JPS63135399A - Cyclic amp derivative - Google Patents

Abstract

NEW MATERIAL:A compound expressed by formula I (R is lower alkyl). EXAMPLE:Adenosine 3',5'-cyclic methyl phosphonate. USE:A carcinostatic agent and cyclic adenosine monophosphate (AMP) phosphodiesterase inhibitor having excellent permeability to cells. PREPARATION:A 2'-tetrahydropyranyladenosine expressed by formula II is reacted with an alkyl 0,0-bis-(1-benzotriazolyl)phosphonate expressed by formula III. N-Methylimidazole is added and reacted with the resultant reaction product to afford a compound expressed by formula IV, which is then hydrolyzed with an acid.

Description

Detailed Description of the Invention [Industrial Application Field] The present invention provides a novel compound, cyclic AMP@conductor, represented by the general formula (1) (wherein R represents a lower alkyl group). The present invention also relates to a method for producing the same, an anticancer agent containing the same, and a cyclic AMP phosphodiesterase inhibitor.

[Prior art]

Cyclic hormones are known as intracellular transmitters of hormones, and are involved in metabolic regulation, protein and nucleic acid synthesis regulation, secretion,
It is said to be a substance related to neurotransmission, differentiation, and canceration. However, it is also known that cyclic AMP has poor cell permeability and exhibits significantly low effects when administered externally to a living body. Also, cyclic AM
It is known that P is rapidly degraded by the action of cyclic AMP phosphogesterase within the M5 vacuole. In order to improve the cell permeability of this cyclic AMP, dibutyryl cyclic compound was synthesized, but it quickly becomes cyclic AMP in cells and is therefore degraded by the action of cyclic AMP phosphogesterase. It has the disadvantage of being

[Purpose of the invention]

An object of the present invention is to provide a novel cyclic AMP derivative that has good cell permeability and is not degraded by the action of cyclic AMP phosphogesterase.

[Disclosure of the invention]

As a result of intensive research, the present inventor discovered the general formula (wherein R
represents a lower alkyl group. ) has anticancer activity and is not only not degraded by the action of cyclic AMP phosphogesterase, but also has inhibitory activity of cyclic phosphogesterase (usually cyclic AMP phosphogesterase). It has been found that compounds having inhibitory activity against phosphogesterase are also expected to have diuretic effects, cardiotonic effects, stimulant effects, blood sugar increasing effects, etc.).

The present invention is based on this knowledge.

Accordingly, the present invention provides novel cyclic AMP derivatives useful as anticancer agents and cyclic AMP phosphogesterase inhibitors.

The invention also provides a novel method for manufacturing cyclic AMPts conductors (1).

Furthermore, the present invention provides cyclic AMP derivatives (1)
The object of the present invention is to provide an anticancer agent and a cyclic AMP phosphogesterase inhibitor, which contain as an active ingredient.

The cyclic AMP derivative (1) according to the present invention can be synthesized, for example, according to the following reaction formula. That is, 2'-tetrahydropyranyl adenosine represented by formula (II) and a-carboxylated 02 represented by general formula (In)
By reacting o-bis-(1-benzotriazolyl)phosphonate and then adding N-methylimidazole to this reaction product and reacting, a compound represented by the general formula (g) is produced, and then , produced by acid hydrolysis.

(In the formula, R represents a lower alkyl group.) The compound of formula (II) used as a raw material in this method is, for example, a commercially available N6-penzoyl-5'-0-(4,4'-dimethoxytrityl)- It is obtained by first treating 2'-0-tetrahydropyranyladenosine with ammonia and then with acetic acid to remove the two protecting groups on the base moiety and the 5' hydroxyl group. In addition, 3',5'-0-(
Tetrainpropyldisiloxane 1,3-diyl)-adenosine and then treatment with dihydrobyran and p-toluenesulfonic acid to give 2'-〇-tetrahydropyranyl31. It can also be obtained by synthesizing s/-0-(tetraisopropyldisiloxane 1,3-diyl)-adenosine and further treating this with tetra-n-butylammonium fluoride.

The compound of formula (2), which is one of the raw materials, was prepared by the method of Marugg et al. (Nucleic A
c1d Res, 14, 2171, 1986).

The reaction between the compound of formula (It) and the compound of formula
This is done by adding methylimidazole and leaving it for 18 to 30 hours.

Further, hydrolysis of the compound of formula (I) is achieved by leaving it at room temperature for 40 to 50 hours with a strong alcohol such as hydrochloric acid.

Examples of the production of compounds according to the present invention are listed below.

