temperature and reaction time will differ depending on the starting compounds used, bases, solvents, etc., but are usually 0 to 150 ° C., preferably 15 to 120 ° C. and 1 to 24 hours.

When using dialkyl sulphates as the alkylating agent, the reaction is preferably carried out in a solvent in the presence of a base. Examples of the base used include inorganic bases such as alkali metal hydroxides such as sodium hydroxide and potassium hydroxide, alkali metal carbonates such as sodium carbonate and potassium carbonate, sodium methoxide, sodium ethoxide, potassium. It is carried out in the presence of an alkali metal alcoholate such as -t-butoxide and a metal hydride compound such as sodium hydride and potassium hydride. The ratio of dientrol to base is usually dientrol
1.6 to 2.2 mol, preferably 1.8 per mol
To 1.9 mol. Further, the ratio of the diene strol and the dialkyl sulfates is usually 0.8 to 1.2 mol, preferably 0.9 to 1.
It is 1 mol. The reaction is usually performed in the presence of a solvent. The solvent used is not particularly limited as long as it does not affect the reaction and varies depending on the type of base, for example, alcohols such as methanol, ethanol and propanol, aromatic hydrocarbons such as benzene, toluene and xylene. Examples thereof include ethers such as diethyl ether, tetrahydrofuran and dioxane, and alcohols are preferable. The reaction conditions such as reaction temperature and reaction time will differ depending on the starting compounds used, bases, solvents, etc., but are usually 0 to 150 ° C., preferably 50
~ 100 ℃ for 1 ~ 24 hours.

When using diazo hydrocarbons as the alkylating agent, the reaction is carried out in the presence of a solvent. The ratio of diene strol to diazo hydrocarbons is usually 1 to 10 mol, preferably 1.5 to 2 mol, per 1 mol of diene strol. The solvent to be used is not particularly limited as long as it does not affect the reaction, ethers such as diethyl ether, tetrahydrofuran, dioxane and dimethoxyethane; esters such as ethyl acetate; amides such as dimethylformamide. Preferred are sulfoxides such as dimethyl sulfoxide. The reaction conditions such as reaction temperature and reaction time will differ depending on the starting compounds used, the solvent, etc., but are usually -10 to 100 ° C, preferably 0 to 20 ° C.
0.5 to 24 hours.

(2) Second Step The second step is achieved by subjecting the diene strol monoalkyl ether obtained in the first step to a ring closure reaction.

The reaction is carried out by contacting the diene strol monoalkyl ether with an acid. As the acid used, for example, mineral acids such as hydrochloric acid, hydrobromic acid, sulfuric acid and nitric acid, or organic acids such as formic acid, acetic acid, propionic acid, oxalic acid and malonic acid are preferably used. The reaction is suitably carried out in the presence of solvent. The solvent used is not particularly limited as long as it does not affect the reaction, for example, alcohols such as methanol, ethanol, propanol, benzene, toluene, aromatic hydrocarbons such as xylene, diethyl ether, tetrahydrofuran,
Examples thereof include ethers such as dioxane, and alcohols are preferable. The reaction conditions such as reaction temperature and reaction time will differ depending on the starting compounds used, the solvent, etc., but are usually 0 to 150 ° C., preferably 20 to 100 ° C.
1 to 24 hours at ℃.

Alternatively, the reaction is carried out by heating the diene strol monoalkyl ether. For example, it is carried out by placing dienestrol monoalkyl ether in a glass tube substituted with nitrogen gas, sealing it, and then heating. The reaction conditions such as reaction temperature and reaction time vary depending on the raw material compound used, solvent, etc.,
Usually 20 to 300 ℃, preferably 150 to 250
1 to 24 hours at ℃.

The compound (3) or the compound having the general formula (II) can also be achieved by heating the compound having the general formula (I). The reaction conditions are the same as those described above.

After completion of the reaction, the target compound thus obtained in each step is subjected to adsorption column chromatography using a carrier such as silica gel or florisil, if necessary;
Reversed-phase partition chromatography using silica gel having octadecyl group, octyl group, etc .; Partition column chromatography using cellulose such as Avicel (manufactured by Asahi Kasei Co., Ltd.) or Sephadex LH-20 (manufactured by Pharmacia). Or it can be separated and purified by an extraction method or the like utilizing the difference in the distribution ratio of the mixed impurities to the solvent.

