DD290330A7 - P-CYTIDINE-5'-DIPHOSPHORSAEURE ALKYLESTER AND METHOD FOR THE PRODUCTION THEREOF - Google Patents

Abstract

The invention relates to a process for the preparation of P-cytidine-5-diphosphorsÄure alkyl ester of the general formula I and their salts, in which R is an alkyl radical having 10-18 C-atoms. The aim is to develop a process for producing these compounds. The compounds are prepared by reacting alkyl phosphates or their salts with cytidine-5-monophosphate or an activated derivative thereof. Formula I

Description

ί-%

in which R is an alkyl radical having 10-18 C atoms, and salts thereof, characterized in that alkyl phosphates of the general formula II or their salts,

O u ROP-OH - υ

OH

in which R has the abovementioned meaning, as such or in the form of the salts with cytidine-5'-monophosphate or an activated derivative of this compound in the presence of an organic base in the ratio 1: 0.6 to 1.8 at 20 ⁰ C to 6O ⁰ C condenses, the resulting condensation product is usually isolated and optionally converted into a salt.

2. P-Cytidine-δ'-diphosphoric acid hexadecyl ester.

3. P-Cytidine-B'-diphosphoric acid dodecyl ester.

Field of application of the invention

The invention relates to a process for preparing P-cytidine-B'-diphosphorsäure-alkyl ester of general formula I, in which R is an alkyl radical with 10-18 C-Atomn, and of their salts. These are novel derivatives of cytidine 5'-diphosphate. Field of application of the invention is the chemical or pharmaceutical industry.

0 0

Il

R-O-P-O-P-O-CH

ο o

Characteristics of the known state? of the technique

Condensations of nucleoside phosphates with phosphatidic acids are known. Thus, for example, React cytidine C monophosphate morpholide with diacylglycerophosphoric acid to form cytidine 5'-diphosphate diglycerides (B. Agranoff et al., Biochem., Prep., 10 (1963) 47).

Cytosine arabinoside acylglycerol adducts have also been synthesized in an analogous manner (S. Bottka et al., Tetrahedron, 35 (1979) 2909). Processes for the preparation of compounds of the general formula I type are hitherto unknown.

Object of the invention

It is an object of the invention to develop a process for the preparation of the compounds of general formula I.

Explanation of the essence of the invention

The object of the invention is to develop a method for the production of new physiologically active derivatives of the cytildin type as potential cytostatics.

According to the invention, the P-Cytidln-B'-diphosphorsäure-alkyl esters of general formula I are prepared by alkylphosphato of the general formula II, in which R is as defined above

O II R-O-P-OH Il

I oh

has reacted with cytidine-o'-monophosphate or an activated derivative of dloser compound. The alkylphosphato are dissolved several times in anhydrous pyridine (dried over CaH ₂ ) and the solutions are each evaporated in vacuo to dryness in order to remove traces of moisture. If the corresponding morpholidate, which is usually present as a 4-morpholine-N, N'-cyclohexylcarboxamidine salt, is used for the reaction, it is expedient to use this several times with toluene in order to remove any water of crystallization and adhering moisture To stamp the vacuum. Subsequently, both reaction components are combined. The reaction mixture is re-evaporated with pyridine, taken up in pyridine and stirred under exclusion of atmospheric moisture until the reaction is complete. The molar ratio between alkyl phosphate and nucleoside component is 1: 0.6 to 1.8. The reaction temperature is between 20 ⁰ C and 60 ⁰ C, preferably at 35 ⁰ C to 40 ⁰ C. The reaction time is generally between 24 hours and 10 days. If the reaction component used in the deficit can no longer be detected chromatographically, the reaction mixture is worked up in the customary manner. It is concentrated to dryness in vacuo, the residue obtained is re-rotated several times with toluene to remove pyridine and then taken up in a mixture of chloroform-methanol 0.01 N HCl. The aqueous phase is separated off and extracted again several times with chloroform-methanol. After reextraction of the combined organic phases with water, they are concentrated to dryness in vacuo. The resulting free acid I is converted by treatment with aqueous methanolic ammonia in the diammonium salt, which has good water solubility.

