JPH0533927B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Field of Application] The present invention relates to pharmaceuticals, particularly anticancer agents. [Prior Art] Under the circumstances where deaths due to cancer are increasing, various chemotherapy and immunotherapy are being performed in conjunction with surgical therapy. In chemotherapy, cytosin arabinoside, 5-fluorouracil, and the like are clinically used as antimetabolite antineoplastic agents, as they are effective for acute leukemia and the like. [Problems to be solved by the invention] However, the conventional anticancer drugs still do not have a fully satisfactory therapeutic effect and have various problems such as causing side effects. It has been demanded. [Means for solving the problem] From this perspective, the present inventors have conducted many years of research and found that a new nucleoside compound has antitumor effects against tumor cells such as L1210 leukemia or KP388 leukemia. found that it has an effect,
The present invention has been completed. The present invention relates to an anticancer agent containing 2'-deoxy-2'-methylidenecytidine or a pharmaceutically acceptable acid addition salt thereof as an active ingredient. 2′-deoxy-2′-methylidenecytidine has the formula The pharmaceutically acceptable acid addition salts include inorganic acid salts such as hydrochloride, hydrobromide, sulfate, and phosphate, or maleate, fumarate, tartrate, and succinate. Examples include organic acid salts such as salts, citrates, p-toluenesulfonates, and pamoates. A typical active ingredient compound of the present invention is 2'-deoxy-2'-methylidenecytidine hydrochloride 1/2 hydrate [melting point 148-155°C (decomposition point). Hereinafter, it is also referred to as compound A]. [Effect] Next, the antitumor action of the active ingredient compound of the present invention will be explained using pharmacological experiments. Example 1 Compound A was dissolved in dimethyl sulfoxide and then serially diluted with distilled water for injection to a final concentration of 20 times.
The test solution had a concentration of ml. 96 10μ of this test solution
Place the L1210 leukemia cell suspension at 1×10 ⁵ cells/ml in a hole tissue culture plate (Falcon No. 3072).
Add 190 μ and place in a carbon dioxide incubator.
Cultured at 37°C for 48 hours. As a control group, cells cultured in the same manner with only 10μ of distilled water for injection were used. After culturing, the number of viable cells was counted using an erythrocyte counter using the trypan blue staining method to determine the tumor growth inhibition rate, and the 50% tumor growth inhibition concentration (IC ₅₀ , μg/ml) of Compound A was 0.12 μg. /ml. The final concentration of dimethyl sulfoxide was 0.01% or less, and in the control group in which dimethyl sulfoxide alone was diluted in the same manner, no effect on tumor cell proliferation was observed at all. Example 2 10 ⁶ P388 leukemias (derived from the National Cancer Institute) were intraperitoneally transplanted into 3 female CDF ₁ mice (8 weeks cold) per group, and compound A was administered 5 times a day from the day after transplantation.
Intraperitoneal injection (IP) was administered for consecutive days. The median survival time (MST) was determined, and based on this, the survival rate (T/C, %) was determined using the following formula, and the results are summarized in Table 1. Life extension rate (T/C, %) = MST of treatment group/MST of control group ×
100

