CS263955B1 - Method of heterocyclic bases' n-(3-hydroxy-2-phosphonylmethoxypropyl)derivatives production - Google Patents

Abstract

A new process for the preparation of N- (3- -hydroxy-2-phosphoinylmethoxypropyl) derivatives the purine and pyrimidine bases which lie in the reaction of N- (2,3-dihydroxypropyl) derivatives with chloromethanephosphonyldichloride v phosphoric acid trialkyl esters, hydrolysis and simultaneous isomerization to 3‘-chloromethanephosphonylesters N- (2,3-dihydroxypropyl) - heterocyclic base derivatives and their conversion to the subject compounds by treatment aqueous alkali hydroxide solutions.

Description

The present invention provides a process for the preparation of N- (3-hydroxy-2-phosphinylmethoxypropyl) heterocyclic base derivatives.

9- (S- (3-Hydroxy-2-phosphonylmethoxypropyl) adenine (HPMPA) is a highly potent virostatic with an effect primarily against DNA viruses (herpesviruses, poxviruses, adenoviruses, US Patent No. 263,252).

Previous processes for its preparation consisted either of condensation of the sodium salt of specifically protected 9- (S - (2,3-dihydroxypropyl) adenine) with p-toluenesulfonyloxymethanphosphoic acid diester followed by deprotection and alkaline hydrolysis (cf. No. 263 951), or in the reaction of N-substituted 9- (S) - (2,3-dihydroxypropyl) adenine with chloromethanephosphonyldichloride followed by treatment with aqueous alkali hydroxides to the 9- (S) - (2,3-dihydroxypropyl) transitional 3'-O-chloromethanephosphonyl ester adenine (Czech author's certificate no. 233 665).

The latter method is apparently simpler, but provides a mixture of HPMPA with its isomer [9- (S) - / 2-hydroxy-3-phosphonylmethoxypropyl / adenine], which is biologically ineffective and must be separated. This disadvantage is overcome by another process which increases the representation of the above intermediate, 3'-O-chloromethanephosphonyl ester, by acidic isomerization of the reaction mixture and removes the unnecessary 2'-O-isomer by a separation procedure that allows its reuse (cf. 263 263). No. 263,954), thus allowing a substantial increase in HPMPA yield.

In order to achieve a high degree of esterification of the starting material with chloromethanephosphonyldichloride in the presence of pyridine or another tertiary base (cf. author's certificate no. 233 665) or its partial hydrolysis product in pyridine (cf. author's certificate no. 263 954), a considerable excess of reagent Czech Author's Certificate No. 263 954); moreover, because of the risk of side reaction at the amino group, the starting material must always be protected by an acyl or amidine function.

These disadvantages are overcome by the present invention for the preparation of N- (3-hydroxy-2-phosphonylmethoxypropyl) heterocyclic base derivatives of the general formula I-CH 2 CH-OCmP (O (OH) 2 CH 2 OH (I) wherein

R ¹ is a purin-9-yl or pyrimidin-1-yl residue, characterized in that the N - (1,2,3-dihydroxypropyl) derivative of the heterocyclic base of formula II

R ^{1 =} CH ² CH (OH) CH 2 OH (Π) wherein

R ¹ has the same meaning as in formula I, is reacted with 1.03 to 2 molar equivalents of the formula III chlormethanfosfonyldichloridu

ClCH2P (O) Cl 2 (III) in trialkyl phosphate esters of formula IV (R ² O] 3 P = O (IV) wherein

R ² is methyl or ethyl, at temperatures of 0 to 30 ° C, the mixture is precipitated with 5 to 10 volumes of ether based on IV, the precipitated product is dissolved in water to a final concentration of 25 to 150 mmol. . The I- ¹ mixture is heated to 100 to 110 degrees Celsius for 8 to 24 h to form the 3'-O-chloromethanephosphonyl ester of formula V

R ^{1 =} CH ² CH (OH) JCH 2 OP (O) (OH)> CH 2 Cl (V) wherein

R ¹ has the same meaning as in formula I, is isolated on a column of hydrophobized silica gel, heated to 50 to 100 ° C in 1 to 6 moles. Even ^-1 aqueous caustic solution and the compound of formula I is isolated, preferably by desalting or- chromatography.

