HK1116489A - Anticancer compound, intermediate therefor, and processes for producing these - Google Patents

Description

Anticancer compound and intermediate and production method thereof

Technical Field

The invention relates to 2-acetyl-2, 3 dihydro-5-hydroxy naphtho [2, 3-b ] furan-4, 9-dione, a preparation method thereof and a method for preparing 2- (1-hydroxyethyl) -5-hydroxy naphtho [2, 3-b ] furan-4, 9-dione with anticancer activity from the compounds. The present invention also relates to an anticancer agent comprising a racemate or an alpha-isomer of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione as an active ingredient

Background

The molecular formula of the 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione is as follows:

is a compound having optical activity contained in a trumpet flower, a natural yew tree (Taheebo) (tabebia avellandea lorentz ex greseb), lorentz, alligator alternanthera, has a β isomer and is considered to have excellent anticancer activity (for example, see: patent document 1). However, a method for obtaining 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-B ] furan-4, 9-dione other than the extraction from the above-mentioned plants is not known, and therefore, 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-B ] furan-4, 9-dione has not been completely used as a drug because of the scarcity of the above-mentioned plants and the very low yield (0.05%) (see, for example, non-patent document 1).

[ patent document 1] JP2669762

[ non-patent document 1] Shinichi Ueda et al, phytochemistry, 1994, Vol.36(2), p.323-325

Disclosure of Invention

Therefore, it would be desirable to inexpensively and easily synthesize 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione, which can be used as a drug.

5-hydroxynaphthalene-1, 4-dione (also referred to as juglone) can be obtained at a relatively low cost, and the present inventors have found that 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione can be synthesized inexpensively and easily in a high yield by using juglone as a starting material to complete the present invention. Accordingly, the present invention relates to a method for preparing 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione by easily obtaining 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione as an intermediate through juglone. The present inventors have successfully practiced the present invention, and 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione extracted according to the present method can be obtained in the form of racemate which can be separated into α -and β -isomers by conventional optical methods. Also, the racemate and the α -isomer are considered to be safer than the β -isomer.

The present invention provides a method for inexpensively and easily preparing 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione. Furthermore, although 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione extracted from plants is a β -isomer, the method of the present invention can provide the compound as a racemic mixture. Further, an enantiomer of the β -isomer (hereinafter referred to as α -isomer), which cannot be obtained from plants, can be obtained, and it is chirally separated from the racemic mixture, for example, by column chromatography or optical methods.

Detailed Description

In a first embodiment, the present invention provides a process for the preparation of a compound of formula (III):

wherein R is C₁-C₆An alkyl group comprising: reacting a compound of formula (I) with a compound of formula (II) in the presence of a base:

wherein R is as defined above, X¹And X²Each independently is a halogen atom.

In another embodiment, the present invention provides a method of preparing a compound of formula (III):

wherein R is C₁-C₆An alkyl group comprising: reacting a compound of formula (I) with a compound of formula (II) in the presence of a base:

wherein R is as defined above, and X1 and X2 are each independently a halogen atom, to provide a compound of formula (VII)

Wherein R is as defined above, followed by oxidation of the resulting compound of formula (VII) with an oxidizing agent.

As a preferred embodiment, the present invention provides a process for the preparation of a compound of formula (IV).

Wherein R is as defined above, and the wavy line means a racemate further comprising the step of reducing the compound of the general formula (III) obtained in any one of the above-mentioned methods with a reducing agent.

In addition, the present invention provides a process for preparing a racemic mixture of formula (IV):

wherein R is as defined above, the wavy line means a racemate comprising: reducing the compound of formula (III) by a reducing agent.

Wherein R is as defined above, said compound (III) can be prepared directly from a compound of formula (I) or via a compound of formula (VII).

In another embodiment, the present invention provides a process for preparing the α -and β -isomers of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione, comprising: resolving a compound of general formula (IV):

wherein R is as defined above, and the wavy line means a racemate.

