This application claims priority to Indian patent application No. 2378/CHE/2008 filed on September 29, 2008, the contents of which are incorporated by reference in their entirety.

FIELD OF THE INVENTION

The present invention relates to novel process for the preparation of pharmaceutically acceptable salt of valganciclovir hydrochloride via a protected valganciclovir intermediate prepared by regioselective aminoacylation of ganciclovir.

BACKGROUND OF THE INVENTION

Valcyte (Valganciclovir HC1 tablets) contains Valganciclovir hydrochloride. Valganciclovir hydrochloride is a hydrochloride salt of the L-valyl ester of ganciclovir that exist as a mixture of two diastereomers. Ganciclovir is a synthetic guanine derivative active against cytomegalovirus (CMV).Valganciclovir hydrochloride is a white to off- white crystalline powder with a molecular formula of Ci4H22N605.HCl and a molecular weight of 390. 83. .The chemical name of Valganciclovir hydrochloride is L-valine, 2-(2- amino-1,6-dihydro-6-oxo-purin-9-yl)-methoxy-3-hydroxy-1-propanyl ester monohydro- chloride. The chemical structure of Valganciclovir hydrochloride is represented as formula (1) shown below:

European patent No. 375329 discloses ester prodrugs of Ganciclovir i.e. Valganciclovir and its physiologically acceptable salts thereof having advantageous bioavailability when administered by an oral route. The patent also teaches about the process for the preparation of Valganciclovir.

Valganciclovir is prepared in various ways. PCT Patent application WO 94/ 29311 discloses a process for the preparation of amino esters of a nucleoside analogue, including acyclovir and Ganciclovir. This process comprises reacting a nucleoside analogue having a esterifiable hydroxyl group in its linear or cyclic ether moiety, with a 2-oxa-4-aza-cycloalkane-1,3-dione of formula (a) wherein R1 represent hydrogen, a Cm alkyl or alkenyl group or other amino acid side chain, and R2 represent hydrogen or a group COOR3 wherein R3 is a benzyl, t-butyl, fluorenylmethyl or an optionally halo substituted linear or branched Cu8 alkyl group. Preferred R] groups include hydrogen, methyl, iso-propyl and isobutyl, yielding respectively the glycine, alanine, valine and isoleucine esters of acyclovir or ganciclovir. Examples 1-3 of the PCT patent application WO 94/029311 discloses only the condensation of acyclovir with the valine-substituted 2-oxa-4-aza-cycloalkane-1,3-dione (Z-valine-N-carboxy anhydride) by conventional procedures. While the amino acid esters of the PCT application include both the acyclovir and Ganciclovir esters, the application does not disclose how to prepare the Ganciclovir esters, much less the mono-esters of Ganciclovir.

WO 1997/27194 describes process for the preparation of Valganciclovir. The process includes the reaction of Ganciclovir with N-Boc-Valine-NCA in the presence of a silylating agent and a base and maintaining for 72 hours at lower temperature gives 2-(2- amino-1,6-dihydro-6-oxo-purin-9-yl)-methoxy-3-hydroxy-1 -propyl-N- (benzyloxycarbonyl)-L-valinate i.e. protected Valganciclovir. This process takes more time for completion of reaction.

U.S. Patent Nos. 5,700,936, 5,756,736, 5,840,890, 5,856,481 discloses process for the preparation of Valganciclovir, which involve divalinate, N,0-bistrityl, monocarboxylate-monovalinate, bis(L-valinate) intermediates and U.S. Patent. Nos. 6,040,446, 6,215,017 & 6,218,568, involve persilyl guanine or glycerol derivatives as intermediates.

The present invention relates to novel process for the preparation of Valganciclovir via N-benzoyloxy carbonyl Valganciclovir intermediate using regioselective enzymatic aminoacylation of Ganciclovir. The process for the preparation of N-benzoyloxy carbonyl Valganciclovir according to the present invention is eco- friendly, safe and cost effective. In this process enzyme can be reused several times and the reaction can be performed at ambient temperature. In commercial scale it does not require anhydrous conditions.

OBJECT OF THE INVENTION

The main object of the present invention is to provide a novel process for the preparation of pharmaceutically acceptable salt of Valganciclovir from protected Valganciclovir prepared by regioselective enzymatic aminoacylation of Ganciclovir.

