NO147186B - PROCEDURE FOR PREPARING SUBSTITUTED PURINE COMPOUNDS - Google Patents

Description

The present invention relates to a method for

synthesis of substituted purine compounds, especially 9-(2-hydroxyethoxymethyl) derivatives of the purines 2-amino-adenine and guanine.

The 9-(2-hydroxyethoxymethyl) derivatives of purines have antiviral activity against various types of DNA and RNA viruses,

both in vitro and in vivo experiments. In particular, these compounds are active as antiviral agents against adenoviruses, such as adenovirus 5 and rhinovirus. They are particularly active as an anti-viral agent against vaccinia and herpes viruses, such as simplex,

zoster and varicella in mammals, and such viruses produce diseases such as herpetic keratitis in rabbits and herpetic encephalitis in mice.

Examples of 9-(2-hydroxyethoxymethyl) derivatives of

purines that show particularly good antiviral activity are 9-(2-hydroxyethoxymethyl)-guanine and 2-amino-9-(2-hydroxyethoxymethyl)-adenine.

A number of methods are known for the production of 9-(2-hydroxyethoxymethyl) derivatives of purine, e.g. they can be prepared by removing a protecting group from the 2-position of the side chain. Alternatively, they can be prepared by converting 2- and/or 6-substituents on the purine ring to another substituent (see British Patent No. 1,523,865).

It has now been found that certain 9-(2-hyrdoxyethoxymethyl) derivatives of purines can be prepared by a new and advantageous synthesis route. The present invention thus provides a method for the preparation of compounds of formula I:

where R is amino or hydroxy; which includes, in the presence of a base, hydrolyzing a compound of formula II: 1 2 where R is hydroxy or -NR • 1 2 and X and X are the same or different, and each may represent an alkyl or phenyl-. 2 group, and R is hydrogen or

provided that when R is

hydroxy, then R is hydrogen. When X and X are alkyl groups, they preferably have from 1 to 4 carbon atoms, preferably they are of the same type and each represents a methyl group.

The base used during the hydrolysis may be an aqueous or alcoholic primary or secondary aliphatic amine, an alkoxide in alcohol, or an aqueous or alcoholic hydroxide, e.g. sodium alkoxide or hydroxide, and the preferred base is an aqueous primary aliphatic amine, e.g. aqueous methylamine. Depending on the base used, the hydrolysis can be carried out at temperatures from room temperature up to the temperature found in a steam bath. In general, the lower the reaction temperature, the longer the reaction time will be, but in return you get fewer side reactions.

An intermediate of formula II wherein R is

and where R<2> is can be completely hydrolyzed in one step, e.g. using aqueous methylamine. Alternatively and more advantageously, the intermediate product of formula II can be deacylated in 2 steps, in the first step a simple group will be removed from the 2 and 6 positions on the purine ring by mild hydrolysis at room temperature by e.g. uses butylamine in a lower alcohol. The second step in the hydrolysis, i.e. to remove the remaining ones

performed using stronger bases,

e.g. aqueous methylamine.

The intermediates of formula II are new and can be prepared by reacting guanine or 2-aminoadenine where the 2 and 9, or 2, 6 and 9 positions, respectively, are acylated, with a diester of 2-oxa-1,4-butanediol in the presence of a catalytic amount of a strong acid, e.g. sulfuric acid, sulfonic acid such as p-toluenesulfonic acid, methanesulfonic acid or trifluoromethanesulfonic acid, sulfamic acid, bis-(p-nitrophenyl)phosphate or polyphosphoric acid.

The acylated purine can in turn be produced by

reacting a suitable purine with an acid anhydride such as acetic anhydride, or another acylating agent, e.g. an acid halide.

To prepare the diester of 2-oxa-1,4-butanediol,

dioxolane can be reacted with an acid anhydride in the presence of a catalytic amount of a strong acid, e.g. of the type specified above.

The following examples illustrate the invention.