Although the following raw material production examples and examples are for explaining the present invention, the present invention is not limited by these production examples and examples.

Raw material production example (1) Production of 2'-tetrahydropyranyl adenosine N6-penzoi-5'-0-(4,4'-dimethoxytrityl), -27-o-tetrahydropyranyl adenosine (Dojinsha g) 1 8 WIt of pyridine was added to .19 and dissolved therein, and 10 ml of aqueous ammonia was further added thereto, followed by stirring at room temperature for 6 hours. After the reaction solution was concentrated to dryness, 80% acetic acid aqueous solution was added thereto for 15 minutes, and the mixture was left at room temperature for 20 minutes. After concentrating the reaction solution to dryness, it was subjected to silica gel column chromatography to obtain 4.8 t of crude crystals.
(yield 94%).

(2) Production of 2'-tetrahydropyranyladenosine (alternative method) Add anhydrous pyridine 40- to adenosine 2.67f,
Furthermore, 3,46 t of 1,3-dichloro-1,1,3,3-tetrinepropyldisiloxane (manufactured by Tokyo Kasei Co., Ltd.) was added, stirred at room temperature for 3 hours, and the reaction solution was subjected to silica gel column chromatography. 6151-o-(tetraisopropyldisiloxane 1,3-diyl)-adenosine 4.8F (yield 94%) was obtained. This 3', 5'
-0-(tetraisopropyldisiloxane 1,6-diyl)-adenosine 1.018f, dihydrobilane (
(manufactured by Tokyo Kasei Co., Ltd.) 2,018g and p-toluenesulfone! The mixture was stirred for 20 minutes at room temperature in anhydrous dioxane 10, and the reaction mixture was separated using a water-chloroform system to obtain 2/-o-tetrahydropyranyl-3',
5'-0-(tetraisopropyldisiloxane 1.5
-';yl)-adenosine 1.1t (yield 95%) was obtained. This 2'-〇-tetrahydropyranyl-3', 5'
-0-(tetrainpropyldisiloxane 1,5-diyl)-adenosyl/tetra-n-butylammonium fluoride (adjusted to a concentration of 1 mol/L in 900 μL)
A tetrahydrofuran solution (manufactured by Tokyo Kasei Co., Ltd.) was added thereto, stirred at room temperature for 1 hour, and the reaction solution was subjected to silica gel column chromatography to obtain 505 g of crude crystals (yield: 95 g).
h) was obtained.

(3) Production of methyl-〇,0-bis-(1-benzotriazole) phosphonate 1.16 ml of 1-hydroxybenzotriazole and 0.84 ml of anhydrous pyridine were mixed with 27.41 tL of anhydrous dioxane.
It was dissolved in t. This dioxane solution was mixed with anhydrous dioxane 8
．． The mixture was added dropwise to 559 g of methylphosphonic acid dichloride (manufactured by Aldrich Co., Ltd.) dissolved in A.3-3, and reacted at room temperature for 2 hours. The reaction solution was filtered under reduced pressure under anhydrous conditions.
．． A 11M dioxane solution was obtained.

Example 1 The 2'-tetrahydropyranyladenosine 367 prepared in the above raw material production example (1) was prepared in the same manner as in (3). Adding a 0.11 M dioxane solution of methyl-○,○-bis-(1-benzotriazole) phosphonate 18-
It was left at room temperature for 1 hour. Then, 0.42-N-methylimidazole was added and reacted at room temperature for 5 hours. Add dichloromethane-dioxane (9:1) mixture IC to the reaction solution.
1d and separated with 10 sil of IM-triethylammonium acetate to remove the dichloromethane layer.
After washing three times with water, the dichloromethane layer was concentrated, and the concentrated solution was subjected to silica gel column chromatography to obtain crude crystals of 2'-tetrahydropyranyl adenosine S/, S/-cyclic methylphosphonate (80 to 9) (harvested). Rate 18.
5%). To this was added 5-, a solution of 0.01N hydrochloric acid and dioxane-water (9:1), and the mixture was left to stand at room temperature for 45 hours. This reaction solution was subjected to silica gel column chromatography to give crystals of adenosine 3',5'-cyclic methylphosphonate 68119 (yield: 99%).
%) was obtained.

FD-MASS: (M/Z) 528 (M+
1)'HNMR (CD30D) δ: 8.487 (
s, IH) 8.443 (s, IH) 6.154 (s, IH) t756 (d, J = 18.31Hz, 3H) Stone λm
a: c: 258.9 nm (g = 11,500) The results of pharmacological effects testing of the representative compound of the cyclic AMP derivative (1) according to the present invention obtained in this manner are as follows. .