SUMMARY OF THE INVENTION

The main aspect of the present invention is to provide a novel process for the preparation of pharmaceutically acceptable salt of Valganciclovir from protected Valganciclovir formula (4), prepared by regioselective enzymatic aminoacylation of Ganciclovir.
DETAILED DESCRIPTION OF THE INVENTION

The main aspect of the present invention relates to novel process for the preparation of pharmaceutically acceptable salt of Valganciclovir from protected Valganciclovir of formula (4), prepared by regioselective enzymatic aminoacylation of Ganciclovir.

In one embodiment, the present invention encompasses the process for the preparation of pharmaceutically acceptable salt of Valganciclovir comprising the steps of:

a. dissolving ganciclovir in a suitable solvent,

b. adding Z-valine NCA and an enzyme,

c. isolating the protected Valganciclovir, and

d. deprotecting the protected valganciclovir in presence of a catalyst and suitable solvent, and

e. converting valganciclovir to its pharmaceutically acceptable salt.

In another embodiment, the process for the preparation of pharmaceutically acceptable salt of Valganciclovir is as shown in scheme-I

Scheme I

According to the present invention, ganciclovir is dissolved in a suitable solvent and treated with Z-valine NCA and enzyme and maintained till the completion of the reaction monitored by HPLC. The reaction mixture is filtered and the solvent is evaporated. The resultant mixture is dissolved in a chlorinated solvent and treated with dilute hydrochloric acid. Separating the layers and bringing the pH of the solution to acidic precipitates the protected valganciclovir. The protected valganciclovir is dissolved in suitable solvent,and treated with the suitable organic acid. The resultant mixture is maintained and treated with palladium on carbon followed by purging of hydrogen gas. Distilling out the solvent and adding mixture of water and alcoholic solvent such as isopropyl alcohol and cooling the reaction mixture to lower temperature and maintained till solid is seperated. Filtering the separated mass and dissolving the solid in alcoholic solvent followed by distillation leaves the residue which is again dissolved in a mixture of ester solvent such as ethyl acetate and hydrocarbon such as cyclohexane and distilled out thus giving the desired product, pharmaceutically acceptable salt of valganciclovir.

According to the present invention, the solvent for dissolution of ganciclovir is selected from water, ketones such as acetone, methyl isobutyl ketone, methyl ethyl ketone and cyclohexanone; alcohols such as methanol, ethanol, isopropanol, n-propanol, n- butanol, tertiary-butyl alcohol, cyclohexanol; chlorinated solvents such as dichloromethane, chloroform, carbon tetrachloride; hydrocarbon solvents such as toluene, xylene, n-hexane, n-heptane, cyclohexane; esters such as ethyl acetate, n-propyl acetate, n-butyl acetate, t-butyl acetate; ethers such as diethyl ether, dimethyl ether, diisopropyl ether nitriles such as acetonitrile, propionitrile; or polar aprotic solvents like dimethyl sulfoxide, dimethyl formamide and dioxane or a mixture of thereof. More preferably solvents are dimethyl sulfoxide, dimethyl formamide or a mixture thereof.

According to the present invention, the enzyme is selected from the group of (Hydrolase's) preferably Lipozymes, Porcine Pancreatic Lipase, CAL-A, lyophilized Candida lipolytica Lipase, Geotrichum candidum Lipase, Pseudomonas aroginosa Lipase, Aspergillus niger Lipase, Pseudomonas cepacia Lipase, Pseudomonas fluorescens Lipase, Candida rugosa Lipase, Rhizopus delemar Lipase, Rhizopus oryzae Lipase, Penicillium camembertii Lipase, Penicillium camembertii Lipase, Mucor javanicus Lipase, Penicillium roqueforti Lipase, Pseudomonas cepacia Lipase, CAL-B, lyophilized microbial, lyophilized Lipase, Thermomyces sp. Lipase Alcaligines sp., Chromobacterium viscosum Lipase,Candida utilis Lipase,Rhizopus niveus Lipase,Pseudomonas sp. Lipoprotein Lipase Shermomuces lanuginosus Lipase, Rhizomucor miehei Lipase,Pseudomonas species Lipase, Wheat Germ Lipase,Rhizopus
arrhizus Lipase,Pancreatic Lipase 250, Candida antarctica lipase B {Novozyme-435), preferably Candida antarctica lipase B {Novozyme-435) or lypozyme.

According to the present invention, the recovered enzyme can be reused thus making the invention economical.

According to the present invention, the chlorinated solvent is selected from dichloromethane, chloroform, carbon tetrachloride preferably dichloromethane.