For example, a mixture of compounds having the above general formulas (I) and (II) is extracted from the reaction mixture with a solvent such as ethyl acetate. The solvent is distilled off from the extract, and the compound having the general formulas (I) and (II) is isolated and purified from the obtained residue by utilizing its physicochemical properties. For example, it can be collected using an adsorbent. Examples of the adsorbent used include silica gel, activated carbon, or adsorbent resins such as Amberlite XAD-2, XAD-4, XAD-7 (manufactured by Rohm and Haas), Diaion HP 10, HP 20.
, HP 50, CHP20P (manufactured by Mitsubishi Kasei Co., Ltd.), polyamide gel (manufactured by Welmu) and the like are used. A solution containing the compounds having the general formulas (I) and (II) is passed through a layer of the adsorbent as described above to obtain the general formulas (I) and (I).
The impurities contained in the liquid containing the compound having I) are adsorbed and removed, or the impurities represented by the general formulas (I) and (I
It is obtained by adsorbing the compound having I) and then eluting with a suitable solvent. In order to further separate and purify the compounds having the general formulas (I) and (II) thus obtained, cellulose such as Avicel (Asahi Kasei Co., Ltd.), Sephadex LH-20 (Pharmacia) Column chromatography using
An effective method is an extraction method utilizing a difference in distribution ratio of the compound having the general formulas (I) and (II) and impurities mixed with the solvent, or a countercurrent distribution method.

The compounds having the above general formulas (I) and (II) can be separated and purified by repeating the above separation and purification means alone or in appropriate combination and repeatedly used.

[0032]

The compound of the present invention represented by the general formula (I) or (II) is a novel compound which has not been published in the literature and exhibits a strong growth inhibitory action on various human cancer cells and is useful as an antitumor agent. Is.

The above general formula (I) or (II) of the present invention
When the compound having is used as a medicine, it is mixed with itself or an appropriate pharmaceutically acceptable carrier, excipient, diluent according to a conventional method, and powder, granules, tablets, capsules,
It can be safely administered orally or parenterally in the form of injection. The dose varies depending on the target disease, administration route, number of administrations, and the like. Valid for adults, for example 1
The daily dose, when injected, is 0.0 per kg patient body weight.
1 mg to 80 mg, orally 0.5 mg to 120 mg
Is preferably administered once or in several divided doses depending on the symptoms.

[0034]

EXAMPLES The present invention will be described in more detail with reference to Reference Examples, Examples and Test Examples, but the present invention is not limited to these.

Reference Example 1. E, E-Dientrol monomethyl ether dientrol 500 g (1.9 mmol) in methanol 7.
Dissolve in 5 ml, then 1N sodium hydroxide 3.
6 ml was added. Then add 0.25 ml of dimethyl sulfate.
(2 mmol) was added, and the mixture was refluxed at 90 ° C for 7 hours. After the reaction was completed, the reaction mixture was cooled. Then, 200 ml of ethyl acetate was added to the reaction mixture for extraction. The extract was washed with water and dried over anhydrous sodium sulfate. When the solvent was distilled off from the extract, 526 mg of residue was obtained. The obtained residue was subjected to column chromatography using 20 g of silica gel, and the fraction (201 mg, yield 38%) eluted with n-hexane-benzene (3: 7) was recrystallized from methanol. When attached, a target compound having a melting point of 145-148 ° C was obtained.

1) Structural formula

[0037]

[Chemical 3]

2) Mass spectrum (m / z): 280 (M ⁺ ),
265, 251, 159, 135, 121.