In a further preferred embodiment, the present invention provides a process for the preparation of a plurality of the compounds described above, comprising the steps of

Reacting dimethylamine dissolved in a solvent with a compound of formula (V):

to give compounds of the general formula (VI)

Reacting the resulting compound of formula (VI) with 5 to 15 wt% of an acidic solution (W/W) to form a compound of formula (I), and

comprising carrying out any of the above-mentioned processes by using the resulting compound of formula (I).

In another embodiment, the present invention provides a compound of formula (VII)

Wherein R is as defined above. The compound can be used as an intermediate for preparing 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione.

In another embodiment, the invention provides 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione having (+) -optical rotation in methanol (c0.25, 25 ℃), referred to as the α -isomer

In another embodiment, the present invention provides an anticancer agent comprising a racemate of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione as an active ingredient.

In another embodiment, the present invention provides an anticancer agent comprising an α -isomer of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione as an active ingredient.

Oxidizing agents referred to herein include, but are not limited to, manganese compounds (e.g., MnO)₂、KMnO₄) Chromium compounds (e.g.: CrO₃Or Na₂Cr₂O₇) Lead compounds [ such as: PbO, PbO₂Or Pb (OCOCH)₃)₄]And other metal compounds (e.g., HgO, AgO, Ag)₂O、AgNO₃、CuCl₂Or FeCl₃) Halogen and halogen compounds (such as: cl₂、Br₂、I₂、NaClO、KBrO₃Or KIO₄) Oxygen, ozone, peroxides [ such as: h₂O₂、Na₂O₂Or (C)₆H₅CO)₂O₂]Or peracids and salts (e.g. CH)₃CO₃H、C₆H₅CO₃H or K₂S₂O₈)。

The term "halogen" as referred to herein includes fluorine, chlorine, bromine, iodine. Preferably "halogen" is bromine.

The term "C" as referred to herein₁-C₆Alkyl "may be a straight or branched chain alkyl, such as: methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, n-pentyl or n-hexyl. In the present invention, preferred is C₁-C₆Alkyl is methyl.

The term "base" referred to herein may be an organic or inorganic base. Organic bases include, for example, pyridine, DMAP (4-dimethylaminopyridine), quinoline, isoquinoline, triethylamine, diisopropylethylamine, DBU (1, 8-diazabicyclo [5.4.0] undec-7-ene) or DBN (1, 5-diazabicyclo [4.3.0] non-5-ene), but are not limited to those listed above. The inorganic base includes hydroxides, carbonates or bicarbonates of basic or alkaline earth metals, such as sodium hydroxide, potassium hydroxide, calcium hydroxide, sodium carbonate or sodium bicarbonate, but is not limited to the above list. In the present invention, the preferred base is DBU.

The term "reducing agent" as referred to herein includes, for example: sodium borohydride (NaBH)₄) Potassium borohydride, lithium borohydride, sodium aluminum hydride, potassium aluminum hydride, lithium aluminum hydride, zinc borohydride, sodium triacetoxyborohydride, pyridine/boron, sodium cyanoborohydride, sodium mercuric oxide, H₂/Pd、H₂/Pd-C、H₂/Pt、H₂/PtO₂、H₂Rh and H₂Ni/La, but not limited thereto. The preferred reducing agent is sodium borohydride.

The compound of formula (VI) obtained above may optionally be isolated using methods conventional in the art, such as fractional crystallization or chiral column chromatography. Furthermore, the optically active 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-B ] furan-4, 9-dione can be obtained by a known method using a conventional chiral reducing agent such as a chiral borane derivative (e.g., (-) or (+) -B-chlorodi-3-pinylborane or BINAP (R) or (S) -2, 2 '-bis (diphenylphosphino) -1, 1' -binaphthyl)) instead of the above-mentioned reducing agent.

The process of the invention can be illustrated as follows:

r, X therein¹And X²The definition of (a) is the same as above.