The compounds obtained have a structural relationship with the natural cytidine S'-diphosphate diacylglycerols and their alkylglycerol analogue, but differ from them in that instead of the acyl (or alkyglycerol component) an alkanol component is present in the molecule New compounds are also physiologically active, causing, for example, a concentration-dependent proliferation homing of Ehrlich ascites tumor cells (see Table 1).

Table 1 Inhibition of growth of Ehrlich ascites tumor cells (EAT) as well as colony formation of mouse bone marrow (KM) connection

[Inhibition at a βοηζΚμΜ 3 10

30

P-Cytidine-ö'-diphosphoric acid hexadecyl ester P-Cytidine-ö'-diphosphoric dodecyl ester

EAT 16 38 67 97 KM 0 0 29 69 EAT 22 35 70 98 KM 0 0 18 62

example 1

P-Cytlyl-S'-diphosphoric acid hexadecyl ester I (R hexadecyl phosphate II (R = C ₁₈ H ₃₃ ) (150 mq, 0.47 mmol) is taken up four times in about 5 ml pyridine (dried over CaH ₁ ) and the solution each concentrated to dryness in vacuo.

Cytidine 5'-morpholidophosphate (4-morpholine-N, N'-dicyclohexylcarboxamidine salt) (404.2 mg, 0.58 mmol) is triturated four times with 4 ml of toluene and the mixture is rotary evaporated to dryness in vacuo. Thereafter, the reaction components are combined, concentrated again with 5 ml of pyridine and taken up in about 10 ml of pyridine. The solution is stirred for 5 days at 37 ° C under exclusion of atmospheric moisture. Subsequently, the reaction mixture is concentrated to dryness in vacuo. To the residue is added toluene (3x 5 ml), which is distilled off in vacuo. The resulting substance is taken up in a mixture of 15 ml of chloroform-methanol (1: 1, v / v) and 7 ml of 0.01N HCl. The aqueous phase is separated and extracted three times with about 5 ml of chloroform-methanol. The combined organic phases are washed neutral with a little water and concentrated to dryness. The residue is dissolved in a mixture of 45 ml of chloroform, 20 ml of methanol, 9 ml of water, 0.3 ml of 1N methanol. Treated ammonia, the organic phase is separated, extracted again with 5 ml of water. Concentration of the combined aqueous phases gives compound I as diammonium, which is reprecipitated from chloroform-acetone for further purification. The yield is 144 mg. Thin Layer Chromatography (Merck Finished Plate Kieselgel 60 F254): RfO, 31 (chloroform-methanol-glacial acetic acid-water, 25: 15: 2: 4, v / v / v / v). UV absorption : A _m " 272.6 nm (water), X _mtx 282 (0.01 N HCl).

Example 2 P-Cytidine-δ-diphosphoric acid dodecyl ester I (R = C) ₂ H ₂ S) A mixture of dodecyl phosphate II (R = C ₁₂ H ₂ S) (0.25 g, 1 mmol), cytidine-o'-morpholidophosphate (0.859 g, 1.25 mmol) and

dried pyridine (5 ml) is evaporated in vacuo to dryness. The residue is again taken up in 5 ml of pyridine.

The mixture is stirred for 5 days at 37 ⁰ C under exclusion of atmospheric moisture and then concentrated to dryness. The Residue is dissolved in 30 ml of chloroform / methanol / 0.01 N HCl (2: 1: 1, v / v / v). The organic phase is separated and the

aqueous phase several times with 10 ml of chloroform / methanol (2: 1, v / v) extracted. The combined organic phases are after

Enriched addition of methanolic ammonia and the residue purified by reprecipitation from CHCl ₃ / acetone. Yield: 0.177 g (DI-NH ₄ -SaIz), 30%. Thin Layer Chromatography (Merck Finished Plate Kieselgel 60): Ri = O, 32 (chloroform-methanol-glacial acetic acid-water, 25: 15: 2: 4,

Claims (1)

1. A process for the preparation of P-Cytldln-B'-dlphosphorsäurealkylester of general formula I, HH, ANK OO Λ-ν _Μ κ Ii η ^/ IV ROPOPO-CH ₂ ο "ο- - °