3) Elemental analysis: (%) as C ₁₉ H ₂₀ O ₂ Actual value: C, 81.89; H, 7.44 Calculated value: C, 81.39; H, 7.194) ¹ H-nuclear magnetic resonance spectrum: (δ ; Ppm) Nuclear magnetic resonance spectrum (270 MHz) measured by using tetramethylsilane as an external standard in deuterated acetone is as shown below. 1.46 (6H, d, J = 6.3 Hz), 3.81 (3H, s), 5.27 and 5.3
2 (2H, q, J = 6.3 Hz), 6.86 (2H, br.d, J = 8.9 Hz), 6.94 (2H,
br.d, J = 8.9 Hz), 6.99 (2H, br.d, J = 8.9 Hz), 7.07 (2H,
br.d, J = 8.9 Hz), 8.25 (1H, br.s).

Example 1. Reference example 1. Compound obtained in
1 g was dissolved in 30 ml of methanol and 7 ml of 10 N sulfuric acid and heated under reflux for 3 hours. After the reaction was completed, the reaction mixture was diluted with water and then extracted with ethyl acetate. The extract was washed with water, dried and concentrated. 1.2 g of the obtained residue was subjected to column chromatography using 100 g of silica gel using a hexane-benzene (1: 1) mixed solution. The fraction eluted with the eluent hexane-benzene (1: 4) was collected to obtain a mixture of (±) -4′-O-methylindensetrol A and (±) -6-O-methylindensetrol A 1 g (Mixing ratio about 1: 1) was obtained. 100 μl of 2-propanol-hexane (1: 5) solution containing 5 mg of the obtained mixture was added to Chiralcel OJ.
It was injected into a column (manufactured by Daicel). Then, this was subjected to high performance liquid column chromatography, and hexane-
Elution was performed with a benzene (1: 1) mixture.

The fraction eluted at a retention time of 44 minutes was collected. This was repeated 4 times to obtain (-)-(S) -6-O-methylindensetrol A 4 mg as an oily substance.

Then, the fraction eluted at a retention time of 47 minutes was collected. Do this 4 times to get (-)-(S) -4 '
4 mg of -O-methylindensetrol A was obtained as an oil.

Then, the fraction eluted at a retention time of 69 minutes was collected. Do this 4 times to get (+)-(R) -6-
4 mg of O-methylindensetrol A was obtained as an oil.

Finally, the fraction eluted at a retention time of 89 minutes was collected. Do this 4 times to get (+)-(R) -4 '
4 mg of -O-methylindensetrol A was obtained as an oil.

Next, the structural formulas and physical property values of each compound obtained above are as follows. (I) (-)-
(S) -6-O-Methylindensetrol A 1) Structural formula

[0046]

[Chemical 4]

2) Specific optical rotation: [α] _D ²⁴ -265.9 ° (C
0.13, ethanol) 3) Circular dichroism (C = 0.65 mg / ml, ethanol)
[Θ] (nm): -29090 (239), -7810 (253), -32990
(301) (negative maximum).

4) ¹ H-nuclear magnetic resonance spectrum: (δ; pp
m) Nuclear magnetic resonance spectrum (270 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below. 1.14 (3H, d, J = 7.3 Hz), 1.24 (3H, t, J = 7.6 Hz), 2.63
(2H, q, J = 7.6 Hz), 3.82 (3H, s), 6.86 (1H, dd, J = 8.4 Hz)
, 6.92 (2H, bd, J = 8.9 Hz), 7.09 (1H, d, J = 2.3 Hz),
7.23 (2H, bd, J = 8.6 Hz), 7.27 (1H, d, J = 8.6 Hz), 8.39
(1H, bs).

5) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below. 14.2 (q), 17.2 (q), 19.8 (t), 46.3 (d), 5
5.7 (q), 110.2 (d), 112.6 (d), 116.1 (d), 120.2
(d), 127.8 (s), 130.7 (d), 137.6 (s), 137.9
(s), 145.1 (s), 151.2 (s), 157.1 (s), 158.9
(s).

(II) (-)-(S) -4'-O-methylindendrol A 1) Structural formula

[0051]

[Chemical 5]

2) Specific optical rotation: [α] _D ²⁴ -312.6 ° (C
0.12, ethanol) 3) Circular dichroism (C = 0.62 mg / ml, ethanol)
[Θ] (nm): -35340 (240), -9970 (257), -36020
(301) (Negative maximum) 4) Mass spectrum (m / z): 280 (M ⁺ ), 251, 13
Five .