Step 1 in the above protocol is preferably in accordance with Chaker, l.; pautet, f.; fillon, h., was performed as described in chem. pharm. bull (1994, 42, 2238-. Meanwhile, the starting material 5-hydroxynaphthalene-1, 4-dione (also referred to as juglone) in step 1 of the above scheme is commercially available, for example, from tokyo chemical industries co. In addition, details on juglone are described in the Merck Index, 13 th edition, page 5288 and references therein.

The inventors have unexpectedly found that similar results (substituent selectivity and yield) can be obtained even when a solution of dimethylamine in a solvent is used instead of dimethylamine (boiling point: -6 ℃) in the process disclosed in the above-mentioned document. Accordingly, the present invention also provides a process according to step 1, comprising the addition of a solution of juglone in toluene to a solution of dimethylamine in a solvent.

In step 1 of the above scheme, the solvent for juglone is not limited, and includes any solvent commonly used in the art. The preferred solvent is toluene. The solvent for dimethylamine is also not specified, but includes any solvent commonly used in the art. Preferred solvents include water (H)₂O), hexane, Tetrahydrofuran (THF), diethyl ether, toluene, methanol and ethanol.

The reaction of step 1 may be carried out at a temperature in the range of-78 deg.C to the reflux temperature of the solvent, preferably from 40 deg.C to room temperature. In particular, the reaction temperature is most advantageously from-40 to 0 ℃ in terms of selectivity and ease of operation.

The reaction in step 2 of the above scheme may preferably be according to De Oliveira, a.; ferreira, d.t.; raslan, d.s. in Tetrahedron lett, 1988, 29, 155-. The inventors have unexpectedly found that the concentrated hydrochloric acid used in the process described in the above-mentioned document can be replaced with 5 to 15% acidic aqueous solution to achieve similar yields. The aqueous acid solution may be a solution of any acid used in the usual hydrolysis reactions, preferably in an aqueous hydrochloric or sulphuric acid solution. Furthermore, dioxane may optionally be added dropwise to the above aqueous solution. Meanwhile, the concentration of the acid solution is not specified, and may be in any range as long as hydrolysis can be performed, and is preferably 5 to 15% from the viewpoint of safety and ease of handling. Accordingly, the present invention also provides a process of step 2 comprising using 5 to 15% by weight of an aqueous acid solution, preferably 5 to 15% by weight of an aqueous hydrochloric acid solution. Further, although not particularly limited, the reaction of the 2 nd step is preferably carried out under heating under reflux.

The reaction of step 3 in the above scheme can be carried out in a manner similar to that described by Hagiwara et al (Hagiwara, h.; Sato, k.; Nishino, d.; Hoshi, t.; Suzuki, t.; Ando, m., j.chem.soc. perkin trans.1, 2001, 2946-. Although the literature reports a yield of 60% of product, replacing the starting material with the 2-hydroxy juglone described in the literature results in very low yields. According to the intensive studies of the present inventors, it was found that by using the reactant methylvinyl ketone within 24 hours, preferably within 3 hours after the purification by distillation, the yield of the reaction in the step is greatly improved. The present invention therefore also provides an improved process for step 3, i.e. the use of methyl vinyl ketone which has just been purified by distillation, preferably within one hour after purification. In step 3, a mixture of compound (III) and compound (VII) can be obtained. Compound (VII) can be converted to compound (III) by treatment with an oxidizing agent in the reaction of step 4.

The solvent for methyl vinyl ketone and bromine used in step 3 is not specified, but is preferably isopentane or hexane. On the other hand, the solvent for 2-hydroxyjuglone is not specified, but is preferably tetrahydrofuran or diethyl ether. The reaction temperature in step 3 is not specified, but is preferably room temperature.

The reactions in scheme steps 4 and 5 above can be carried out according to methods known in the art, preferably by the methods described in Hagiwara et al (Hagiwara, h.; Sato, k.; Nishino, d.; Hoshi, t.; Suzuki, t.; Ando, m., j.chem.soc.perkin trans.1, 2001, 2946-.