〔Example〕

The present invention will be explained below with reference to production examples of Compound A and formulation examples of the drug of the present invention, but the present invention is not limited thereto. Production example 2'-Deoxy-2'-methylidenecytidine hydrochloride production (1) Synthesis of 1-β-D-ribofuranosyl-4-ethoxy-2-pyrimidone 2',3',5'-tri-O- Acetyl uridine
Dissolve 3.35 g in 50 ml of chloroform, add 8.1 ml of thionyl chloride and 0.5 ml of dimethylformamide.
was added and refluxed for 6 hours and 30 minutes, and then dried under reduced pressure. The residue was dissolved in 20 ml of ethanol, 30 ml of 1N sodium ethoxide was added, and after refluxing for 2 hours, it was neutralized with 1N hydrochloric acid, the precipitated salt was filtered off, and the solution was concentrated to dryness. This was adsorbed on a silica gel column (4 x 31 cm), the fraction containing the target compound was extracted with 16% ethanol-chloroform, and the solvent was distilled off to obtain crude crystals of the target compound. Recrystallize this from ethanol to obtain 2.08 g of the target product (as 1/3 hydrate) (yield 84.2%)
I got it. Melting point: 136-137.5℃ Elemental analysis value: _{C11H16N2O6 ・ 1} / _3H2O Calculated value C: 46.97%, H: 6.09%, _N : 9.96%, O: 36.98% Actual value C: 46.91%, H: 6.02%, N: 9.98%, O: 37.09% (2) 1-(2-oxo-3,5-O-tetraisopropyldisiloxanyl-β-D-ribofuranosyl)-4-ethoxy -Synthesis of 2-pyrimidone 7.04 g of 1-β-D-ribofuranosyl-4-ethoxy-2-pyrrolidone was dissolved in 80 ml of pyridine, cooled on ice, and then 1,3-dichloro-1,
1,3,3-tetraisopropyldisiloxane
9.57 g was added, and the mixture was stirred and reacted at room temperature for 4 hours and 30 minutes. Ice water was added, the solvent was distilled off, and the residue was partitioned between chloroform and water. After drying the chloroform layer, the solvent was distilled off, the residue was adsorbed on a silica gel column (10 x 130 cm), and 40% ethyl acetate-hexane was added. Collect and concentrate the eluted portion, 3′,
5'-O-tetraisopropyldisiloxane
12.3g was obtained. Next, add 2.7 ml of oxalyl chloride to methylene chloride.
It was dissolved in 40 ml and cooled to -70°C. 4.8 ml of dimethyl sulfoxide dissolved in 20 ml of methylene chloride was added dropwise to this over 20 minutes under an argon stream, and the mixture was stirred for 30 minutes. To this, 12.3 g of the above 3',5'-O-tetraisopropyl disiloxane dissolved in 50 ml of methylene chloride was added dropwise, and -
After stirring at 70°C for 2 hours, triethylamine 20
ml and further stirred for 1 hour. The reaction solution was returned to room temperature, water was added and partitioned, the methylene chloride layer was separated, the solvent was distilled off, the residue was dissolved in ethyl acetate, and the mixture was partitioned with water. The ethyl acetate layer was concentrated to dryness, adsorbed on a silica gel column (5 x 28 cm), and fractions containing the target substance eluted with 20% ethyl acetate-hexane were collected, and after evaporation of the solvent,
Crystallized from hexane, 10.2g of target substance (yield
72.1%). Melting point _: 157.5-159℃ Elemental analysis value _{: C24H39N2O7Si2 Calculated} value C: 53.87%, H: 7.86%, _N : 5.46% Actual value C: 53.73%, H: 7.87%, N :5.57% (3) 1-(2-deoxy-2-methylidene-β-
Synthesis of D-ribofuranosyl)-4-ethoxy-2-pyrimidone 232 mg of potassium hydride was added to 2.4 ml of dimethyl sulfoxide under an argon stream, and the mixture was stirred at room temperature for 40 minutes. 2.2 g of methyltriphenylphosphonium bromide was dissolved in 8 ml of dimethyl sulfoxide, and the above mixture of potassium hydride and dimethyl sulfoxide was added dropwise thereto under ice cooling in an argon stream, followed by stirring for 10 minutes. Add to this the above 1-(2
-Oxo-3,5-O-tetraisopropyldisiloxanyl-β-D-ribofuranosyl)-4
A solution of 1.02 g of -ethoxy-2-pyrimidone crystals dissolved in 10 ml of dimethyl sulfoxide was added dropwise under an argon stream and stirred for 2 hours under ice cooling.
To this was added 10 ml of a 1N aqueous ammonium chloride solution, and then 50 ml of ethyl acetate and 40 ml of water were added for distribution. The organic layer was concentrated under reduced pressure, adsorbed on a silica gel column (2.4 x 30 cm), and eluted with a hexane-ethyl acetate mixed solvent to obtain a 2'-methylidened compound. 320 mg of the compound obtained above was dissolved in 10 ml of tetrahydrofuran, 1 ml of tetra n-butylammonium olide was added, and the mixture was stirred at room temperature for 10 minutes. After neutralization with acetic acid, apply a silica gel column (2.4
× 12cm), eluted with chloroform-ethanol, collected the eluate fractions containing the target compound, and deprotected the 2'-methylidene-4-O-ethyl compound.
155 mg (yield 30%) was obtained. Melting point: 157.5-159℃ Elemental analysis value _{: C12H16N2O5} Calculated value C: _53.72 %, H: _6.01 %, N: 10.44% Actual value C: 53.80%, H: 5.99%, N: 10.37 % (4) Synthesis of 2'-deoxy-2'-methylidenecytidine hydrochloride/hemihydrate 150 mg of 2'-methylidene-4-O-ethyl compound was dissolved in 10 ml of ammonia-saturated methanol solution under ice cooling. , and heated at 100°C for 2 days in a sealed tube.
After cooling, add 2 ml of 2N hydrochloric acid, concentrate, and crystallize from ethanol-water to obtain the title compound 125.
mg (yield 81.7%). Melting point: 148-155℃ (decomposition point) Elemental analysis value: _C10H13N3O4・_HCl・1/ _2H2O Calculated value C: _42.14 %, H: 5.31%, _N : 14.74% Actual value C : 42.25%, H: 5.26%, N: 14.69% Formulation Example 1 Tablet consisting of Compound A 50.0mg, finely powdered cellulose 25.0mg, lactose 49.5mg, starch 40.0mg, talc 5.0mg, and magnesium stearate 0.5mg. Sugar-coated tablets or film-coated tablets can be made by sugar-coating or film-coating, if desired. Production Example 2 A capsule prepared by filling a capsule with the following composition: Compound A 50.0mg Lactose 50.0mg Starch 15.0mg Talc 5.0mg. Formulation Example 3 Fine granules consisting of 10% Compound A, 80% lactose, and 10% starch. Formulation Example 4 Granules consisting of 10% Compound A, 55% lactose, 20% finely powdered cellulose, and 15% starch. Production Example 5 Dissolve 50.0 mg of Compound A and 100.0 mg of glucose in purified water to make an injection solution with a total volume of 2 ml. Production Example 6 A suppository consisting of Compound A 100mg Witepsol○R H15 950mg Witepsol○R E75 950mg. However, Uitepsol is a registered trademark owned by Witten GmbH (West Germany). Formulation example 7 Compound A 2g Ethyl parahydroxybenzoate 0.025g Propyl parahydroxybenzoate 0.015g Sodium lauryl sulfate 1.5g Propylene glycol 12.0g Stearyl alcohol 22.0g White petrolatum 25.0g The composition was dissolved in purified water to give a total amount of 100.0g. Use as ointment.

Claims (1)

[Claims] 1. An anticancer agent containing 2'-deoxy-2'-methylidenecytidine or a pharmaceutically acceptable acid addition salt thereof as an active ingredient.