CLIAMS:We Claim:

1. An improved process for the preparation of a pure key intermediate of regadenoson, compound of formula II

Formula II
, which comprises:

i) reaction of 1, 2, 3, 5-tetra-O-acetyl ribofuranose with 2-chloroadenine using stannic chloride in presence of nitrile solvent to provide condensed compound of formula III; and

ii) reaction of the condensed compound of formula III

Formula III

with ammonia in presence of alcohol to provide pure compound of 2-chloroadenosine of formula II.

2. The process of claim 1, wherein said nitrile of step i) is selected from acetonitrile, propionitrile, butyronitrile and acrylonitrile.

3. The process of claim 1, wherein said nitrile of step i) is acetonitrile.

4. The purity of condensed compound of Formula III of claim 1 is greater than or equal to 98% by HPLC.

5. The process of claim 1, wherein the alcohol of step ii) is methanol.

6. The process of claim 1, wherein the ammonia is aqueous solution of ammonia.

7. The process of claim 1, in which the reaction of step ii) is performed at elevated temperature of about 45 °C to about 65 °C.

8. The purity of 2-chloroadenosine of claim 1 is greater than or equal to 98 % by HPLC.

Dated this 28th day of March, 2013 For Wockhardt Limited

(Dr Mandar Kodgule)
Authorized Signatory
,TagSPECI:DESCRIPTION

The present invention provides an improved process for the preparation Key intermediate of regadenoson, the compound of formula II.

Formula II

Regadenoson is chemically known as adenosine, 2-[4-[(methylamino)carbonyl]-1H-pyrazol-1-yl]-, monohydrate. Its structural formula is depicted as Formula I:

Formula I
Regadenoson is a selective A 2 A-adenosine receptor agonist that is a coronary vasodilator. It is currently marketed in the form of a monohydrate as a pharmacologic stress agent indicated for radionuclide myocardial perfusion imaging (MPI) in patients unable to undergo adequate exercise stress.

U.S. Patent Nos. 6,403,567 and 7,183,264 discloses Regadenoson and synthesis of regadenoson. The syntheses disclosed are multi-step processes that proceed via 2-hydrazinoadenosine, which is prepared from the corresponding iodo-derivative (2-iodoadenosine). However, the use of 2-iodoadenosine is not well suited for large-scale syntheses.

U.S. Patent No. 6,252,061 discloses a process for the preparation of intermediate of regadenoson, chloroadenosine, from diaminopurine ribonucleoside using antimony trichloride (SbCl3) in presence of DMSO, dichloroethane and t-butyl nitrite. However, this process involves expensive, hazardous and unsafe reagents, which are not well suited for industrial scale up process.

Various other prior arts, for example, WO 2003/035012, Journal of Organic Chemistry, 71(20), 7773-7779; 2006, Journal of Medicinal Chemistry, 35(24), 4557-61; 1992, Journal of Organic Chemistry, 67(3), 790-802; 2002, provides different process for producing the compound of formula II. However, those processes are involves use of numbers of steps and expensive reagents and not suited for industrial scale up.

Hence, there is a need to develop an improved process for the preparation of pure compound of formula II that is lower cost alternative, simple and industrially applicable, result in high yield and high purity of compound of Formula II as well as pharmaceutically acceptable salt thereof.

The compound of Formula II is a key intermediate for the preparation of regadenoson. The chloroadenosine of Formula II is also well suited and it can be easily handled for the preparation of 2-hydrazinoadenosine.

The present inventors found well suited reaction conditions for preparing the compound of Formula II from 1, 2, 3, 5-tetra-O-acetyl ribofuranose that provided a faster rate of reaction to achieve higher yield and greater purity of compound of formula II.

In an aspect of the present invention provides an improved process for the preparation of a pure key intermediate of regadenoson, compound of formula II

Formula II
, which comprises:
i) reaction of 1, 2, 3, 5-tetra-O-acetyl ribofuranose with 2-chloroadenine using stannic chloride in presence of nitrile solvent to provide condensed compound of formula III; and
ii) reaction of the condensed compound of formula III

Formula III

with ammonia in presence of alcohol to provide pure compound of 2-chloroadenosine of formula II.

The step i) involves reaction of 1, 2, 3, 5-tetra-O-acetyl ribofuranose with 2-chloroadenine using stannic chloride in presence of nitrile solvent to provide condensed compound of formula III.

The suitable nitrile solvents for the reaction include and are not limited to acetonitrile, propionitrile, butyronitrile, acrylonitrile and the like.
The use of stannic chloride for the step i) enhances the reaction to complete the starting material in a shorter time. Further, the stannic chloride is cheap, easily available and safer reagent for the handling and industrial purpose.