Meanwhile, the reaction solvent of step 4 is not specified, but is preferably a halogenated hydrocarbon such as chloroform or dichloromethane. Further, the reaction in step 4 is preferably carried out under heating under reflux, however, this is not limitative.

The reaction solvent in step 5 is not specified, but is preferably a mixture of chloroform and ethanol, particularly a chloroform: ethanol mixture in a volume ratio (v/v) of 4: 1. Further, the reaction of step 5 is preferably carried out at 0 ℃, however, this is not limited.

The invention is illustrated by, but not limited to, the following examples.

In the following example, the following tools are used,

1H-NMR (hydrogen nuclear magnetic resonance): UNITY INOVA 500 (manufacturer varean), nmr solvent: CDCl₃(internal standard material: Tetramethylsilicon (TMS));

melting point apparatus: MP-J3(Yanaco)

Example 1

Preparation of 2-dimethyl juglone

To a solution of 5-hydroxynaphthalene-1, 4-dione (also known as juglone) (171 mg, 1 mmol) in toluene (5 ml) at-20 ℃ was added dimethylamine (0.75 ml, 2.0M in tetrahydrofuran, 1.5 mmol). The mixture was stirred at-20 ℃ for 1 hour. Dimethylamine (0.75 ml of a 2.0M solution in tetrahydrofuran, 1.5 mmol) was then added to it, and the mixture was stirred at-20 ℃ for 30 minutes, after which the solvent was evaporated under vacuum. The residue was purified by silica gel column chromatography (chloroform/ethyl acetate ═ 20/1(V/V)) to isolate 2-dimethylamino juglone (87.2 mg, 40%) and 3-dimethylamino juglone (28.8 mg, 13%).

2-dimethylamino juglone

Melting point: 147 to 148 DEG C

¹H-NMR(CDCl₃)：δ3.25(s，6H)，5.72(s，1H)，7.20(dd，1H，J＝1.2，8.3Hz)，7.45-7.51(m，2H)，13.0(s，1H)。

3-dimethylamino juglone

¹H-NMR(CDCl₃)：δ3.23(s，6H)，5.84(s，1H)，7.15(dd，1H，J＝3.7，6.1Hz)，7.56-7.59(m，2H)，11.9(s，1H)。

Example 2

The reaction was carried out in a similar manner to that described in example 1, except that-40 ℃ was substituted for-20 ℃ to obtain 2-dimethyljuglone (104 mg, 48%) and 3-dimethyljuglone (20 mg, 10%).

Example 3

The reaction was carried out in a similar manner to that described in example 1 except that tetrahydrofuran was replaced with water as a solvent of dimethylamine, and 0.15 ml of an aqueous dimethylamine solution (50% aqueous solution, 1.5 mmol) was used at a reaction temperature of 0 ℃ to obtain 2-dimethylaminocarpenoquinone (97 mg, 45%) and 3-dimethylaminocarpenoquinone (67 mg, 31%). The use of water instead of an organic solvent makes this process more advantageous in terms of environment and safety.

Example 4

Preparation of 2-hydroxy juglone

To a solution of 2-dimethylaminocarpenoquinone (1.95 g, 9mmol) in dioxane (45 ml) was added 10% hydrochloric acid (10 ml) and the mixture was heated to reflux for 30 min. The mixture was cooled to room temperature, and then the reaction solution was extracted with chloroform. The combined organic layers were washed with brine, dried over sodium sulfate, filtered, and the solvent was evaporated under vacuum to give 2-hydroxyjuglone (1.67 g, 97%) as a brown solid.

Melting point: 220 to 221 DEG C

¹H-NMR(CDCl₃)：δ6.31(1H，s)，7.35(1H，dd，J＝8.5，1.2Hz)，7.44(1H，s)，7.59(1H，t，J＝8.5Hz)，7.69(1H，dd，J＝8.5，1.2Hz)，12.33(1H，s)。

Example 5

Preparation of 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione and 2-acetyl-2, 3 dihydro-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione.