5) ¹ H-nuclear magnetic resonance spectrum: (δ; pp
m) Nuclear magnetic resonance spectrum (270 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below. 1.11 (3H, d, J = 7.3 Hz), 1.24 (3H, t, J = 7.6 Hz), 2.62
(2H, q, J = 7.6 Hz), 3.83 (3H, s), 6.78 (1H, dd, J = 8.1 Hz)
, 6.98 (1H, d, J = 8.3 Hz), 7.00 (2H, bd, J = 8.9 Hz), 7.
19 (1H, d, J = 8.6 Hz), 7.30 (2H, bd, J = 8.9 Hz), 8.11 (1H,
bs).

6) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below. 14.2 (q), 17.2 (q), 19.8 (t), 46.2 (d), 5
5.5 (q), 111.5 (d), 114.1 (d), 114.6 (d), 120.4
(d), 130.5 (d), 131.0 (s), 137.6 (s), 138.5
(s), 144.0 (s), 151.4 (s), 156.4 (s), 159.3
(s).

(III) (+)-(R) -6-O-methylindendrol A 1) Structural formula

[0056]

[Chemical 6]

2) Specific rotation: [α] _D ²⁴ + 258.6 ° (C
0.14, ethanol) 3) Circular dichroism (C = 0.71 mg / ml, ethanol)
[Θ] (nm): 28730 (239), 5746 (254), 34281 (30
1) (Positive maximum).

4) ¹ H-nuclear magnetic resonance spectrum: (δ; pp
m) Nuclear magnetic resonance spectrum (270 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below. 1.14 (3H, d, J = 7.3 Hz), 1.24 (3H, t, J = 7.6 Hz), 2.63
(2H, q, J = 7.6 Hz), 3.82 (3H, s), 6.86 (1H, dd, J = 8.4 Hz)
, 6.92 (2H, bd, J = 8.9 Hz), 7.09 (1H, d, J = 2.3 Hz),
7.23 (2H, bd, J = 8.6 Hz), 7.27 (1H, d, J = 8.6 Hz), 8.39
(1H, bs).

5) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below. 14.2 (q), 17.2 (q), 19.8 (t), 46.3 (d), 5
5.7 (q), 110.2 (d), 112.6 (d), 116.1 (d), 120.2
(d), 127.8 (s), 130.7 (d), 137.6 (s), 137.9
(s), 145.1 (s), 151.2 (s), 157.1 (s), 158.9
(s).

(IV) (+)-(R) -4'-O-methylindendrol A 1) Structural formula

[0061]

[Chemical 7]

2) Specific optical rotation: [α] _D ²⁴ + 281.6 ° (C
3) Circular dichroism (C = 0.76 mg / ml, ethanol)
[Θ] (nm): 32970 (240), 9950 (257), 33640 (3
01) (Positive maximum) 4) ¹ H-nuclear magnetic resonance spectrum: (δ; ppm) Nuclear magnetic resonance spectrum (270 MHz) measured by using tetramethylsilane as an external standard in deuterated acetone is As shown. 1.11 (3H, d, J = 7.3 Hz), 1.24 (3H, t, J = 7.6 Hz), 2.62
(2H, q, J = 7.6 Hz), 3.83 (3H, s), 6.78 (1H, dd, J = 8.1 Hz)
, 6.98 (1H, d, J = 8.3 Hz), 7.00 (2H, bd, J = 8.9 Hz), 7.
19 (1H, d, J = 8.6 Hz), 7.30 (2H, bd, J = 8.9 Hz), 8.11 (1H,
bs).

5) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below. 14.2 (q), 17.2 (q), 19.8 (t), 46.2 (d), 5
5.5 (q), 111.5 (d), 114.1 (d), 114.6 (d), 120.4
(d), 130.5 (d), 131.0 (s), 137.6 (s), 138.5
(s), 144.0 (s), 151.4 (s), 156.4 (s), 159.3
(s).

