PL193592B1 - Analogues of polydinucleoside phosphates modified in their polyphosphate chain and method of obtaining them - Google Patents

Abstract

1. Dinucleoside polyphosphate analogs, modified in the polyphosphate chain, with general Formula 1 wherein Z1 is hydroxy or a hydrogen atom, B1 is an adenine, guanine residue, cytosines, thymines, X and Y are oxygen or a sulfur atom, R1 is a polyol residue, preferably glycerol, erythritol, pentaerythritol or bis- (hydroxymethylene) phosphine of the formulas 2, 3, 4, 5 where Z2 is a hydrogen atom or a phosphoryl residue or thiophosphoryl or dithiophosphoryl or 5'-phosphorylated nucleosides or 5'-thiophosphorylated nucleosides or 5'-dithiophosphorylated nucleosides or 3'-phosphorylated nucleosides or 3'-thiophosphorylated nucleosides or 3'-dithiophosphorylated nucleosides.

Description

Description of the invention

The invention relates to dinucleoside polyphosphate analogs, modified in the polyphosphate chain, of the general formula 1, in which Z ¹ is a hydroxyl group or a hydrogen atom, B ¹ is an adenine, guanine, cytosine, thymine residue, X and Y are an oxygen or sulfur atom, R ¹ is a polyol residue, preferably glycerol, erythritol, pentaerythritol or bis (hydroxymethylene) phosphinic acid of formulas 2, 3, 4, 5 where Z ² is a hydrogen atom or a phosphoryl or thiophosphoryl or dithiophosphoryl residue or 5'-phosphorylated nucleosides or 5'-thiophosphoryl nucleosides or 5'-dithiophosphorylated nucleosides or 3'-phosphorylated nucleosides or 3'-thiophosphorylated nucleosides or 3'-dithiophosphorylated nucleosides, and a method for their preparation.

The occurrence of dinucleoside polyphosphates in living organisms was first described by Zamecnik. Since then, purine and pyrimidine receptors have been tested for their affinity to nucleotides that contain negative charges of various sizes in their molecules (tri-, tetra-, penta-, hexaanions). The physiological functions of dinucleoside polyphosphates are recognized thanks to the synthesis and use of appropriate analogues in biological experiments. Of particular note is the fact that Ap4A and its analogs have potential therapeutic importance. In vitro studies have shown that these compounds are competitive inhibitors of ADP-dependent platelet aggregation. Since natural compounds undergo cellular metabolism, there is a need for analogues of natural compounds with selective and potent inhibitory properties in this process. Recently, it has been shown that AP3A and Ap4A are the preferred substrates for the human protein encoded by the gene "Fhit putative tumor suppressor", the presence of which is closely related to the tumorigenesis process. New chemical compounds with selectivity of action in relation to the FHIT protein may complement cancer diagnostics.

The dinucleoside polyphosphate analogs represented by the general formula 1 are not known to date.

Analogs dinukleozydopolifosforanów modified chain polyphosphate having the general formula 1, wherein Z ¹ is hydroxy or hydrogen, B ¹ is a residue of adenine, guanine, cytosine, thymine, X and Y are oxygen or sulfur, R ¹ represents the residue of a polyol , preferably glycerol, erythritol, pentaerythritol or bis (hydroxymethylene) phosphinic acid of the formulas 2, 3, 4, 5 where Z ² is a hydrogen atom or a phosphoryl or thiophosphoryl or dithiophosphoryl residue or 5'-phosphorylated nucleosides or 5'-thiophosphorylated nucleosides '-dithiophosphorylated nucleosides or 3'-phosphorylated nucleosides or 3'-thiophosphorylated nucleosides or 3'-dithiophosphorylated nucleosides.

₁

A method for the preparation of dinucleoside polyphosphate analogs of general formula I, wherein Z ¹ , 11

B ¹ , X, Y and R ¹ are as defined above, according to the invention, the nucleoside of the general formula 6, in which Z ³ is a hydrogen atom or a blocking group, Z ⁴ is a blocking group,

B ² represents an adenine, guanine, cytosine residue of the formulas 7, 8, 9 where Z ⁵ is a hydrogen atom or an exoamine function blocking group or a thymine residue of the formula 10, is subjected to a condensation reaction with a compound of the general formula 11, where X has the above-mentioned meaning, Y is oxygen or sulfur, R ² represents derivatives of polyols, preferably glycerol, erythritol, pentaerythritol, or bis (hydroxymethylene) a phosphine of the formulas 12, 13, 14, 15, where X and Y are as defined above, in the presence or without nucleophilic reagents such as water, alcohols, preferably 3-hydroxypropionitrile, a salt of phosphoric acid, preferably bis-tri-n-butylammonium, a salt of pyrophosphoric acid, preferably tris-tetra-n-butylammonium, or condensation is performed on nucleoside derivatives of general formula 16 , where X, Y, Z ³ , Z ⁴ and B ² are as defined above, with polyols, e.g. glycol, erythritol, pentaerythritol, bis (hydroxymethylene) phosphinic acid esters, the condensation being carried out at in aqueous organic solvents in the presence of a non-nucleophilic catalyst, after the condensation is complete, the 2 'and 3' hydroxyl blocking groups and the exoamine groups of the nucleosides are removed.