According to the present invention, the solvent used in the deprotection step is selected from methanol, ethanol, isopropyl alcohol, ethyl acetate,cyclohexane ,water or mixtures thereof, preferably methanol, and the catalyst is selected from Palladium, platinum, Raney nickel, preferably palladium on charcoal. The hydrogenation is carried out under hydrogen pressure.

According to the present invention, the pharmaceutically acceptable salt is selected from hydrochloric acid, sulphuric acid, phospharic, maleic, fumaric, citric, tartaric, preferably hydrochloric acid.

The following non-limiting examples illustrate specific embodiments of the present invention. They should not construe it as limiting the scope of present invention in any way.

Examples: Example 1
Ganciclovir (1 g) was dissolved in 5 ml solvent (20% dimethyl sulfoxide /80% dimethyl formamide) by stirring at room temperature, then molecular sieve 4°A (1 g) was added. Novozyme-435 (0.5 g) and Z-Valine NCA (1.2 g) was added and the resulting mixture was stirred at room temperature and the reaction was periodically monitored by HPLC. Reaction was stopped after 28 hours. HPLC analysis showed N-Benzoyloxy carbonyl valganciclovir 4 (46 %), 2(42 %) and 5 (10%). The enzyme was removed by filtration, and the solvent was evaporated with a rotary evaporated at 40° C. The residue was treated with methylenedichloride (40ml) and dil HC1 and the organic and aqueous layer was separated. The organic layers containing compound 5 are combined, dried and concentrated. The N-Benzoyloxy carbonyl valganciclovir 4 was precipitated out at pH 5 from aqueous layer. The solid was filtered and dried.

Example 2
Ganciclovir (lg,) was dissolved in 50 ml solvent (20% dimethyl sulfoxide /80% dimethyl formamide) by stirring at 45° C, then molecular sieve 4 °A (lg) was added. Novozyme- 435 (0.5 g) and Z-Valine NCA (1.2 g) was added and the resulting mixture was stirred at 45°C and the reaction was monitored by HPLC. Reaction was stopped after 24 hours. HPLC analysis showed N-benzoyl ox y carbonyl valganciclovir 4 (48 %), 2 (25 %) and 5 (25.5%). The enzyme was removed by filtration, and the solvent was evaporated with a rotary evaporated at 40 °C. The residue was treated with methylenedichloride (40 ml) and dil HC1 and the organic and aqueous layer was separated. The organic layers containing compound 5 are combined, dried and concentrated. The A-Benzoyloxy carbonyl valganciclovir 4 was precipitated out at pH 5 from aqueous layer. The solid was filtered and dried.

Example 3

Ganciclovir (lg) was dissolved in 50 ml solvent (20% dimethyl sulfoxide /80% dimethyl formamide) by stirring at room temperature. Novozyme-435 (0.5 g) and Z-Valine NCA (1.2 g) was added and the resulting mixture was stirred at room temperature and the reaction was monitored by HPLC. Reaction was stopped after 60 h. HPLC analysis showed JV-Benzoyloxy carbonyl valganciclovir 4 (33.54%), 2 (60 %) and 5 (4.39%). The enzyme was removed by filtration, and the solvent was evaporated with a rotary evaporated at 40 °C. The residue was treated with methylenedichloride (40ml) and dil HC1 and the organic and aqueous layer was separated. The organic layers containing compound 5 are combined, dried and concentrated. The A-benzoyloxy carbonyl valganciclovir 4 was precipitated out at pH 5 from aqueous layer. The solid was filtered and dried.

Example 4

Ganciclovir (1 gram) was dissolved in 50mL solvent (20% dimethyl sulfoxide /80% dimethyl formamide) by stirring at 40 °C. Lipozyme (0.25 g) and Z-Valine NCA 2 (1.2 g, 4.3 mM) was added and the resulting mixture was stirred at 40 °C and the reaction was monitored by HPLC. Reaction was stopped after 24 hours. HPLC analysis showed N- Benzoyloxy carbonyl valganciclovir 4 (48%), 2 (29 %) and 5 (22%). The enzyme was removed by filtration, and the solvent was evaporated with a rotary evaporated at 40 °C.
and aqueous layer was separated. The organic layers containing compound 5 are combined, dried and concentrated. The N-benzoyloxy carbonyl valganciclovir 4 was precipitated out at pH 5 from aqueous, layer. The solid was filtered and dried. Example 5
N-benzoyloxy carbonyl valganciclovir 4(lgm) was dissolved in methanol (10 ml) at room temperature, added DM water (5 ml) and dil HC1 at room temperature. The resulting mixture was stirred for 2 to 3 hours till a clear solution was obtained. To the clear solution added Pd/C at room temperature. Purged H2 gas at 25 to 35°C, after completion of the reaction, filtered the resulting mixture through hiflow bed & washed with methanol (10 ml). Distilled off methanol completely under vacuum at <50°C and added DM water to the residue & raised the temperature. The resulting mixture was dissolved in isopropyl alcohol and stirred for 2 to 3 hours. Cooled the reaction mixture to 5 to 10°C and stirred for 2 hours. Filtered & washed with chilled isopropyl alcohol and water. The wet mass was dissolved in methanol to get a clear solution. Distilled off methanol completely under vacuum at <50°C, to the residue was added mixture of ethyl acetate and cyclohexane. Isolated valganciclovir hydrochloride and dried the wet mass under vacumm at 50 °C.