Example 2. Example 1. (±)-
4'-O-methylindensetrol A and (±)-
Mixture of 6-O-methylindensetrol A 500 mg
Was placed in a glass tube under a nitrogen atmosphere and heated at 240 ° C for 2 hours. After completion of the reaction, the reaction mixture was allowed to cool, and then the obtained solid product was extracted with acetone. The solvent was distilled off from the extract. As a result of measuring a part of the obtained residue by ¹ H-nuclear magnetic resonance spectroscopy, a mixture of (±) -4′-O-methylindensetrol A and (±) -6-O-methylindensetrol A and ( It was confirmed that a mixture of ±) -4′-O-methylindensetrol B and (±) -6-O-methylindensetrol B was present at an approximate ratio of 1: 1. Next, the obtained residue was left to stand overnight at room temperature with acetic anhydride / pyridine (3 ml each) for acetylation according to a conventional method. Then, the reaction solution was poured into ice water and extracted with benzene. The extract was washed with water, dried over anhydrous sodium sulfate, and the solvent was distilled off from the extract to obtain 560 mg of a residue. The obtained residue was mixed with a hexane-benzene (1: 1) mixed solution,
It was subjected to column chromatography using 60 g of silica gel. The fraction eluted with hexane-benzene (1: 2) was collected and recrystallized from ethanol to give pale yellowish white (±) -4'-O-methylindensetrol B and (±)- 6-O-methylindensetrol B
183 mg of an acetate form of Dissolve 150 mg of the obtained acetate body in 17 ml of methanol,
0.6 ml of 5N aqueous sodium hydroxide solution was added, and the mixture was stirred at room temperature for 2 hours. The reaction mixture was poured into ice water, adjusted to pH 7 to 8 with diluted hydrochloric acid, and then extracted with 150 ml of ethyl acetate. The extract was washed with water and dried over anhydrous sodium sulfate. When the solvent was distilled off from the extract, 120 mg of residue was obtained. The obtained residue was subjected to column chromatography using 20 g of silica gel using a hexane-benzene (1: 1) mixed solution. Eluent hexane-benzene (
When the fraction eluted at 1: 4) was collected, (±) -4 '
-O-methylindensetrol B and (±) -6-
84 mg of a mixture of O-methylindensetrol B was obtained.

5 mg of the mixture obtained above was used in Example 1. It was subjected to high performance liquid column chromatography using an optically active column under the conditions described in, and eluted with a hexane-benzene (1: 1) mixed solution.

The fraction eluted at a retention time of 38 minutes was collected. Do this 4 times to get (-)-(S) -4'-O-
4 mg of methylindensetrol B was obtained as an oil.

Then, the fraction eluted at a retention time of 42 minutes was collected. Do this 4 times to get (-)-(S) -6-
4 mg of O-methylindensetrol B was obtained as an oil.

Then, the fraction eluted at a retention time of 66 minutes was collected. Do this 4 times to get (+)-(R) -6-
4 mg of O-methylindensetrol B was obtained as an oil.

Finally, the fraction eluted at a retention time of 84 minutes was collected. Do this 4 times to get (+)-(R) -4 '
4 mg of -O-methylindensetrol B was obtained as an oil.

Next, the structural formulas and physical property values of each compound obtained above are as follows. (V) (−)-(S) -4′-O-methylindensetrol B 1) Structural formula

[0071]

[Chemical 8]

2) Specific optical rotation: [α] _D ²⁴ -332 ° (C 0.1
1, ethanol) 3) Circular dichroism (C = 1.05 mg / ml, ethanol)
[Θ] (nm): -6907 (244) (negative maximum), -4050 (26
4), -6760 (287) 4) ¹ H-nuclear magnetic resonance spectrum: (δ; ppm) Nuclear magnetic resonance spectrum (270 MHz) measured by using tetramethylsilane as an external standard in deuterated acetone is as follows. As shown in. 0.39 (3H, t, J = 7.5 Hz), 1.59 (1H, m), 1.90 (1H, m),
2.16 (3H, d, J = 2.0 Hz), 3.84 (3H, s), 3.87 (1H, bs),
6.69 (1H, dd, J = 7.9 and 2.3 Hz), 6.82 (1H, d, J = 2.3
Hz), 7.02 (2H, bd, J = 8.9 Hz), 7.24 (1H, d, J = 7.9 Hz),
7.36 (2H, bd, J = 8.9 Hz), 8.09 (1H, bs).

5) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below.

8.5 (q), 11.7 (q), 24.0 (t), 5
1.0 (d), 55.5 (q), 106.9 (d), 112.2 (d), 114.6
(d), 123.8 (d), 129.9 (s), 130.9 (d), 133.6
(s), 137.8 (s), 146.4 (s), 149.3 (s), 157.6
(s), 159.5 (s).

(VI) (-)-(S) -6-O-methylindendrol B 1) Structural formula

[0076]

[Chemical 9]

2) Specific rotation: [α] _D ²⁴ -279 ° (C 0.0
7, ethanol) 3) Circular dichroism (C = 0.75 mg / ml, ethanol)
[Θ] (nm): -5600 (239), -2980 (261), -5800 (29
0) (Negative maximum) 4) Mass spectrum (m / z): 280 (M ⁺ ), 251 5) ¹ H-nuclear magnetic resonance spectrum: (δ; ppm) Tetramethylsilane is used as an external standard in deuterated acetone. The nuclear magnetic resonance spectrum (270 MHz) measured by using is as shown below. 0.39 (3H, t, J = 7.5 Hz), 1.61 (1H, m), 1.94 (1H, m),
2.19 (3H, d), 3.84 (3H, s), 3.84 (1H, bs), 6.75 (1H, d
d, J = 8.1 and 2.5 Hz), 6.92 (1H, d, J = 2.5 Hz), 6.9
4 (2H, bd, J = 8.6 Hz), 7.28 (2H, bd, J = 8.9 Hz), 7.33 (1
H, d, J = 8.3 Hz), 8.44 (1H, bs).

6) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below.

8.5 (q), 11.7 (q), 24.0 (t), 5
1.0 (d), 55.6 (q), 105.4 (d), 110.7 (d), 116.4
(d), 120.9 (s), 123.8 (d), 131.1 (d), 133.2
(s), 138.9 (s), 147.1 (s), 149.5 (s), 157.3
(s), 160.3 (s).

(VII) (+)-(R) -6-O-methylindendrol B 1) Structural formula

[0081]

[Chemical 10]

2) Specific rotation: [α] _D ²⁴ + 262 ° (C 0.1
0, ethanol) 3) Circular dichroism (C = 0.98 mg / ml, ethanol)
[Θ] (nm): 5660 (239), 3010 (261), 5930 (290)
(Positive maximum) 4) ¹ H-nuclear magnetic resonance spectrum: (δ; ppm) Nuclear magnetic resonance spectrum (270 MHz) measured by using tetramethylsilane as an external standard in deuterated acetone is as shown below. Is. 0.39 (3H, t, J = 7.5 Hz), 1.61 (1H, m), 1.94 (1H, m),
2.19 (3H, d), 3.84 (3H, s), 3.84 (1H, bs), 6.75 (1H, d
d, J = 8.1 and 2.5 Hz), 6.92 (1H, d, J = 2.5 Hz), 6.9
4 (2H, bd, J = 8.6 Hz), 7.28 (2H, bd, J = 8.9 Hz), 7.33 (1
H, d, J = 8.3 Hz), 8.44 (1H, bs).

5) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below.

8.5 (q), 11.7 (q), 24.0 (t), 5
1.0 (d), 55.6 (q), 105.4 (d), 110.7 (d), 116.4
(d), 120.9 (s), 123.8 (d), 131.1 (d), 133.2
(s), 138.9 (s), 147.1 (s), 149.5 (s), 157.3
(s), 160.3 (s).