The present process is based on the known fact that phosphoric acid chlorides react with alcohols in trimethyl or triethyl phosphate in high yields to ester chlorides which can be hydrolyzed to the corresponding esters alkaline or acid (M. Yoshikawa, T. Kato, T. Takenishi: Bull). Soc. Chem. Japan 42, 3, 505 (1969).

In contrast to the ribonucleosides or deoxyribonucleosides which, when reacted with phosphorus oxychloride under these conditions, only give the esters of the primary alcohol function, the 2,3-dihydroxypropylderivatives of the formula II preferentially esterify the secondary alcohol group at the 2-position. 15 to 20 hours and using 1.03 to 1.2 molar equivalents for purine base derivatives, or 1.5 to 2.0 mol equivalents for pyrimidine base derivatives.

Precipitation of the reaction mixture with ether gives a mixture of isomeric chloromethanephosphonic acid ester chlorides which can be converted by hydrolysis in a neutral medium to the isomeric chloromethanephosphonic acid esters of compounds of formula II. However, to substantially increase the proportion of the 3'-isomer of the formula V required, said precipitate of isomeric ester chlorides may be heated in 0.5 to 10 moles. I ^-1 hydrochloric acid according to the principle of MS. author's certificate No. 263 953.

6 3 9 5 6

A marked simplification of the process is the utilization of the intrinsic acidity resulting from ester chloride hydrolysis: isomerization to form the compound of formula V occurs simply by boiling a solution of the reaction intermediate in water. Usually a boiling time of 15 to 24 hours is required to achieve an equilibrium mixture. From this mixture, the pure 3‘-isomer of formula V is isolated after concentration by preparative liquid chromatography according to the procedure described in U.S. Pat. In many cases, the content of the compound of formula V in the reaction mixture exceeds 90% and the product can be used directly for the next preparation step.

Substances of formula V (or said high-enriched enriched mixture) are converted to compounds of formula I by the intiramolecular cyclization reaction, which proceeds at a slightly elevated temperature in aqueous alkali (sodium, sodium, After completion of the reaction, which usually takes 5 to 8 hours at 70 ° C, the mixture is freed of excess alkali and, in the case of a previously purified intermediate of formula V, the compound of formula I is isolated after desalting as free acid or better soluble alkali salt. When using a crude mixture with an enriched compound of formula V, it is appropriate to purify the desalted reaction mixture by anion exchange chiromatography (preferably DEAE-modified polymers) and isolate the compounds I as alkali salts.

The advantage of this process is that it can be used directly for compounds of formula II without protecting the heterocyclic base, significant savings of reagent of formula III, simplicity of implementation, easy purification of compound of formula V, and good final yield of compound of formula I. HPMPA mentioned antivirals (I, ^R1 = adenine-9-yl), as well as derivatives thereof substituted on the adenine ring, and in many other analogs of purine and pyrimidine.

The present invention is further illustrated by the following non-limiting examples of the preparation of the compounds of formula I and exemplified in Tables 1 and 2.

Example 1

A mixture of 627 mg (3 mmol) of 9- (S) - (2,3-dihydroxypropyl) adenine and 10 ml of triethyl phosphate is heated to dissolve, cooled with ice and 0.40 ml (3.09 mmol of chloromethanephosphonyl dichloride) is added with stirring. 20-25 ° C for 24 hours, 100 ml of ether are added with stirring, the mixture is filtered off with suction, washed with ether and the precipitate is dried for 30 min at 13 Pa. This product is dissolved in 20 ml of water and refluxed for 10 hours. After cooling, the mixture is applied to a 100 ml cation exchange column in an H ⁺ cycle (e.g. Dowex 50 X8), the column is eluted with water and the UV-absorbing eluate is evaporated at 40 ° C / 2 kPa. alkaline with triethylamine to dissolve and concentrate the solution in vacuo (see above) to a volume of 1.5 ml.

