CS263953B1 - Method of 9-(s) or 9-(rs)-(3-hydroxy-2-phosponylmethoxypropyl)adenine production - Google Patents

Abstract

An improved process for the production of viral preparation 9- (S) -, resip. 9- (RS) - (3-hydroxy-2-phosphonylmethoxypropyl) adenine rests in the isomerization of 2'-O-chloromethylphosphonyl- and 3‘-O-chloromethylphosphonyl esters 9- (S) -, respectively. 9- (RS) - (2,3 dihydroxypropyl) - adenine in the acid medium it provides 70 to 80% equilibrium mixture the required 3‘-isomer. This mixture can be divided, preferably by preparative liquid chromatography and the 2‘-isomer is recycled to isomerization. 3‘-O-chloromethylphosphonyl-9- - (2,3-dihydroxypropyl) adenine provides action alkali metal hydroxides 9- (3- -hydroxy-2-phosphonylmethoxypropyladienine.

Description

The present invention relates to a process for the preparation of 9- (Sj- or 9- (RS) - (3-hydroxy-2-phosphonylmethoxypropyl) adenine.

- (S) - (3-Hydroxy-2-phosphorylmethoxypropyl-adenine (HPMPA) is a nucleotide analogue that has a strong antiviral effect, specifically directed against DNA viruses, characterized by its action on herpesviruses, including those strains and their mutants, which resist previously known virostatics, such as acyclovir (cf. No. 263 952). A racemate of this substance has a similar effect, whereas its (R) -enantemometer and (R) -, (S) - i (RS) The isomeric form of 9- (2-hydroxy-3-phosphonylmethoxypropyl Jadenine) is almost ineffective, so the technical problem to solve is the preparation of the isomerically pure 2-isomer of HPMPA [(Sj-form or racemate), or a mixture of both isomers with the highest forms.

HPMPA has so far been prepared, for example, by the reaction of N ⁸ , O ³ '-disubstituted 9- (2,3-dihydroxypropyl) adenines with p-toluenesulfonyloxymethanephosphonic acid diester, followed by cleavage of the protecting and ester groups (CSN 263 263) or by reaction N ⁸ -substituted 9- (2,3-dihydroxypropyljaidenine with chloromethylphosphonyldichloride and subsequent alkaline hydrolysis (cf. No. 233 665).

While the former provides pure 2 poskytuje-phosphonylmethoxy derivative HPMPA, it requires prior introduction of protecting groups both in the heterocyclic ring and in the 3 polohy position, as well as their subsequent removal.

The second method is simpler, but always provides a mixture of both isomers, of which the desired HPMPA is present in the mixture at most 30-40%. Both isomers need to be separated and the by-product is no longer usable. Therefore, neither of the two prior art processes is a technically advantageous way of preparing the active ingredient.

The above-mentioned disadvantages are overcome by the present invention for the preparation of 9- (S) - and 9- (RS) - (3-hydroxy-2-phosphonylmethoxypropyl) adenine of the formula I

CH-CH-CH-OH and _from p (o) (oh) _z (I) and its alkali metal and ammonia salts by the mixture of isomeric chloromethylphosphonyl esters 9- (Sj- and 9- (RSj- (2)). (3-dihydroxypropyl) adenine of formula II

(II) where

R ¹ is hydrogen and

R ² is -P (O) (OHjCHzCl or R ² is hydrogen and R ¹ is -P (O) (OHJCH2CI group, solubilized to a final concentration of 25 to l 50 mmol. 1 ^_1, based on the compound of formula II , mineral or organic acid concentration of 0.5 to 10 moles. 1 ^_1, preferably in hydrochloric acid, the mixture is kept at 40 to 80 ^K C for 8-24 h, after removal of excess acid the compound of formula II ( R ¹ is hydrogen, R ² is -P (Oj (OHJCH ² Cl group) is isolated by ion exchange or hydrophobized slag gel chromatography, heated in aqueous solutions to a final concentration of 1 mole ^-1 to 6 moles l · ¹ alkali hydroxides at temperature 40-100 ° C and the compound of formula I is obtained by neutralization and desalting.

The invention is based on the fundamental improvement and streamlining of the known process for the preparation of the compound of formula I and its 3'-isomer, i.e. 9- (2-hydroxy-3-phosphonylmethoxypropyl kernin) cyclic esters of formula II in an alkaline environment according to US Patent No. 233,665 .