To a solution of methyl vinyl ketone (10.5 g, 150mmol) in pentane (150 ml) at-15 deg.C was added a solution of bromine (25 g, 156mmol) in pentane (30 ml). The mixture was stirred at-15 ℃ for 10 minutes, then the solvent was evaporated under vacuum to give a colorless oil. This oil was then added to a solution of 2-hydroxy juglone (4.75 g, 25mmol) in tetrahydrofuran (250 ml) after which DBU was further added at 0 ℃ and the mixture was stirred at room temperature overnight. 10% hydrochloric acid was added to the mixture, and the reaction mixture was extracted with chloroform. The combined organic layers were washed with brine, dried over sodium sulfate, filtered and the solvent was evaporated under vacuum. The residue was purified by silica gel column chromatography (eluent: chloroform/ethyl acetate ═ 9/1(v/v)) to give an orange solid mixture (6.14 g, 95%) comprising 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione and 2-acetyl-2, 3-dihydro-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione in a ratio of 1: 5. The solid mixture was separated by silica gel column chromatography (eluent: chloroform) to give 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione and 2-acetyl-2, 3-dihydro-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione. 2-acetyl-2, 3-dihydro-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione

Melting point: 175 to 182 ℃ (decomposition)

¹H-NMR(CDCl₃)：δ2.39(3H，s)，3.39(2H，d，J＝9.5Hz)，5.30(1H，t，J＝9.5Hz)，7.26(1H，dd，J＝8.0，1.0Hz)，7.56(1H，t，J＝8.0Hz)，7.65(1H，dd，J＝8.0，1.0Hz)，12.18(1H，s)。

2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione

Melting point: 208 to 220 ℃ (decomposition)

¹H-NMR(CDCl₃)：δ2.67(3H，s)，7.33(1H，dd，J＝8.5，1.0Hz)，7.60(1H，s)，7.67(1H，t，J＝8.3Hz)，7.81(1H，dd，J＝7.4，1.0Hz)，12.13(1H，s).

Example 6

Preparation of 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione

To a solution of 2-acetyl-2, 3 dihydro-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (2.4 g, 9.4mmol) in chloroform (50 ml) was added 20 g of manganese dioxide (manufactured by Aldrich corp., 85% active manganese dioxide, < 5 μm), and the resulting suspension was heated to reflux for 1 day. The mixture was cooled to room temperature and then filtered. The filtrate was evaporated in vacuo and the residue was purified by silica gel column chromatography (eluent: chloroform) to give 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (0.718 g, 33%).

Melting point: 208 to 220 ℃ (decomposition)

¹H-NMR(CDCl₃)：δ2.67(3H，s)，7.33(1H，dd，J＝8.5，1.0Hz)，7.60(1H，s)，7.67(1H，t，J＝8.3Hz)，7.81(1H，dd，J＝7.4，1.0Hz)，12.13(1H，s)。

Alternatively, in the process of example 6 above, manganese dioxide (Aldrich corp., 90% manganese dioxide for batteries, < 10 microns) can be used instead (Aldrich corp., 85% active manganese dioxide, < 5 microns) (see example 7 below).

Example 7

To a solution of 2-acetyl-2, 3 dihydro-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (0.5 g, 1.95mmol) in chloroform (50 ml) was added 10 g manganese dioxide (aldrich corp., 90% manganese dioxide for battery, < 10 μm), and the resulting suspension was heated to reflux for 3 days. The mixture was cooled to room temperature and then filtered. The filtrate was evaporated in vacuo and the residue was purified by purified silica gel column chromatography (eluent: chloroform) to give 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (0.216 g, 44%) and 2-acetamido-2, 3-dihydro-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (0.255 g, 51%).

The present process provides a process for the preparation of 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione at a lower cost than example 6.

