JPH0613546B2 - Nucleic acid derivative and anticancer agent containing the derivative as an active ingredient - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial field of use] The present invention is a nucleic acid derivative having a specific anticancer activity,
The present invention relates to a compound expected to be used as an anticancer agent.

[Prior Art and Problems] Conventionally, many researchers have studied anticancer agents consisting of nucleic acid derivatives, and especially for fluorinated pyrimidine derivatives, 5-fluorouracil and tegafur have been subjected to strict examination for their usefulness. , Anticancer compounds such as doxyfluridine are clinically used.

However, from the viewpoint of side effects and resistance acquisition, development of a more effective and safe anticancer agent has been desired.

[Means for Solving the Problems] In order to obtain an anticancer agent having an excellent antitumor effect, the present inventor has synthesized various derivatives using 5-fluorouracil as a starting material, and has conducted extensive studies on the antitumor effect. The present invention has been completed by finding a compound having an excellent antitumor effect. That is, the present invention is [In the formula, R ₁ represents a lower alkyl group having 1 to 3 carbon atoms, and R ₂ represents a pharmacologically acceptable salt such as H, Na or (Et) ₃ NH. ] The nucleic acid derivative represented by
It is called the compound of the present invention. ) And a derivative thereof as an active ingredient.

The compound of the present invention can be obtained, for example, by synthesizing as follows.

It can be obtained by using commercially available 5-fluorouridine as a raw material, protecting the hydroxyl groups at the 2'-position and the 3'-position, reacting the hydroxyl group at the 5'-position with methyl phosphoric acid to perform methyl phosphorylation, and further removing the protecting group.

In protecting the hydroxyl group, any method may be used as long as it does not affect the 5'-position hydroxyl group, and it is particularly preferable to react with acetone in the presence of p-toluenesulfonic acid.

Methyl phosphorylation can be achieved by a usual method, and the reaction is completed in 10 to 48 hours at room temperature using N, N'-dicyclohexylcarbodiimide as a condensing agent in a solvent such as anhydrous pyridine.

Deprotection depends on the method of protecting the hydroxyl group described above, but when the reaction with acetone is performed in the presence of p-toluenesulfonic acid, acetic acid is added and heating is performed at 100 ° C.

The compound of the present invention is obtained as described above, and the compound of the present invention also includes pharmacologically acceptable salts such as sodium salt and potassium salt.

Next, the fact that the compound of the present invention exhibits a carcinostatic action will be described with reference to experimental examples.

Experimental Example 1 HeLa tumor cells, human T24 tumor cells and mouse L1210 tumor cells were dispersed in Eagle's minimum medium containing 10% fetal bovine serum in advance, and 3 × 10 ³ cells (100 μl) were plated per well on a coaster 3596 plate, After 3 hours, the compound obtained in Example 2 described later and 5-fluorouracil (manufactured by Sigma) were dissolved in serum-free Eagle's minimal medium at various concentrations and added to each tumor cell. After culturing these tumor cells for 2 days, MTT [3-, which has the property of reacting specifically with living tumor cells and developing color
(4,5-Dimethylthiazol-2-yl) -2,5-
Diphenyltetrazolium bromide] was added, the reaction was stopped 4 hours later, the absorbance was measured, and the tumor cell growth inhibition rate was calculated. The 50% growth inhibitory concentration for each tumor cell is shown below.

Experimental Example 2 1 × 10 ⁵ mouse leukemia L1210 cells were intraperitoneally transplanted into BDF1 male mice (6 weeks old), and from the next day, 0.2 ml of physiological saline solution of the compound obtained in Example 2 and 5-fluorouracil was added. 5 mg / kg and 10 mg / kg were administered intraperitoneally 5 times each day. As a control, physiological saline alone was intraperitoneally administered, and the survival rate was calculated from the survival days of the mice.

As a result, the life extension rate of 5-fluorouracil was 71%, and the life extension rate of the compound of the present invention was 90%.

From these results, the excellent antitumor activity of the compound of the present invention was confirmed.

Next, the dose and formulation of the compound of the present invention will be explained.

No toxicity was observed when the compound of the present invention was administered to mice at 60 mg / kg for 10 days.

The compound of the present invention can be administered to animals and humans as it is or together with a conventional pharmaceutical carrier. The administration form is not particularly limited and may be appropriately selected and used as needed, and examples thereof include oral agents such as tablets, capsules, granules, fine granules and powders, parenteral agents such as injections and suppositories. .

In order to exert a desired effect as an oral agent, it varies depending on the age, body weight and degree of disease of the patient, but usually 50 to 300 mg of the compound of the present invention is divided into several times a day in adults. It seems that it is appropriate to take.

In the present invention, oral preparations such as tablets, capsules and granules are produced according to a conventional method using starch, lactose, sucrose, mannitol, carboxymethyl cellulose, corn starch, inorganic salts and the like.

In addition to the above-mentioned excipients, a binder, a disintegrant, a surfactant, a lubricant, a fluidity promoter, a flavoring agent, a coloring agent, a fragrance and the like can be appropriately used in this type of formulation. Specific examples of each are as follows.

[Binder] Starch, dextrin, gum arabic powder, gelatin,
Fudroxypropyl starch, methyl cellulose, sodium carboxymethyl cellulose, hydroxypropyl cellulose, crystalline cellulose, ethyl cellulose, polyvinylpyrrolidone, macrogol.

[Disintegrant] Starch, hydroxypropyl starch, sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, carboxymethyl cellulose, low-substituted hydroxypropyl cellulose.

