CS199666B2 - Process for preparing 9-substituted purines - Google Patents

Description

The present invention relates to 9-substituted purines and pharmaceutically acceptable salts thereof, and processes for their preparation. In particular, the present invention relates to esters of 9- (2-hydroxyethoxymethyl) guanine a derivatives

2,6-diaminopurine and pharmaceutically acceptable salts thereof.

It has now been found that the 9-substituted purines of formula I

R *

CM 10 O C 6 O ⁶ OC.R ¹⁰ o

ω kde

R ¹ is hydroxyl or amino,

R ² is hydrogen, unbranched or branched alkyl having 2-16 carbon atoms, have antiviral activity against various species of DNA and RNA viruses in vitro and against DNA viruses in vivo. In particular, the compounds are effective against herpes, vaccinia and rhino viruses, wherein the herpes viruses include simplex, zoster and varicella in mammals that cause diseases such as herpes keratitis in rabbits and herpes encephalitis in mice.

According to the present invention, compounds of formula I are prepared wherein R ¹ and R ² are as defined above, or salts thereof, especially in the form of pharmaceutically acceptable salts.

Compounds of formula I wherein R ¹ is hydroxyl or a salt thereof are preferred as well as compounds wherein R ² is a hydrogen atom or a branched or unbranched alkyl group having 2 to 8 carbon atoms, and most preferably compounds having 2 to 4 carbon atoms .

The most preferred compounds are

9- (2-formyloxyethoxymethyl) guanine,

9- (2-propionyloxyethoxymethyl) guanine,

9- [2- (2,2-dimethylpropionyloxy) ethoxymethyl] guanine a

2-amino-9- (2-formyloxyethoxymethyl) adenine, especially for their remarkable antiviral activity against Herpes.

Salts which are particularly suitable for therapeutic use are the salts of pharmaceutically acceptable organic acids such as lactic, acetic, malonic or p-toluenesulfonic acid, as well as salts of pharmaceutically acceptable mineral acids such as hydrochloric acid or sulfuric acid. When R ¹ is hydroxyl, pharmaceutically acceptable alkali metal salts, preferably sodium salts, may be used. Other salts may also be prepared and then converted into salts directly usable for therapeutic purposes.

The present invention relates to a process for the preparation of a 9-substituted purine of formula I as defined above, characterized in that the alcohol of formula V

C / ^ O '(V) where

R ¹ has the abovementioned meanings, is esterified by reaction with an appropriate acylating agent to introduce the group R ^a, and if the product jo base, converting optionally the product into a salt with an acid or an alkali metal salt or where the product is a salt of a compound of formula I is converted optionally a salt or other salt thereof.

In the process of the present invention, the alcohol of formula V is reacted with a suitable acylating agent such as the corresponding carboxylic acid, anhydride, acyl halide, mixed carboxylic acid and carbonic anhydride or carboxylic acid ester. Compounds of formula V can be prepared as described in German DOS 2,539,963.

The method applies to intermediates that can be prepared from simply substituted purines. These purines are, of course, readily available by commonly known literature and textbook procedures such as "Heterocyclic Compound - Fused Pyrimidines Part II. Purines D. J. Brown (1971) by Wiley-Interscience".

The biological effect of the compounds of the present invention is demonstrated by the following test.

The plate inhibition assay was performed on VERO cells in which a confluent monolayer of cells was infected with sufficient virus to form a uniform coating on the plate. After addition of the solid overcoat layer, a filter paper disc impregnated with a test compound solution (50 µg / disc) is placed in the center of the plate on the solid cover layer. Cultures are incubated for 96-120 hours at 37 ^Q C in a 5% carbon dioxide. After this time, the cells are fixed with 10% formalin and then stained with 0.05% methyl red. The results are summarized in Table 1.

Table 1

Compound Effect on herpes

9- (2-Formyloxyethoxymethyl) guanine - η 2 -amino-9- (2-hydroxyethoxymethyl) adenine +

9- (2-hexanoyloxyethoxymethyl) guanine - (9- (2-propionyloxyethoxymethyl) guanine-η 9 - [2- (2,2-dimethylpropionyloxy) ethoxymethyl] guanine +

9- (2-palmitoyloxy-ethoxymethyl) guanine +

9- (2-butyryloxyethoxymethyl) guanine +

-j— | - zone greater than 25 mm in diameter -f- zone 5 to 25 mm + zone 0 to 5 mm - negative

According to another feature, the present invention relates to a pharmaceutical composition comprising a compound of formula I as defined above or a pharmaceutically acceptable salt thereof together with a pharmaceutically acceptable carrier. According to a feature, the pharmaceutical composition comprises a compound of Formula I in an effective unit dosage form.

As used above, the term "effective unit dose" refers to a predetermined antiviral amount sufficient to counteract viruses in vivo. Pharmaceutically acceptable carriers are substances useful for drug applications and may be solid, liquid or gaseous and which are otherwise inert and pharmaceutically suitable and compatible with the active ingredients.

The pharmaceutical compositions may be administered parenterally, orally, as suppositories or pessaries, topically applied as ointments, creams, aerosols, powders, or as drops in the eyes or nose, and the like. depending on whether the preparation is to be used for the treatment of internal or external viral infection.