Preparation of compounds of formula (I) by reaction of 9- (2,3-dihydroxypropyl adenine) with chloromethanesulfonyldichloride in pyridine (cf. author's certificate no. 233 665) or its hydrolysis product with an equimolar amount of water in pyridine [I. Rosenberg, A. Holý, Collect. Chem. Commun., 50, 1, 507 (1985)] provides the 3'-isomer of Formula II (R ¹ is hydrogen, R ² is -P (Oj (OHJCH ² Cl group)) I, only in minor amounts (30 to 40%) relative to the unused 2'-isomer [II, R ² is hydrogen, R ¹ is a -P (O) (OH) CH 2 Cl 2 group] (hereinafter 2'- ll).

The present invention is directed to the isomerization of the latter 2‘-II isomer to the 3‘-isomer of formula 3‘-II, which takes place in aqueous solutions of organic, but preferably mineral acids (hydrochloric acid, sulfuric acid, etc.) at slightly elevated temperatures. This process results in the equilibrium of a high mixture (75-80 percent) of the 3‘-isomer of Formula 3‘-II, equal to 263953. The composition of this mixture is dependent on the acid concentration and reaction time. These data are shown in Table 1. The resulting mixture does not change when the mixture is treated in an acidic or slightly alkaline environment.

The mixture of compounds of formula II thus enriched in the 3'-isomer of formula 3'-II can be effectively separated into the individual isomers after removal of the excess acid by various means, most preferably ion exchange chromatography on strongly acidic cation exchange or hydrophobized silica gel, or can be used directly as an enriched mixture for cyclization . Separation at the level of compounds of formula II is much more efficient than chromatographic purification of isomers of compound of formula I.

A further advantage of the present invention is that the by-product, the 2 is-II isomer can be re-converted by isomerization to the 3 vzorce-II compound in an acidic environment, thereby increasing the degree of utilization of the starting material and other reagents.

The isomerization of the mixture of compounds of formula II proceeds equally well regardless of the method for its preparation. The isomerically pure 3'-O-chloromethanesulfonyl ester of formula 3'-II is subjected to an alkaline treatment according to the conditions of the original procedure (cf. No. 233 665) to give an isomerically pure product of formula I (HPMPA); Similarly, the 2'-II gives the pure 3'-isomer of the compound of formula I [9- (2-hydroxy-3-phosphonylmethoxypropyl) adenine] Further purification of the compound of formula I is no longer necessary, of excess hydroxide and alkaline chloride by desalination, eg on cation exchangers.

There is no racemization or configuration change at the 2-position carbon atom during the procedure. The process of the present invention does not require special equipment or chemicals and is a significant improvement in the state of the art for the preparation of the compound of formula I. High purity of the product, high yield and increased process efficiency are particularly important. Separation of the isomeric compounds of formula (II) can easily be achieved on a large scale by preparative liquid chromatography with the appropriate volume of the stationary phase and at suitably selected column parameters.

In the following, the process according to the invention is exemplified without limiting it in any way.

Example 1

A mixture of 522 mg (2.5 mmol) of 9- (RS) - (2,3-dihydroxypropyl) adenine, 10 ml of dimethylformamide and 3 ml of dimethylformamide dimethyl acetal is stirred in a sealed flask for 16 h at room temperature and evaporated at 40 ° C / 13 Pa. Dissolve in 20 ml 50% aqueous pyridine and add about 1 g solid carbon dioxide, evaporate after 1 h and co-distill 3 times with 20 ml pyridine at 40 ° C / 13 Pa. To a solution of 1.87 g (10 mmol) of chloromethanephosphonyl chloride in 2.0 ml of pyridine is added 180 [mu] l of water at 0 [deg.] C., after 30 min in an ice bath, the mixture is filtered under exclusion of moisture to the above residue and left at room temperature for 2 h. mol. I ¹ triethylammonium hydrogen carbonate, pH 7.5, concentrated at 40 ° C / 2 kPa and the residue evaporated three times with 50 ml of ethanol under the same conditions.

The residue is left to stand at room temperature for 16 hours in 100 ml of 5% aqueous ammonia and evaporated to dryness at 40 ° C / 2 kPa. According to liquid chromatography analysis:

column 4 X 200 mm Separon CGX C18 ACS (10 µm), mobile phase 5% methanol in 0.05 mol.

. I ^-1 triethylammonium ^hydrogen carbonate,

PH 7.5 (v / v, elution rate 0.7 ml / min ^-1 , detection at 254 nm contains 95.5% mixture of compound II containing 33% of 3'-H (t = 18.8 min) and 67% of 2'-II (t = 14.7 min) This residue was loaded onto a 100 mL ion exchange column (e.g., Dowex 50 X8) in an H ⁺ cycle in 20 mL of water and eluted with water (2 to 2 mL). 3 ml. min ^-1) (300 mL) and a radius of ^2.5!% aqueous ammonia (1 l). the ammonia eluate is evaporated at 40 ° C / 2 kPa and the residue in 40 ml of 1 mol. I ^"1 hydrochloric acid heated The mixture is applied to 100 ml of cation exchange resin (e.g. Dowex 50 X8) in an H ⁺ -cycle and the column is eluted with water (3 mL min ^-1 ) to detect eluate at 254 nm.