(1) Anticancer effect of DT [cells (Proc, Nat, Acad, S), which are transformed cancer cells of 3T3 cells, which are mouse fibroblasts.
ci.

USA 80,562.1983), T24 cells derived from human bladder cancer (Nature 298,343.198
2), human melanoma-derived A375 a cysts (T, N, C,
I 72, 913.1984) as well as Burkitt's lymphoma-derived Daudi cells (Cancer Ftes.

28.1300, 1958), and each was placed in a 96-well multiplate with one hole per hole.
After 3 hours, the test substance was dissolved in a serum-free medium and added in an amount of 10 μt at a final concentration of 0.6 to 1.3 mM per well. After culturing this in a CO2 incubator at 67°C for 48 hours, cell proliferation was performed using the MTT method (J, Immunol.

Method 65, 55.1981 J-Immun
ol, Method 70.257.1984). The results are shown in Table 1.

When similar experiments were conducted using 5T5 cells, which are normal cells, none of the compounds showed cytotoxicity.

(2) Cyclic AMP phosphogesterase inhibitory effect Using bovine heart cyclic AMP phosphogesterase (manufactured by Boehringer Mannheim), the method of Pichard et al. (J, Biol, Chem, 251, 5)
The inhibitory effect was investigated according to the cyclic AMP phosphodiesterer heptaactivity assay method according to 726.1976). i.e. 5mM MgSO4, 50pM C as final concentration
Add 10,000 d of 5H-cyclic AMP2Q to 100 pL of 40mM) J-HCl buffer prepared to give aCl2.
prp, 1 μM cyclic AMP, cyclic A
After adding 0.2μ2 of MP phosphogesterase and various concentrations of test substances and reacting at 60°C for 20 minutes,
Inactivate the enzyme by treating at 0°C for 6 minutes, return to room temperature,
Add 0μ2 of snake venom (manufactured by Sigma), react for 10 minutes at 30°C, cyclically decompose 5'-A, IJP produced by the action of AP phosphogesterase into adenosine, and add AGIX2 resin (manufactured by Bioland). ) of 1:3
After adding water suspension 1- to stop the decomposition reaction, the mixture was thoroughly mixed and centrifuged (5000 rpm) for 5 minutes. Take 100 μt of this supernatant, measure its emission count with a liquid scintillation counter, and measure the 50% inhibitory concentration (IC5
0) was calculated. The results are shown in Table 2.

Table 2 Also, the compound produced in Example 1 itself was not degraded at all by cyclic AMP phosphogesterase.

Compound (1) according to the present invention can be used as an anticancer agent or as a cyclic AMP phosphogesterase inhibitor, and can be administered by either oral or parenteral routes. As a dosage form,
Examples of oral preparations include tablets, capsules, powders, buccal tablets, etc.; examples of parenteral preparations include injections. The additives such as excipients, disintegrants, lubricants, dyes, diluents, etc. used in these V4 fissures are those normally used in medicine, and all can be used. must not have an adverse effect on compound (1).

Patent applicant Akira Kaji Agent: Patent attorney Takashi Minami.

,1

Claims (1)

[Claims] (1) A cyclic AMP derivative represented by the general formula (I) ▲There are numerical formulas, chemical formulas, tables, etc.▼(I) (In the formula, R represents a lower alkyl group.) (2) Formula (II) ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ (II) 2'-tetrahydropyranyl adenosine represented by and general formula (III) ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ (III) (wherein, R represents a lower alkyl group) is reacted with an alkyl O,O-bis-(1-benzotriazolyl)phosphonate represented by
By adding methylimidazole and causing a reaction, a compound represented by the general formula (IV) ▲ Numerical formula, chemical formula, table, etc. ▼ (IV) (in the formula, R represents a lower alkyl group) is produced, and this Cyclic AMP represented by the general formula (I) ▲Mathematical formula, chemical formula, table, etc.▼(I) (In the formula, R represents a lower alkyl group.) Method for producing derivatives. (6) General formula (I) ▲There are mathematical formulas, chemical formulas, tables, etc.▼(I) (In the formula, R represents a lower alkyl group.) An anticancer agent containing a cyclic AMP derivative represented by the following as an active ingredient . (4) General formula (I) ▲There are mathematical formulas, chemical formulas, tables, etc.▼(I) (In the formula, R represents a lower alkyl group) A cyclic AMP derivative containing as an active ingredient Click AMP phosphodiesterase inhibitor.