Example 1 - 9-(2-hydroxyethoxymethyl)guanine

A mixture of 120 g of acetic anhydride, 15 g of acetic acid and 5.0 g of p-toluenesulfonic acid was cooled to 10°C, and 70 g of dioxolane was added with stirring and cooling at such a rate that the temperature of the mixture did not exceed 40°C. The mixture was then cooled to room temperature, and 300 ml of toluene and 50 g of diacetylguanine were added. The reaction mixture was then heated to reflux for 16 hours. It was then cooled to room temperature and 50 ml of chloroform was added and the solid product removed by filtration. The filter cake was washed with chloroform and dried to give 2-acetamido-9-(2-acetoxymethyl)hypoxanthine. The dried filter cake was added to 350 ml of 40% aqueous methylamine, and the mixture heated under reflux for 40 minutes, then cooled and filtered. The filtrate was evaporated under reduced pressure to a thick suspension. This was cooled and filtered and the filter cake was washed with ethanol and dried, thereby obtaining 27 g of 9-(2-hydroxyethoxymethyl)guanine, purity greater than 90%, melting point 255-257°C, yield = 56%.

Example 2 - 9-(2-hydroxyethoxymethyl)guanine

A mixture of 50 g of diacetylguanine, 59.8 g of 2-oxa-1,4-butanediol and 1.2 g of p-toluenesulfonic acid in 350 ml of toluene was heated with stirring and refluxed for 16 hours. The mixture was cooled to room temperature, filtered, after which the filter cake was washed with toluene to give 2-acetamido-9-(2-ace-toxyethoxymethyl)-hypoxanthine. It was then dried and 350 ml of 40% aqueous methylamine was added. The mixture was heated under reflux for 40 minutes, cooled to room temperature and filtered. The filtrate was evaporated under reduced pressure to a thick suspension. To this was added 200 ml of ethanol and the whole was cooled, filtered and washed with ethanol and dried, thereby obtaining 36 g of 9-(2-hydroxyethoxymethyl)guanine, more than 90% pure, yield = 75%.

Example 3 - 2-acetamido-9-(2-acetyloxyethoxymethyl) hypozantine

(production of starting material)

A mixture of 1.0 g of diacetylguanine, 0.82 g of 2-oxa-1,4-butanediol diacetate and 23 mg of p-toluenesulfonic acid in 4 g of mineral oil was kept at 115°C under reduced pressure stirring overnight. The mineral oil was then poured off. The residue was treated with chloroform and then extracted with boiling methanol. The methanol extract was concentrated to 50 ml, cooled and filtered. The filtrate was evaporated to dryness, whereby 0.43 g of solid was obtained. This was purified by column chromatography (silica gel, 10 g in chloroform eluted with 1:1 chloroform:acetone), after which a recrystallization from ethanol gave 0.14 g of 2-acetamido-9-(2-acetyloxyethoxymethyl)hypoxanthine, melting point 202, 5-204.5°C.

Example 4 - 2-amino-9-(2-hydroxyethoxymethyl)adenine

(a) 89.0 g of 2-formamidoadenine was placed in a 5-liter flask equipped with an air stirrer and a condenser (CaCl2" drying tube) to which was added 4 L of acetic anhydride. The mixture was refluxed and kept under reflux for 60 hours. Then the excess of anhydride was removed by distillation at atmospheric pressure until about 3.5 liters of distillate had been removed. Distillation under reduced pressure was continued until most of the

the remaining anhydride. The dark brown residue was viscous on cooling to room temperature and was dissolved in dichloromethane.

It was filtered to remove suspended solids, after which the solution was evaporated in vacuo to give 211.0 g (> 100%) of 2,6-bis-(diacetylamino)-9-acetylpurine. The yield was 96.7% based on nmr, and the rest of the material was acetic anhydride. (b) 174 g of penta-acetylpurine was combined with 126.8 g of 1,4-diacetoxy-2-oxabutane in a flask equipped with a stirrer and drying tube. The mixture was placed in an oil bath at 130°C and stirred for a few minutes to homogenize the mixture. The acid catalyst, i.e. 2.74 g of para-toluenesulfonic acid, was then added in one portion and the heating was continued in vacuum for 4 hours, whereby an almost quantitative conversion to the products was then obtained. The reaction mixture was cooled to room temperature and stored under dry nitrogen. (c) The melt product (208.5 g) was dissolved in ethanol (5 ml/g) at room temperature and transferred to a flask equipped with a dropping funnel, stirrer and thermometer. 14 0.4 g of n-butylamine was then added dropwise over 2 hours, and the exothermic reaction was regulated by means of a water bath. The maximum temperature that allowed the reaction to proceed was only approx. 30°C. The product began to separate from the mixture during the actual addition. After the addition was complete, the mixture was stirred at room temperature for 3 hours and then placed in a cold room overnight. The product was removed by filtration and a paste-like mass was obtained which was stirred with 1 x 500 ml of acetone and then filtered again. The product was dried in vacuo at 65°C for 3 hours and then at room temperature overnight to give 154.2 g (91.7%) of a light brown hard solid. This product was purified by dissolving it in hot dimethylformamide (10 ml/g) at 100-110°C to give an opaque brown solution. After cooling overnight at 5°C, the product was removed by filtration, washed with 1 x 250 mL of acetone and air dried to give 121.7 g (78.8%) of 2,6-diacetamido-9-(2-acetoxyethoxymethyl ) purine. (d) 121.7 g of 2,6-diacetamido-9-(2-acetoxyethoxymethyl)purine was added to a stirred solution of aqueous methylamine (608.5 ml of a 40% solution) over 5 minutes. The addition was followed by a mild exothermic reaction which raised the temperature of the mixture to 35°C and all solids dissolved within a few minutes. After stirring for 2 1/2 hours, thin layer chromatography of the mixture showed complete reaction. The mixture was then concentrated in vacuo on a water bath at 45-50°C and a thick mass of brown crystals was obtained. These were stirred with 545 ml of acetone for fifteen minutes to remove N-methylacetamide and then vacuum filtered. The filter cake was washed with acetone (1 x 200 ml) and air dried, whereby 74.7 g (96.0%) were obtained