Example 8

Preparation of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione

To a solution of 2-acetyl-5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (694 mg, 2.73mmol) obtained in example 5 in chloroform (100ml) and ethanol (25 ml) was added sodium borohydride (515 mg, 13.7mmol) at 0 ℃. The mixture was stirred for 30 minutes, and then the reaction was quenched by adding 10% hydrochloric acid to the mixture. The aqueous layer was extracted twice with chloroform, and the extract was washed successively with water and brine. The mixture was evaporated in vacuo and purified by silica gel column chromatography (eluent: chloroform) to give 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (516 mg, 74%) as a racemic mixture as yellow crystals.

Melting point temperature: 148 to 149 deg.C

¹H-NMR(CDCl₃)：δ1.66(3H，d，J＝6.8Hz)，2.31(1H，brs)，5.05(1H，m)，6.84(1H，s)，7.27(1H，dd，J＝8.3，1.0Hz)，7.62(1H，t，J＝8.0Hz)，7.75(1H，dd，J＝8.0，0.9Hz)，12.18(1H，s)。

The resulting racemic mixture can be separated by chiral column chromatography under the following conditions to give its enantiomers.

Column: SUMICHIRAL OA-4500(4.6mm * X250 mm)

Mobile phase: ethane/2-propanol/methanol 95: 4: 1

Detecting light: UV254nm

Flow rate: 1.0 mL/min

Temperature: room temperature (constant temperature about 25 deg.C)

Injection amount: 5 μ L (0.1mg/mL methanol)

Retention time: 30.8 and 34.4 minutes.

The enantiomers of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione can also be separated with "hexane/ethanol 95: 5" instead of "hexane/2-propanol/methanol 95: 4: 1" under the above conditions as the mobile phase and in this case the holding times are 24.9 and 27.4 minutes. Under the same conditions, the retention time for chiral column chromatography to obtain the naturally derived 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (also known as β -isomer) was 27.4 minutes.

It has been found that, considering the respective retention times of the enantiomers obtained by HPLC separation of the synthesized 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione under the above conditions using an optically active column, the unnatural form (. alpha. -isomer) elutes first and the natural form (. beta. -isomer elutes later). The properties are shown in Table 1 below.

TABLE 1

	Non-natural form (alpha-isomer)	Natural type (beta-isomer)
			Purity of	≥99％	≥99％
Melting Point	172-173.5℃	171-172℃

Anticancer activity and toxicity to normal cells were examined by using the racemate, α -isomer and β -isomer of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione obtained in example 8, as shown below.

Test 1: anticancer activity

(1) Anticancer Activity against PC-3 cells (prostate cancer cells) (1)

First, PC-3 cells (manufactured by Department of Laboratory Products of Dainippon pharm. Co. Ltd.) were seeded in a 35mm dish containing DMEM (Dulbecco's modified Eagle medium) medium at a concentration of 20% and 1X 10⁵Fetal bovine serum in ml. The cells were cultured at 36 ℃ under 5% carbon dioxide for 1 day, and all of the cells were confirmed to be adhered to the bottom of the culture dish. The resulting pc-3 cells were divided into three groups, to the first group was added (. + -.) 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ]]Furan-4, 9-dione (racemate) at concentrations of 0.5mM, 0.05mM and 0.005mM, and doxorubicin (wakenyaku, inc.) at concentrations of 0.5mM, 0.05mM and 0.005mM was added to the second group as a control. After these groups were cultured at 36 ℃ for 3 days, viable cells were counted to calculate cell viability. The results are shown in Table 2.

TABLE 2

Comparison of anticancer Activity on PC-3 cells-1
			Concentration (mM)	Racemate treatment (% viability)	Adriamycin treatment (% viability)
0.5	0	0
			0.05	0	0
0.005	50	42

(2) Anticancer Activity against PC-3 cells (2)

The PC-3 cells obtained in a similar manner to (1) above were divided into three groups, to the first group (. + -.) -2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (racemate) was added at concentrations of 0.5mM, 0.05mM, 0.005mM and 0.0005mM, to the second group was added non-natural type 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (α -isomer) at concentrations of 0.5mM, 0.05mM, 0.005mM and 0.0005mM, and to the third group was added natural type 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione (β -isomer) at a concentration of 0.5mM, 0.05mM, 0.005mM and 0.0005mM, mitomycin was added as a control to the fourth group at concentrations of 0.5mM, 0.05mM, 0.005mM and 0.0005 mM. After these groups were cultured at 36 ℃ for 3 days, viable cells were counted to calculate cell viability. The results are shown in Table 3.