For internal infections, the compositions may be administered orally or parenterally in doses, calculated on the free base, of from about 0.1 to 250 milligrams per kg, preferably 1 to 50 mg / kg of mammalian body weight, and when used in unit dosage forms in humans. they may be administered several times a day in an amount of from 1 to 250 mg per unit dose.

For oral administration, the fine powders or granules may contain diluents, dispersants and / or surfactants and may be drinkable as a gulp, in water or syrup, may be in solid form in capsules or wafers or in the form of non-aqueous solutions or suspensions. in which suspending agents may be present, in the form of tablets, which may contain binders and lubricants, or in the form of suspensions in water or syrup. If desired or necessary, flavoring, preservative, suspending, thickening or emulsifying agents may also be present. Tablets and granules are most preferred and may be coated.

For parenteral administration or for administration in the form of drops, for example for ocular infections, the compounds may be present in the form of an aqueous solution at a concentration of from about 0.1 to 10%, preferably from 0.1 to 1%, most preferably from 0.2% / obj. The solution may contain antioxidants, buffers and the like.

Alternatively, for infection of the eye or other external tissue such as the mouth and skin, the compositions are preferably applied to the infected part of the patient's body in the form of an ointment, for example in a water-soluble ointment or cream base, for example as an oil in an aqueous cream base. 0.1 to 10%, preferably from 0.3 to 3%, most preferably 1% w / v.

According to a further feature, the present invention relates to a method of treating viral infections in mammals, characterized in that an effective unit dose as defined above is administered to a compound of formula I or a pharmaceutically acceptable salt thereof. The administration is preferably topical or oral or parenteral.

The present invention is illustrated by the following examples.

Example 1

A solution of 9- (2-hydroxyethoxymethyl jguanine (4.73 g) prepared according to the procedures described in DOS 2,539,983 in 97% formic acid (24 mL) was stirred overnight at room temperature. The amber solution was diluted with about 200 mL of anhydrous ether and The resulting white precipitate was filtered, dried and recrystallized from anhydrous dimethylformamide to give 9- (2-formyloxyethoxymethyl) guanine (3.6 grams, mp 225-275 ° C).

Example 2

A solution of 2-amino-9- (2-hydroxyethoxymethyl) adenine (0.5 g) prepared according to the procedure of DOS 2,539,963 in 97% formic acid (2.5 ml) was stirred in an ice bath for three hours and then at room temperature overnight. Anhydrous ether (80 mL) was added and the mixture was cooled, filtered and dissolved in hot acetonitrile (125 mL), the remaining solids were filtered off and the filtrate was applied to a column containing silica gel (14 g) in acetonitrile.

The column was eluted with anhydrous acetone. The eluate was evaporated and the remaining solids were recrystallized from acetonitrile to give 2-amino-9- (2-formyloxyethoxymethyl) adenine (93 mg, mp 238-240 ° C).

Example 3

Preparation 9- ^! (2-hexanoyloxyethoxymethyl) guanine

9- (2-hydroxyethoxymethyl) guanine (1.0 g) was prepared as described in DOS 2,539,963 and dissolved in anhydrous dimethylformamide (70 mL) by heating on a steam bath. The solution was cooled to room temperature and anhydrous pyridine (10 mL) and hexanoic anhydride (9.4 g) were added. After stirring at room temperature for 4 days, the reaction was complete as judged by thin layer chromatography in 10% methanol in chloroform.

The yellow reaction solution was diluted with ethyl acetate to 800 mL and cooled for several days. The fine white powder which precipitated was filtered off, dried and recrystallized from anhydrous acetonitrile. 400 mg (28%) of white granules of m.p. 221-223 ° C are obtained.

Example 4

Preparation of 9- (2-tridecanoyloxyethoxymethyl) guanine

9- (2-Hydroxyethoxymethyl) guan (300 mg) prepared according to German DOS 2 539 963 was heated in anhydrous dimethylformamide (20 ml) and anhydrous pyridine (30 ml) on a steam bath until dissolved. The solution was cooled to room temperature and tridecanoyl chloride (0.93 g) was added. The solution was stirred at room temperature until thin layer chromatography showed that the reaction was complete and the starting material disappeared (18 hours).

The solution was evaporated and the residual yellow oil crystallized from acetonitrile to give a cream-yellow solid (375 mg). NMR analysis of the crude product indicated the presence of several compounds. The clear mixture is then purified by column chromatography.