[Surfactant] Sodium lauryl sulfate, soybean lecithin, sucrose fatty acid ester, polysorbate 80.

[Lubricant] Talc, waxes, hydrogenated vegetable oil, sucrose fatty acid ester, magnesium stearate, calcium stearate, aluminum stearate, polyethylene glycol.

[Fluidity promoter] Light anhydrous silicic acid, dried aluminum hydroxide gel, synthetic aluminum silicate, magnesium silicate.

The compound of the present invention can also be administered as a suspension, emulsion, syrup, elixir,
These various dosage forms may contain a flavoring agent and a coloring agent.

In order to exert a desired effect as a parenteral agent, it depends on the age, body weight, and degree of disease of the patient, but usually the adult compound is intravenously administered at a dose of 0.5 to 100 mg / day as an adult. , Intravenous drip, subcutaneous injection, and intramuscular injection may be appropriate.

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

Other parenteral agents include external preparations, coating agents such as ointments, suppositories for rectal administration, etc., and they are manufactured by a conventional method.

Hereinafter, examples of producing the compounds and salts of the present invention will be shown and described in more detail, but the present invention is not limited thereto.

Example 1 400 mg of 5-fluorouridine (manufactured by Sigma) and 40 mg of p-toluenesulfonic acid were dissolved in 20 ml of anhydrous acetone, and the mixture was stirred at room temperature for 3 hours. The resulting reaction solution was cooled with ice,
Baking soda was added and left overnight. Excess acetone was added to this, the precipitated salt was filtered off, the obtained residue was washed with acetone, the washing solution and the filtrate were mixed, and concentrated under reduced pressure to give a chloroform-soluble part of the obtained residue 2. 460 mg of ', 3'-isopropylidene-5-fluorouridine were obtained.

This 2 ', 3'-isopropylidene-5-fluorouridine (460 mg), methylphosphoric acid (336 mg) and N, N'-dicyclohexylcarbodiimide (3.15 g) were dissolved in anhydrous pyridine (5 ml), and the mixture was stirred at room temperature for 2 days. The precipitated insoluble matter was filtered off, washed with water, the filtrate was extracted with petroleum ether, and the aqueous layer was concentrated under reduced pressure to give crude 2 ', 3'-isopropylidene-
1.1 g of 5-fluorouridine-5'-O-methyl monophosphonate was obtained.

1.1 g of this crude 2 ', 3'-isopropylidene-5-fluorouridine-5'-O-methylmonophosphonate was dissolved in 10% acetic acid and stirred at 100 ° C for 1.5 hours.
The resulting reaction solution was diluted with water, extracted with ether,
The aqueous layer of the obtained petroleum ether extract was concentrated under reduced pressure. The obtained residue was subjected to anion exchange chromatography [developing solvent: 0.05 to 0.35 M, triethylamine carbonate buffer solution], and the eluted UV absorbing portion at 260 nm was collected to give 5-fluorouridine-5'-. 600 mg of O-methyl monophosphonate triethylamine salt was obtained.

Example 2 600 mg of the 5-fluorouridine-5'-O-methylmonophosphonate triethylamine salt obtained in Example 1 was dissolved in 21 ml of methanol, and 2.4 g of sodium perchlorate was dissolved in 17 ml of acetone while stirring. And add
The deposited precipitate was centrifuged to obtain 433 mg of crystals. Due to the physicochemical properties shown below, 5-fluorouridine-5 '
It was determined to be -O-methyl monophosphonate sodium salt.

Mass spectrum m / e: 385 (M ⁺ , + Na) Infrared absorption spectrum ▲ ν ^KBr _max ▼ cm ⁻¹ : 3444, 1710, 1262, 1180, 1120, 1058 Ultraviolet absorption spectrum λ _max nm (ε): 0.495 ( 8960) Proton nuclear magnetic resonance spectrum (δ ppm in CDCl ₃ ): 1.33 (3H, d, J = 16.4 Hz), 4.06 to 4.30 (5H, m), 5.92 to 5.97 ( 1H, m), 8.12 (1H, d, J = 6.6 Hz) Example 3 Crystalline cellulose 39.5 g Magnesium stearate 0.5 g Carboxymethyl cellulose calcium 5 g Compound obtained in Example 2 5 g Total 50 g According to the prescription, a part of and is uniformly mixed, compression-molded, crushed, and the remaining amount of is added and mixed, and compression-molded with a tableting machine to give a tablet of 200 mg. It was obtained.

Each tablet contains 20 mg of the compound obtained in Example 2, and 3 to 15 tablets for adults are to be taken in several divided doses per day.

Example 4 Distilled water for injection Appropriate amount Glucose 200 mg Compound obtained in Example 2 3 mg Total amount 5 ml After dissolving and in distilled water for injection, the mixture was poured into an ampoule of 5 ml and autoclaved at 121 ° C. for 15 minutes. Then, an injection was obtained.

Claims (2)

[Claims] 1. A formula [In the formula, R ₁ represents a lower alkyl group having 1 to 3 carbon atoms, and R ₂ represents a pharmacologically acceptable salt such as H, Na or (Et) ₃ NH. ] The nucleic acid derivative represented by these. 2. A formula [In the formula, R ₁ represents a lower alkyl group having 1 to 3 carbon atoms, and R ₂ represents a pharmacologically acceptable salt such as H, Na or (Et) ₃ NH. ] An anticancer agent comprising a nucleic acid derivative represented by the following as an active ingredient.