0.2 ml portions of this solution are loaded onto a column of octadecylsilica gel (7 µl) (8 x 500 mm) equilibrated with 0.05 mol. Even ^-1 triethylammonium, pH 7.5 and the column is washed first with the same buffer to a decrease in absorption at 254 nm and then with methanol same Putna (1: 9 obj.j, elution rate 2 ml / min. Elutes first 3'- isomer (compound of formula VJ, followed by 2'- isomer.

The fractions of compound V are combined after complete application, evaporated in vacuo (see above) and the residue is heated in 20 ml of 2 moles. I ^"1 NaOH 5 hours at 80 ° C. The mixture was neutralized by addition of a cation-exchange resin in the H ⁺ -cycle, evaporated in vacuo and dried by codistillation with ethanol. The product was suction filtered from ethanol / ether (1:10), washed with ether and dried. 500 mg of sodium salt of the compound of formula I (HPMPA) is obtained.

Example 2

A mixture of 627 mg (3 mmoles] 9- (RSj- (2,3-dihydroxypropyljadeninu and 10 ml of trimethyl phosphate was treated in the same way as in Example 1. After column chromatography on cation exchanger was obtained chlormethanfosfonát formula V treated with 1 mol. I ^-1 with lithium hydroxide according to Example 1 and after neutralization of excess hydroxide by cation exchange resin in the H ⁺ -cycle, the mixture was evaporated and dried by codistillation with ethanol in vacuo .The residue was stirred with acetone-ethanol (4: 1j (25 ml) for 30 min). acetone, ether and dried to give a 75% yield of the lithium salt containing 86% of the compound of Formula I (R ¹ = adenin-9-yl).

Example 3

A mixture of 3 mmol of 1- (RS) - (2,3-dihydroxypropyljuracil) and 10 ml of triethyl phosphate is treated after treatment with 0.6 ml of chloromethanephosphonyldichlodir as described in Example 1. After boiling in water for 10 hours, the acidic solution is neutralized with 1M sodium hydroxide. the mixture was evaporated to dryness in vacuum (see above], and the residue heated in 20 ml of 6 mol. I ^-1 NaOH 5 hours at 60 ° C. the mixture was neutralized with a cation exchanger in the H ⁺ form, filtered, concentrated in vacuo to a volume of 20 ml and this solution was loaded onto a 100 ml column of anion exchange resin (e.g. Sephadex a-25) in HCO 3 ^- cycle. the column was eluted in the absorption loss of 0.02 mol. I ^-1 triethylammonium, pH 7.5 and then increasing concentrations of the same buffer to final values of 0.2 mole.l ^-1 over 2 liters (elution rate 3 ml. min ^-1 ).

The UV-absorbing product fraction was evaporated in a water-jet vacuum, the residue codistilled with methanol (3 x 50 ml) and the residue in 10-15 ml of water was applied to a cation exchange column.

6 3 9 5 6

R in Na @ ⁺ -cyclile. This column is eluted with water, the UV-absorbing eluate is evaporated, the residue is dried by codistillation with ethanol and suction filtered from a mixture (100 ml) of methanol and ether (1:10), washed with ether and dried. Yield 67% of the sodium salt of l- (RS) - [3 (2) -hydroxy-2 (3) -fosfonylmethoxypropylj-uracil and containing 72% of the compound of formula I (R ¹ = uracildL-yl).