In an embodiment, the use of stannic chloride in presence of acetonitrile at a ambient temperature, for example, 25 to 35 °C provides greater purity and yield of condensed product of formula III.

The reaction is performed at a temperature of about 20 to about 40 °C, or at about 25 to about 30 °C. The reaction may be stirred for a period of about 3 to 6 hours or more without increasing the formation of impurities.

After completion of the reaction, the reaction mixture is quenched with quenching agent, for example, water, organic solvent, acid or combination thereof. The organic solvent includes chlorinated solvent such as dichloromethane, dichloroethane, chloroform and the like; esters such as ethyl acetate, isopropyl acetate, 2-methoxyethyl acetate and the like. The acid includes hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid and the like.

The organic layer of resultant reaction mixture may be separated and subjected for precipitation of the solid using suitable techniques such as concentration, for example, buchi distillation and spray drying; anti solvent technique using suitable alcohol solvent or recrystallization using alcohol solvent. The suitable alcohol solvent is selected from methanol, ethanol, isopropyl alcohol, n-butanol and the like.

The resultant condensed product of formula III, Acetic acid 3, 4-diacetoxy-5-(6-amino-2-chloro-purin-9-yl)-tetrahydro-furan-2-ylmethyl ester, may have purity greater than or equal to 98%.

The yield of the condensed product of formula III obtained from the present invention is greater than or equal to 90 %.

The step ii) involves reaction of the condensed compound of formula III with ammonia in presence of alcohol to provide pure compound of 2-chloroadenosine of formula II.

The reaction is conducted in presence of alcohol such as methanol, ethanol, isopropyl alcohol, n-butanol and the like.

The condensed product of formula III obtained from the step i) of the present invention is treated with any source of ammonia includes aqueous solution of ammonia at a temperature of about 25 to about 70 °C or at about 45-65°C. The reaction may be maintained at the same temperature for certain period of time, for example, 3-4 hours.

The resultant product from the present invention provides greater yield and purity, for example, at least 90 %.

In another aspect, the present invention provides a process for the preparation of 2-chloroadenosine having purity greater than or equal to 98 % by HPLC, which comprise reaction of the compound of formula III

Formula III

with ammonia in presence of alcohol at an elevated temperature.

The reaction is conducted in presence of alcohol such as methanol, ethanol, isopropyl alcohol, n-butanol and the like.

The compound of formula III is reacted with any source of ammonia includes aqueous solution of ammonia. The suitable elevated temperature for the reaction is about 40 to about 70 °C or 50 to about 65 °C. The reaction may be maintained at the same temperature for certain period of time, for example, 3-4 hours.

After completion of the reaction, the reaction mixture may be concentrated to provide suspension and then cooled to enhance the yield of precipitated product of Formula II (2-chloroadenosine).

The isolated product may be recovered by using suitable technique such as filtration by vacuum, filtration by gravity, centrifugation, decantation, and the like.

The resultant solid optionally dried at a suitable temperature of about 30 to about 50 ° or more for a period of 1 hour to 3 hours or more to stabilize the compound.

The purity of 2-chloroadenosine obtained from the present invention may be greater than 98 % determined by HPLC method.

The present invention is further illustrated by the following example, which does not limit the scope of the invention. Certain modifications and equivalents will be apparent to those skilled in the art and are intended to be included within the scope of the present application.

EXAMPLES

Example -1: Preparation of Chloroadenosine

Stage- I: process for condensed product (Acetic acid 3, 4-diacetoxy-5-(6-amino-2-chloro-purin-9-yl)-tetrahydro-furan-2-ylmethyl ester)

To a stirred solution of 1, 2, 3, 5-tetra-o-acetyl ribofuranose (100 g) and 2-chloroadenine (60 g) in acetonitrile (400 ml) was added stannic chloride (150 g). The reaction mixture was stirred at ambient temperature for 6 hours and after wards quenched in a mixture of water and dichloromethane. The pH of the reaction mixture was carefully adjusted to ~ 12 and layers were separated. The DCM layer was washed with water concentrated under vacuum and the residue was re-crystallized with methanol to get the desired product.

Yield: 88 g
Purity: 98.5% by HPLC

Stage- II: 2-chloroadenosine

Condensed product (50 g) was suspended in aqueous ammonia solution and then heated for 4 hours at a temperature of about 50-60 °C. The ammonia solution was partially distilled under vacuum and the resulting slurry obtained after cooling was filtered and dried under vacuum to get the desired product.

Yield: 32.5 g
HPLC purity: 99.5% by HPLC