(VIII) (+)-(R) -4'-O-
Methylindensetrol B 1) Structural formula

[0086]

[Chemical 11]

2) Specific optical rotation: [α] _D ²⁴ + 340 ° (C 0.1
2, ethanol) 3) Circular dichroism (C = 1.05 mg / ml, ethanol)
[Θ] (nm): 6480 (244) (positive maximum), 3920 (264)
, 5350 (287) 4) ¹ H-nuclear magnetic resonance spectrum: (δ; ppm) Nuclear magnetic resonance spectrum (270 MHz) measured by using tetramethylsilane as an external standard in deuterated acetone is as shown below. Is. 0.39 (3H, t, J = 7.5 Hz), 1.59 (1H, m), 1.90 (1H, m),
2.16 (3H, d, J = 2.0 Hz), 3.84 (3H, s), 3.87 (1H, bs),
6.69 (1H, dd, J = 7.9 and 2.3 Hz), 6.82 (1H, d, J = 2.3
Hz), 7.02 (2H, bd, J = 8.9 Hz), 7.24 (1H, d, J = 7.9 Hz),
7.36 (2H, bd, J = 8.9 Hz), 8.09 (1H, bs).

5) ¹³ C-nuclear magnetic resonance spectrum: (δ;
ppm) Nuclear magnetic resonance spectrum (67.8 MHz) measured using tetramethylsilane as an external standard in deuterated acetone is as shown below.

8.5 (q), 11.7 (q), 24.0 (t), 5
1.0 (d), 55.5 (q), 106.9 (d), 112.2 (d), 114.6
(d), 123.8 (d), 129.9 (s), 130.9 (d), 133.6
(s), 137.8 (s), 146.4 (s), 149.3 (s), 157.6
(s), 159.5 (s).

Test Example 1. Antitumor activity Below, the result of having measured the antitumor activity about the compound of the present invention is described. The measurement is carried out by the method of Array (Michael C. Alley) et al. (Cancer Research, 48, 589-601 (198
8 years)), and the antitumor effect of (−)-(S) -4′-O-methylindensetrol B was examined. That is,
Various types of cancer cells were added to a 96-well culture plate at different cell numbers.
100 μl medium (RPMI-1640 + 10% FCS) was suspended. However, MKN-1, MKN-7, MKN-74 is 1x
10 ⁴ , RERF-LC-MA, SBC-5, WiDr, SW-480
2 × 10 ³ and MCF-7 and KU-2 were suspended 5 × 10 ³ .
After culturing these suspensions at 37 ° C. under 5% CO ₂ for 24 hours, 100 μl of medium containing the compound was added, and the cells were further cultured under the same conditions for 72 hours. After culturing, 50μg / 50μ
l MTT was added, and the cells were further incubated at 37 ° C under 5% CO _{2 for} 4 hours. The culture solution was removed, 150 μl of DMSO was added and stirred, and the cell growth rate was calculated from the absorbance at 540 nm. The concentrations of compounds required to inhibit cell proliferation by 50% (IC ₅₀ ; μg / ml) are shown in Table 2.

[0091]

[Table 2] Human tumor cell growth inhibitory effect of (-)-(S) -4'-O-methylindensetrol B ─────────────────────── ───────────── Cell-derived tissue type Cell doubling time IC ₅₀ (μg / ml) (hours) (72 hours) ──────────────── ──────────────────── MKN-1 Gastric cancer Adenosquamous carcinoma 31 15.5 MKN-7 Gastric cancer Differentiated adenocarcinoma 43 17.25 MKN-74 Gastric cancer Well-differentiated tubular adenocarcinoma 36 4.9 RERF-LC-MA Lung cancer Small cell cancer 24 0.585 SBC-5 Lung cancer Small cell cancer 21 5.05 MCF-7 Breast cancer IDC ¹⁾ 36 7.0 WiDr Colon cancer Adenocarcinoma 20 1.57 SW-480 Colon cancer Adenocarcinoma 27.3 6.7 KU-2 Renal Cancer Adenocarcinoma 22 0.66 ─────────────────────────────────── 1) infiltrating ductal carcinoma Table 2. As described above, various types of (-)-(S) -4'-O-methylindensetrol B are available. Against human tumor cells, it showed excellent growth inhibitory effect.

[0092]

The indenstrol derivative of the present invention is
It has an excellent antiproliferative effect on tumor cells and is useful as an antitumor agent.

Claims (1)

[Claims] 1. A general formula: (In the formula, R ¹ and R ² represents an alkyl group having 1 to 5 carbon number of hydrogen atoms or carbon. However, R ¹ and R ²
Except when both are a hydrogen atom or an alkyl group. )
The indensetrol derivative shown by or its salt.