Example 5

9- (2-Propionyloxyethoxymethyl) guanine

A mixture of 9- (2-hydroxyethoxymethyl) guanine (1.0 g) and anhydrous dimethylformamide (50 ml) was heated on a steam bath until most of the solid dissolved. The solution was then cooled to room temperature. Anhydrous pyridine (10 mL) and propionic anhydride (2.9 mL) were added and the mixture was stirred at room temperature overnight. Additional propionic anhydride (1.0 mL) was added and the mixture stirred for an additional 18 hours. The reaction mixture was diluted with ethyl acetate, cooled and the resulting solid was filtered off. Crystallization from dimethylformamide gave 9- (2-propionyloxyethoxymethyl) guanine (0.9 g) m.p. 223 DEG-226 DEG C .;

Example 6

9- (2- (2,2-Dimethylpropionyloxy) ethoxymethyl] guanine

A mixture of 9- (2-hydroxyethoxymethyl) guanine (2.46 g), anhydrous pyridine (400 mL) and pivalic anhydride (6.5 mL) was heated on a steam bath for a total of 33 days. Additional pyridine (150 mL) was added on day 11 and dimethylformamide (50 mL) was added on day 27. The volatiles were removed under reduced pressure and the remaining solid was triturated with ethyl acetate. The insoluble solid was filtered off and dissolved in a mixture of methanol and acetone. Silica gel (3 g) was added and the solvent was evaporated. The residue was applied to a silica gel column (180 g) in acetone. Elution with acetone first yields fractions of Ν, Ο-diacylated material, followed by fractions containing the monoacylated product. Acetone was evaporated from this last fraction and the residue was recrystallized from dimethylformamide / acetonitrile / ethyl acetate to give 9- (2- (2,2-dimethylpropionyloxyjethoxymethyl) guanine (0.5 g), mp 245-246 ° C.

Example 7

Preparation of 9- (2-palmitoyloxyethoxymethyl) guanine

9- (2-Hydroxyethoxymethyl) guanine (0.3 g) prepared according to German DOS 2 539 963 was partially dissolved in anhydrous dimethylformamide (15 ml) and pyridine (30 ml) and heated on a steam bath for 30 minutes. The mixture was cooled with an ice bath and palmitoyl chloride (1.1 g) was added with stirring. The mixture was cooled to room temperature, stirred for 20 hours until all solid material dissolved. Thin layer chromatography showed complete conversion of the starting material.

The solution was diluted to 300 mL with ethyl acetate and then cooled overnight. A white solid, 0.49 g, was collected by filtration.

The crude material was recrystallized once with ethyl acetate and once with acetone. Analytically pure 9- (2-palmitoyloxyethoxymethyl) guanine (190 mg) (31%), mp 190-210 ° C, was obtained.

NMR, DV and mass spectra are in accordance with the desired product.

Example 8

Preparation of 9- (2-butyryloxyethoxymethyl) guanine

A solution of sodium methoxide in anhydrous methanol (10 mL) was added to a methanolic solution of 9- (2-hydroxyethoxymethylguguine) at room temperature. Removal of the solvent in vacuo gave the purine sodium salt as a white powder.

The sodium salt prepared above was suspended in a mixture of ethyl butyrate and anhydrous dimethylformamide (20 ml) and heated to boiling with stirring. After 3 1/2 days the bulk of the solid is dissolved, the reaction mixture is concentrated to dryness in vacuo and the residue is dissolved in 2 N acetic acid (15 mL). Removal of the solvent in vacuo gave a semi-solid which, upon treatment with water (1.5 mL) and SD3A (15 mL) solidified to remove the solvents in vacuo. This product was suspended in SD3A (25 mL), filtered and dried in vacuo to give 195.2 mg of a tan solid tt 225-227.5 ^and C.

Example 9

Tablets containing a mixture of 9- (2-formyloxyethoxymethyl) guanine (100 mg), lactose (200 mg), starch (50 mg), polyvinylpyrrolidone (5 mg) and magnesium stearate ¹ (4 mg) are prepared by wet granulation.

Example 10

Tablets containing a mixture of 2-amino-9- (2-formyloxyethoxymethyl iadenine (100 mg), lactose (200 mg), starch (50 mg), polyvinylpyrrolidone (5 mg) and magnesium stearate (4 mg) are prepared by wet granulation.

Tablets containing a mixture of 9- (2-propionyloxyethoxymethyl) guanine (100 mg), lactose (200 mg), starch (50 mg), polyvinylpyrrolidone (5 mg) and magnesium stearate (4 mg) are prepared by wet granulation. Example 12

Tablets containing a mixture of 9- (2- (2,2-dimethylpropionyloxy) ethoxymethyl) guanine (100 milligrams), lactose (200 mg), starch (50 mg), lpyrrolidone (5 mg), and magnesium stearate (4 mg) were prepared by wet granulation.

Claims (2)

SUBJECT VYNALEZU A process for the preparation of 9-substituted purines of formula I CH (O-CH ₃ -C) - / O-CP 6 i Z t ⁰ (I) wherein R ¹ is hydroxy or amino, R ² is hydrogen, unbranched or branched alkyl having 2-16 carbon atoms and their salts, characterized in that the alcohol of formula V wherein R ¹ has the abovementioned meanings, is esterified by reaction with an appropriate acylating agent to introduce the group R ^2, and if the reaction product is a base, converting the product into a salt with an acid or an alkali metal, or if the product salt of a compound of formula I is converted optionally, a salt thereof or another salt thereof. 2. The process according to claim 1, wherein the acylating agent is a corresponding carboxylic acid, anhydride, acyl halide, mixed carboxylic acid anhydride and carbonic acid or carboxylic acid ester.