TABLE 3

Comparison of anticancer Activity on PC-3 cells-2
					Concentration (mM)	Racemate treatment (% viability)	Alpha-isomer treatment (% viability)	Beta-isomer treatment (% viability)	Mitomycin treatment (% viability)
0.5	0	0	0	0
					0.05	0	0	0	0
0.005	30	30	10	10
					0.0005	50	60	40	40

(3) Anticancer Activity against A-549 cells (Lung cancer cells)

The anticancer activity of A549 cells was studied in a similar manner to that in (2) above, and the results are shown in Table 4.

TABLE 4

Comparison of anticancer Activity on A-549 cells
					Concentration (mM)	Racemate treatment (% viability)	Alpha-isomer treatment (% viability)	Beta-isomer treatment (% viability)	Mitomycin treatment (% viability)
0.5	0	0	0	0
					0.05	10	10	10	0
0.005	50	40	30	10
					0.0005	70	70	60	50

(4) Anticancer Activity against MCF-7 cells (Breast cancer cells)

The anticancer activity of MCF-7 cells was studied in a similar manner to that described in (2) above, and the results are shown in Table 5.

TABLE 5

Comparison of anticancer Activity on MCF-7 cells
					Concentration (mM)	Racemate treatment (% viability)	Alpha-isomer treatment (% viability)	Beta-isomer treatment (% viability)	Mitomycin treatment (% viability)
0.5	0	0	0	0
					0.05	20	30	20	10
0.005	50	60	50	30
					0.0005	90	90	70	60

Test 2: toxicity assay for human skin normal cells (cell system FB cells), human normal hepatocytes (cell system Hc cells), human intestinal normal cells (cell system IE cells) and human normal lung cells (MRC-5)

First, each of the normal human cells was seeded in a 35mm culture dish containing DMEM (Dulbecco's modified Eagle medium) medium containingHas a 20% concentration of 1 × 10⁵Per ml fetal bovine serum. The cells were cultured at 36 ℃ under 5% carbon dioxide for 1 day, and all of the cells were confirmed to adhere to the bottom of the culture dish. Racemic, α -isomer and β -isomer were each dissolved in dimethyl sulfoxide to give solutions at concentrations of 0.5mM, 0.05mM, 0.005mM and 0.0005mM, and 2. mu.l of the above solution was added to each cell. The cells were cultured at 36 ℃ for 3 days, and viable cells were counted with 0.25% trypan blue to calculate cell viability. The results are shown in tables 6 to 9. The commercial anticancer drug mitomycin was also tested as a control in toxicity tests on normal cells of human skin and the results are shown in table 6.

TABLE 6

Toxicity to normal cells of human skin
					Concentration (mM)	Racemate treatment (% viability)	Alpha-isomer treatment (% viability)	Beta-isomer treatment (% viability)	Mitomycin treatment (% viability)
0.5	20	30	10	0
					0.05	50	40	20	0
0.005	80	70	50	30
					0.0005	100	100	100	60

TABLE 7

Toxicity to normal cells of human liver
				Concentration (mM)	Racemate treatment (% viability)	Alpha-isomer treatment (% viability)	Beta-isomer treatment (% viability)
0.5	30	20	10
				0.05	60	40	20
0.005	80	70	50
				0.0005	100	100	100

TABLE 8

Toxicity to normal cells of human intestine
				Concentration (mM)	Racemate treatment (% viability)	Alpha-isomer treatment (% viability)	Beta-isomer treatment (% viability)
0.5	40	40	30
				0.05	60	60	50
0.005	80	80	70
				0.0005	100	100	100

TABLE 9

Toxicity to normal cells of human lung
				Concentration (mM)	Racemate treatment (% viability)	Alpha-isomer treatment (% viability)	Beta-isomer treatment (% viability)
0.5	30	30	20
				0.05	60	50	40
0.005	80	80	70
				0.0005	100	100	100

Mitomycin, a typical anticancer drug, is currently available which exhibits strong toxicity to normal cells, as shown in table 6.