Example 4

A mixture of 10 mmol of 9- (RS) - (2,3-dihydroxypropyl) -2-methylthioadenine and 35 ml of triethylphosphate was ice-cooled, 1.4 ml of chloromethanephosphonyl dichloride was added and stirred at 20 ° C for 20 hours. 200 ml of ether are added, the precipitate is filtered off with suction, washed with ether and dissolved in 70 ml of 0.5 mol. 1 - ¹ hydrochloric acid. The solution was heated to reflux for 20 hours, evaporated to dryness at 40 degrees C / 2 kPa and applied to a column of 200 ml of a cation exchange resin in a H ⁺ -cycle in 20 ml of water. Elution with water first removes hydrochloric acid, then washes with UV-absorbing compound of formula V. This eluate is evaporated in vacuo (see above) and treated with 40 mL of 2 moles. I ^"1 sodium hydroxide according to Example 1 to give 82% yield of the sodium salt of 9- (RS) - [3- (2-hydroxy-2 j (3) -f onylmethoxypropylj OSF-2-methylthloadeninu containing 89% isomer of Formula I.

263935

Preparation of N- (3-hydroxy-2-phosphonylmethoxypropyl) heterocyclic base derivatives of formula I

Number R ¹ Absolute Number of Equivalents Method of Total Yield ³ 'Representation of configuration on compounds III of Example% Isomer I

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Table 2

Properties of N- (3-hydroxy-2-phosphonylmethoxypropyl) heterocyclic base derivatives of formula (I) # ³ 'RF ^b ' Eup ^c) , ^d) Contents ^(e) tCNNOINCOINO-oolithiol OO Η O Λ Η φ Η HQO inf. ? in ”in” what d cm ”cm” what ”

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Explanatory notes to Table 2:

^a) Compound number of the formula I according to Table 1 ^b) Paper chromatography in 2-propanol-conc. aqueous ammonia-water ^c) Electrophoretic mobility (relative to uridine-3'-phosphate) in paper electrophoresis at 0.05 mol. I ^-1 triethylammonium bicarbonate pH 7,5 (20 V / cm) ^d) Elution constant in liquid chromatography column (200 X 4 mm) of ACS G18 silica gel (10 μ] in a 0,05 mol mixture. I ^-1 triethylammonium bicarbonate pH 7, 5 and methanol (percentages by volume in parentheses) elution rate 1 ml.min ^-1 , k = (t _R / t _o - 1), where t _R is the elution time of the substance and t _{0 the} elution time of the dead volume of the column ^e) spectrophotometrically at pH 2 based on A _max values for compounds of the formula

Claims (1)

A process for the preparation of the heterocyclic base N- (3-hydroxy-2-phosphonylmethoxypropyl) derivatives of the general formula I R ¹ = CH ² CH-OCH 2 P (O) (OH) 2 CH 2 OH (I) wherein R ¹ is a purin-9-yl or pyrimidin-1-yl radical, characterized in that the N + 2,3-dihydroxypropyl derivative of the heterocyclic base of formula II R ^{1 =} CH ² CH (OH) CH ² OH (II) wherein R ¹ has the same meaning as in formula I · is reacted with 1.03 to 2 molar equivalents of the formula III chlormetbanfosfonyldichloridu dCH2P (O) Cl 2 (III] in the trialkyl phosphate esters of formula IV OF THE INVENTION (R ² O) 3P = O (IV) where R is ^a methyl or ethyl group, at temperatures of 0 to 30 ° C, the mixture was precipitated from 5 to 10 volumes of ether, relative to the compound of Formula IV, the precipitated product is dissolved in water to a concentration of 25-150 mM. . The I- ¹ mixture is heated at a temperature of 10 ° to 110 ° C for 8 to 24 h, the resulting S, O-chloromethanephosphonyl ester of formula V R ^{1 =} CH ² CH (OH) CH 2 OP (O) (OH) CH 2 Cl (V) wherein R ¹ has the same meaning as in formula I, is isolated on a column of hydrophobized silica gel, heated to 50 to 100 ° C in 1 to 6 moles. Γ ¹ aqueous caustic solution and the compound of formula I is isolated, or preferably odsolenlm · chromatography.