The main UV-absorbing fraction was evaporated at 40 ° C / 2 kPa (76% 3'-II content, 24% 2'-II content) and the residue was heated to 80 degrees Celsius in 40 ml of 2 moles for 6 h. I ^"1 sodium hydroxide. After cooling, the mixture is neutralized by addition of an ion exchanger (Dowex 50 X8) in an H + -cycle to pH 7.0, filtered, washed with 100 ml of water and the filtrate is evaporated at 40 ° C / 2 kPa. Precipitated from ^: 5 ml of methanol by addition of 100 ml of ether After aspiration and drying in vacuo, 700 mg of the disodium salt of the racemic compound of the formula I (81%) is obtained, containing 76% of the substance I and 24% of its 3'-isomer.

627 mg (3 mmol) of 9- (S) - (2,3-dihydroxypropyljadeninu was converted according to Example 1 a mixture of substances of the formula II, which is isomerized by heating with 40 ml of 10 moles. I ^"1 HCl 8 hours at 60 ° C The mixture is directly loaded onto a 200 ml cation exchange column (e.g., Dowex 50 X8) in an H ⁺ -cycle and eluted with water (3 ml / min) to give a 81% blend of the compounds of Formula II from the main UV-absorbing strip. 3'-II This residue was dissolved in 10 ml of water by addition of trlethylamine and the solution was concentrated in vacuo at 40 ° C / 2 kPa to a volume of 1.5 ml. 3 ml portions of this solution were applied to column 8. XX 500 mm octadecyl-silica gel (e.g. Separon SIX C18, 7 µm), equilibrated 0.05 mol. With triethylammonium ^hydrogen carbonate, pH 7.5.

The column is eluted first with the same buffer to remove salts and impurities and then with a 10% methanol solution in the same buffer (elution rate 2 ml / min.) The eluates of the two isomers II are collected separately. evaporated at 40 ° C / 2 mbar. the purity determined by HPLC (see example 1) · in both cases higher than 99%. the residues of both isomers were separately treated with 10 ml of 2 moles. I ¹ sodium hydroxide according to example 1 to give with 500 mg (56% based on II) of HPMPA of the formula I as the disodium salt of more than 99%.

SUBJECT

Process for the preparation of 9- (S) - and 9- (RS) - (3-hydroxy-2-phosphonylmethoxypropyl adenine of the formula I)

CHoCH-CH-OH

I Z.

α H, P (δ) (OH).

(I) and their alkali metal and ammonia salts, characterized in that a mixture of isomeric chloromethylphosphonyl esters of 9- (Sj- and 9- (RS) - (2,3-dihydroxypropyl adenine) of formula II nh ₂

Table 1

Effect of hydrochloric acid concentration on isomerization of a mixture of compounds of formula II (25 mmol. I ¹ ) for 24 h at 37 ° C

Concentration%

HCl 2'-II 3'-II (mol 1 - ¹ )

0 84.6 15.4 0.1 79.0 21.0 0.2 .74.0 26.0 0.5 57.7 42.3 1.0 27.0 73.0 2,0 21.0 79.0 5.0 15.0 85.0 10.0 13.0 87.0 Note: When performing isomerization at 2.5 mole. 1 ¹ sulfuric acid under otherwise identical conditions, the content of 3‘-II after 24 h is 81%.

Claims (1)

where R ¹ is hydrogen and R ² is -P (O] (OHjCHzčl or R ² is hydrogen and R ¹ is - P (O} (OH) CH2 group, is brought into solution with a final concentration of 25-150 mmol .l ^-1 compound of formula II a mineral or organic acid having a concentration of 0.5 to 10 moles, ^i.e. , preferably in hydrochloric acid, the mixture is maintained at a temperature of 40 to 80 ° C for 8 to 24 hours, after removal of the excess acid, the compound of formula II, wherein R ¹ is hydrogen and R ² is -P (O) (OH) CH2Č1 group isolated ion exchange chromatography or hydrophobic silica, is heated in aqueous solutions of the resultant "concentration of 1 mol. 1 to 6 molů.l ^{_1" 1} alkali hydroxides at 40 to 100 ° C and the compound of formula I is obtained by neutralization and desalting. CH ₂ CH-CH ₂ 0? ^< or * 01) Severography, n. Race 7, Most Gena 2,40 Kčs