impure hydrogenated 2-amino-9-(2-hydroxyethoxymethyl)adenine as light brown crystals. These were dried in vacuum at 80°C for 18 hours, and 69.3 g (89.0%) of dry product was obtained.

Example 5

2-acetamido-9-(2-acetyloxyethoxymethyl)hypoxanthine (6.19 g) was suspended in 1N sodium hydroxide (40 mL) and heated at reflux for 1 hour. The mixture was cooled, neutralized with 6N hydrochloric acid and the product filtered off. The solid was washed with ice water (10 mL) and with acetone (15 mL) and dried under vacuum overnight. Weight 4.13 g (91.8%) of 9-(2-hydroxyethoxymethyl)guanine, m.p. 255-257°C.

Example 6

2-acetamido-9-(2-acetyloxyethoxymethyl)hypoxanthine

(6.19 g) was slurried in a solution of sodium methylate (3.24 g) in methanol (40 ml). The mixture was heated at reflux for 1 hour and allowed to cool. Glacial vinegar (3.96 g) was added and the mixture poured into water (40 ml). After cooling,

0°C, the resulting precipitate was filtered, washed with cold water (15 mL) and with acetone (15 mL) and dried under vacuum to give 4.03 g (89.6%) of 9-(2-hydroxyethoxymethyl)guanine ,

m.p. 255-257°C.

Example 7

2-acetamido-9-(2-acetyloxyethoxymethyl)hypoxanthine

(6.19 g) was added to a solution of potassium hydroxide (3.96 g) in ethanol (40 mL) and heated at reflux for 1

hour. The mixture was allowed to cool and glacial acetic acid (3.96 g) was added. The mixture was poured into water (40 ml) and 50 ml of water was added. The pH was adjusted to 7.0 with aqueous sodium hydroxide and the mixture cooled with ice-water and filtered. The solid washed with cold water (20 ml) and with acetone (20 ml)

and dried under vacuum to give 4.0 g (88.9%) of 9-(2-hydroxyethoxymethyl)guanine, m.p. 255-257°C.

Example 8

2-acetamido-9-(2-acetyloxyethoxymethyl)hypoxanthine

(6.19 g) was slurried with triethylamine (5 g) and water (40 mL) and the mixture heated at reflux for 2 h. The mixture was allowed to cool to 0°C and was filtered. The solid was washed with water (15 ml) and with acetone (15 ml) and dried under vacuum to give 4.34 g (96%) of 9-(2-hydroxyethoxymethyl)guanine, m.p. 255-257°C.

Claims (2)

1. Process for the preparation of compounds of formula I: where R is amino or hydroxy, characterized in that a compound of formula II is hydrolysed in the presence of a base: where R"*" is hydroxy or 1 2 and X and X are the same or different and each may represent an alkyl or 2 phenyl group, and R is hydrogen or provided that when R is hydroxy, then R <2>hydrogen. 2. Method according to claim 1, characterized in that an aqueous or alcoholic, primary or secondary aliphatic amine, an alkoxide in alcohol or an aqueous or alcoholic hydroxide is used as the base.