As protecting groups for the 2 'and 3' hydroxyl groups, known blocking groups are used, preferably selected from the group consisting of acyl, benzoyl, 4,4'-dimethoxytriphenylmethyl, benzyl, trialkylsilyl, preferably trimethylsilyl.

As protecting groups for exoamino groups, known exoamino function blocking groups are used, preferably selected from the group consisting of phenoxyacetyl, isopropoxyacetyl, isobutyryl, benzoyl, (dialkylamino) methylene, (dialkylamino) ethylidene.

As non-nucleophilic condensation catalysts, preferably non-nucleophilic alkoxides such as potassium tert-butoxide, or amines such as imidazole, 1-methylimidazole, 4-dimethylaminopyridine, triethylamine, preferably 1,8-diazabicyclo [5.4.0] undec- 7-en (DBU).

PL 193 592 B1

The reactions are carried out in an anhydrous organic solvent, preferably selected from the group consisting of acetonitrile, methylene chloride, N, N-dimethylformamide, pyridine, dioxane, tetrahydrofuran.

The process according to the invention is a stereoselective process and the less mobile diastereoisomer of the compound of formula 16 is used to obtain the product with the Rp configuration on the phosphorus atom, and the more mobile diastereoisomer of the compound of the formula 16 is used to obtain the product of the Sp configuration.

The process according to the invention is a universal process that can be used for the direct synthesis of dinucleoside polyphosphate analogs of the general formula 1, and it can also advantageously be a stereoselective process allowing to obtain dinucleoside polyphosphate thiophosphate analogs of a specific configuration (Rp or Sp) at each phosphorus atom, ensuring 100% purity isomeric. The method of the invention produces oligonucleoside polyphosphate analogs modified in the polyphosphate chain of the general formula 1, in which Z ¹ represents a hydroxyl group or a hydrogen atom, B ¹ represents adenine, guanine, cytosine, thymine, X and Y represent an oxygen or sulfur atom, R ¹ is a polyol residue, e.g. glycerol, erythritol, pentaerythritol or bis (hydroxymethylene) phosphinic acid of the formulas 2, 3, 4, 5 where Z ² is a hydrogen atom or a phosphoryl or thiophosphoryl or dithiophosphoryl residue or 5'-phosphorylated nucleosides or 5 ' -thiophosphorylated nucleosides or 5'-dithiophosphorylated nucleosides or 3'-phosphorylated nucleosides or 3'-thiophosphorylated nucleosides or 3'-dithiophosphorylated nucleosides.

The process of the invention is illustrated in the following non-limiting examples.

Example I 123

O ¹ , O ² , O ³ -Tri (5'-thiophosphoryladenosyl) glycerol

To a solution of 0.1 mmol of glycerol in 1 ml of acetonitrile, 0.3 mmol of DBU was added and 0.3 mmol of 5'-O- (2-thione-1,3,2-oxathiaphospholane) -N ⁶ , N ⁶ , O ^{2 was added dropwise. ',} O ^3' -tetrabenzoilo-adenosine dissolved in 1.5 ml of acetonitrile. The reaction mixture was left for 5 hours. at room temperature and then concentrated under reduced pressure. 10 ml of a 25% aqueous ammonia solution was added to the residue and left for 12 hours. at room temperature and then concentrated under reduced pressure. The residue was dissolved in 5 ml of water and chromatographed on DEAE Sephadex A-25 with a linear gradient of triethylammonium carbonate from 0.1M to 0.8M. Fractions containing the product were concentrated under reduced pressure to give the product as an amorphous triethylammonium salt powder.

80% yield, ³¹ P NMR (D2O) δ 54 ppm (85% H3PO4 as external standard); MS-MALDI (M-1) m / z 1127.

Example II

O ¹ , O ² , O ³ , O ⁴ -Tetra (5'-thiophosphorythymidyl) pentaerythritol

To a solution of 0.5 mmol of 3'-O-acetylthymidine in 3 ml of methylene chloride, 0.5 mmol of DBU was added, followed by dropwise addition of 0.12 mmol of tetra (2-thione-1,3,2-oxathiaphospholane) pentaerythritol dissolved in 1 ml methylene chloride. The reaction mixture was left for 24 hours. at room temperature and then concentrated under reduced pressure. The crude product was purified by Sephadex A-25 chromatography with a linear gradient from 0.3 to 1 M aqueous triethylammonium carbonate.

45% yield, ³¹ P NMR (D2O) δ 52 ppm; MS-MALDI (M-1) m / z 1416.