We claim

1. A process for the preparation of pharmaceutically acceptable salt of valganciclovir by regioselective enzymatic aminoacylation comprising the steps of:

a. dissolving ganciclovir in a suitable solvent,

b. adding Z-valine NCA and an enzyme,

c. isolating the protected valganciclovir,

d. deprotecting the protected valganciclovir in presence of a catalyst and suitable solvent, and

e. converting valganciclovir to its pharmaceutically acceptable salt.

2. The process according to claim 1, wherein the solvent for dissolution of galciclovir is selected from water, ketones such as acetone, methyl isobutyl ketone, methyl ethyl ketone and cyclohexanone; alcohols such as methanol, ethanol, isopropanol, n-propanol, n- butanol, tertiary-butyl alcohol, cyclohexanol; chlorinated solvents such as dichloromethane, chloroform, carbon tetrachloride; hydrocarbon solvents such as toluene, xylene, n-hexane, n-heptane, cyclohexane; esters such as ethyl acetate, n-propyl acetate, n-butyl acetate, t-butyl acetate; ethers such as diethyl ether, dimethyl ether, diisopropyl ether nitriles such as acetonitrile, propionitrile; or polar aprotic solvents like dimethyl sulfoxide, dimethyl formamide and dioxane or a mixture of thereof. More preferably solvents are dimethyl sulfoxide, dimethyl formamide or a mixture thereof.

3. The process according to claim 2, wherein the solvent is a mixture of dimethyl sulfoxide and dimethyl formamide.

4. The process according to claim 1, wherein the enzyme is selected from the group of (Hydrolase's) are Lipozymes, Porcine Pancreatic Lipase, Candida antarctica lipase A (CAL-A), lyophilized Candida lipolytica Lipase, Geotrichum candidum Lipase, Pseudomonas aroginosa Lipase, Aspergillus niger Lipase, Pseudomonas cepacia Lipase, Pseudomonas fluorescens Lipase, Candida rugosa Lipase, Rhizopus dele mar Lipase, Rhizopus oryzae Lipase, Penicillium camembertii Lipase, Penicillium camembert Lipase, Mucor javanicus Lipase, Penicillium roqueforti Lipase, Pseudomonas cepacia Lipase, Candida antarctica lipase B (CAL-B), lyophilized microbial, lyophilized Lipase, Thermomyces sp. Lipase A Icaligines sp., Chromobacterium viscosum Lipase,Candida utilis Lipase, Rhizopus niveus Lipase, Pseudomonas sp. Lipoprotein Lipase, Thermomuces lanuginosus Lipase, Rhizomucor mxehei Lipase,Pseudomonas species Lipase, Wheat Germ Lipase, Rhizopus arrhizus Lipase, Pancreatic Lipase 250, Candida antarctica lipase B {Novozyme-435).

5. The process according to claim 4, wherein the enzyme is Candida antarctica lipase B (Novozyme-435) or Lypozyme.

6. The process according to claim 1, wherein the protected valganciclovir is precipitated at about pH 4.5 to 5.5.

7. The process according to claim 1, wherein the solvent in the deprotection step is selected from methanol, ethanol, isopropyl alcohol, ethyl acetate, cyclohexane, water or mixtures thereof.

8. The process according to claim 1, wherein the catalyst is selected from Palladium, platinum or Raney nickel.

9. The process according to claim 1, wherein the pharmaceutically acceptable salt is selected from hydrochloric acid, sulphuric acid, phosphoric acid, maleic acid, fumade acid , citric acid or tartaric acid

10. The process according to claim 9, wherein the salt is hydrochloric acid.