Claims (18)

1. A process for preparing a compound of formula (III):

wherein R is C₁-C₆An alkyl group, a carboxyl group,

comprising reacting a compound of formula (I) and a compound of formula (II) in the presence of a base:

wherein R is as defined above, X¹And X²Each independently is a halogen atom.

2. A process for preparing a compound of formula (III):

wherein R is C₁-C₆An alkyl group, a carboxyl group,

comprising reacting a compound of formula (I) and a compound of formula (II) in the presence of a base:

wherein R is as defined above, X¹And X²Each of which is independently a halogen atom,

to give a compound of general formula (VII):

wherein R is as defined above, and wherein,

the resulting compound (VII) is subsequently oxidized by an oxidizing agent.

3. The method of claim 2, wherein the oxidizing agent is manganese dioxide.

4. The process of any one of claims 1 to 3 wherein R is methyl.

5. The process of any one of claims 1 to 4 wherein X¹And X²Each is a bromine atom.

6. The process of any one of claims 1 to 5, wherein the base is DBU.

7. A process for preparing a compound of formula (IV):

wherein R is C₁-C₆Alkyl, the wavy line indicates a racemate,

the method comprises the following steps: reducing a compound of formula (III) with a reducing agent:

wherein R is as defined above.

8. A process for preparing a compound of formula (IV):

wherein R is C₁-C₆Alkyl, the wavy line indicates a racemate,

the method comprises the following steps: reacting a compound of formula (I) with a compound of formula (II) in the presence of a base:

wherein R is as defined above, X¹And X²Each of which is independently a halogen atom,

to obtain a compound of general formula (III):

wherein R is as defined above, and wherein,

the compound obtained subsequently from the reducing agent is of the general formula (III).

9. A process for preparing a compound of formula (IV):

wherein R is C₁-C₆Alkyl, the wavy line indicates a racemate,

the method comprises the following steps: reacting a compound of formula (I) with a compound of formula (II) in the presence of a base:

wherein R is as defined above, X¹And X²Each independently isA halogen atom(s),

to obtain a compound of general formula (VII):

wherein R is as defined above, and wherein,

the resulting compound (VII) is then oxidized by an oxidizing agent to give a compound of the general formula (III):

wherein R is as defined above, and wherein,

and reducing the resulting compound (III) with a reducing agent.

10. The process of any one of claims 7 to 9, wherein the reducing agent is NaBH₄。

11. The process of any one of claims 7 to 10, further comprising the step of resolving the compound of formula (IV):

wherein R is as defined above, the wavy line indicates a racemate,

to obtain the alpha-and beta-isomers of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione.

12. The process of any one of claims 1 to 6, 8 to 11, further comprising the step of reacting dimethylamine dissolved in a solvent with a compound of formula (V):

to give a compound of general formula (VI):

and a step of reacting the obtained compound of the general formula (VI) with 5 to 15% (w/w) of an aqueous acid solution to obtain a compound of the general formula (I).

13. The method of claim 12, wherein the aqueous acid solution is an aqueous hydrochloric acid solution.

14. A compound of the general formula (VII):

wherein R is C₁-C₆An alkyl group.

15. A racemic mixture of formula (IV):

wherein R is C₁-C₆Alkyl, the wavy line indicates racemate.

2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione, which has (+) -optical rotation in methanol (c0.25, 25 ℃).

17. An anticancer agent comprising a racemate of 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione as an active ingredient.

18. The anticancer agent according to claim 17, which comprises 2- (1-hydroxyethyl) -5-hydroxynaphtho [2, 3-b ] furan-4, 9-dione having a (+) -optical rotation in methanol (c0.25, 25 ℃) as an active ingredient.