Example III

^{O 1} , O ² -bis (5'-O-thiophosphoryladenosylmethylene) phosphinic acid triethylammonium salt

To a solution of 0.5 mmol of bis (hydroxymethylene) phosphinic acid methyl ester in 5 ml of acetonitrile was added 1 mmol of DBU followed by dropwise addition of 1 mmol of 5'-O- (2-thione-1,3,2-oxathiaphospholane) -N ⁶ , N ⁶ , O ^{2 '} , O ^3' -tetrabenzoyl adenosine dissolved in 2 ml of acetonitrile. The reaction mixture was stirred for 5 h. at room temperature, then concentrated under reduced pressure and the residue was dissolved in 10 ml of a 25% aqueous ammonia solution and left for 12 hours. at room temperature to remove benzoyl groups. After removing the ammonia under reduced pressure, the crude product was dissolved in 5 ml of water and chromatographed on DEAE Sephadex with a linear gradient of 0.2 to 0.8 M triethylammonium carbonate to give the product, after drying on a vacuum line, in the form of an amorphous powder.

Yield 80%, ³¹ P NMR δ 59.26 ppm, 44.93 ppm; ³ JPP 34Hz; MS-MALDI (M-1) 815.

Claims (6)

Patent claims 1. Analogs of dinucleoside polyphosphates, modified in the polyphosphate chain, of the general formula 1, where Z ¹ is a hydroxyl group or a hydrogen atom, B ¹ is an adenine, guanine, cytosine, thymine residue, X and Y are an oxygen atom or a sulfur atom, R ¹ represents a polyol residue, preferably glycerol, erythritol, pentaerythritol or bis (hydroxymethylene) phosphinic acid of formulas ₂ 2, 3, 4, 5 where Z ² is a hydrogen atom or a phosphoryl or thiophosphoryl or dithiophosphoryl residue or 5'-phosphorylated nucleosides or 5'-thiophosphorylated nucleosides or 5'-dithiophosphorylated nucleosides or 3'-phosphorylated nucleosides or 3'-phosphorylated nucleosides or 3'-phosphorylated nucleosides or 3'-dithiophosphorylated nucleosides. 2. A method for the preparation of dinucleoside polyphosphate analogs of general formula I, wherein 11 Z ¹ represents a hydroxyl group or a hydrogen atom, B ¹ represents an adenine, guanine, cytosine, thymine residue, X and Y represents an oxygen or sulfur atom, R ¹ represents a polyol residue, e.g. glycerol, erythritol, taerythritol pen ₂ or bis acid ( hydroxymethylene) phosphine of the formulas 2, 3, 4, 5, where Z ² is a hydrogen atom or a phosphoryl or thiophosphoryl or dithiophosphoryl residue or 5'-phosphorylated nucleosides or 5'-thiophosphorylated nucleosides or 5'-dithiophosphorylated nucleosides or 3'-dithiophosphorylated nucleosides or 3 nucleosides 3'-thiophosphorylated nucleosides or 3'-dithiophosphorylated nucleosides, characterized in that the nucleoside of general formula 6 in which Z ³ is a hydrogen atom or a blocking group, Z ⁴ is a blocking group, B ² is an adenine, guanine, cytosine residue of Formulas 7, 8, 9 where Z ⁵ is a hydrogen atom or an exoamine blocking group or a thymine residue of formula 10 is subjected to a condensation reaction with a compound of general formula 11, where X is as defined above, Y is t dichloride or sulfur, R ² is a polyol derivatives, e.g. .: glycerol, erythritol, pentaerythritol, or bis (hydroxymethylene) a phosphine of the formulas 12, 13, 14, 15, wherein X and Y have the abovementioned meaning, in the presence or absence of a nucleophilic reagent such as water, alcohols, preferably 3-hydroxypropionitrile, a salt of phosphoric acid, preferably bis-tri-n-butylammonium, a salt of pyrophosphoric acid, preferably tris-tetra-n-butylammonium, or condensation is carried out on nucleoside derivatives of general formula 16, wherein X , Y, Z ³ , Z ⁴ and B ² are as defined above, with polyols, e.g. glycol, erythritol, pentaerythritol, bis (hydroxymethylene) phosphinic acid esters, whereby condensation is carried out in anhydrous organic solvents in the presence of a non-nucleophilic catalyst, and upon completion of condensation, the 2 'and 3' hydroxyl blocking groups and the exoamino groups of the nucleosides are removed. 3. The method according to p. A method according to claim 2, characterized in that known protecting groups for the 2 'and 3' hydroxyl groups are selected from the group consisting of acyl, benzoyl, 4,4'-dimethoxytriphenylmethyl, benzyl, trialkylsilyl, preferably trimethylsilyl. 4. The method according to p. The process of claim 2, wherein known exoamino function blocking groups are used as protecting groups for exoamino functions, selected from the group consisting of phenoxyacetyl, isopropoxyacetyl, isobutyryl, benzoyl, (dialkylamino) methylene, (dialkylamino) ethylidene. 5. The method according to p. A process as claimed in claim 2, characterized in that non-nucleophilic alkoxides such as potassium tert-butoxide or amines such as imidazole, 1-methylimidazole, 4-dimethylaminopyridine, triethylamine, preferably 1,8-diazabicyclo [5] are used as non-nucleophilic condensation catalysts. 4.0] undec-7-ene (DBU). 6. The method according to p. The process of claim 2, wherein the reactions are carried out in an anhydrous organic solvent selected from the group consisting of acetonitrile, methylene chloride, N, N-dimethylformamide, pyridine, dioxane, tetrahydrofuran.