DESC:Field of Invention:

The present invention relates to an efficient industrial stereoselective process for the preparation of (9-[(R)-2-[[(S)-[[(S)-l- (isopropoxycarbonyl)ethyl]amino]phenoxyphosphinyl]methoxy]propyl] adenine and its salts.

Background and Prior Art:

9-[(R)-2-[[(S)-[[(S)-l-(isopropoxycarbonyl)ethyl]amino]phenoxyphosphinyl]methoxy]propyl] adenine, an isopropylalaninyl monoamidate phenyl monoester, is a prodrug of (R)-9-(2-phosphonomethoxypropyl)adenine.

The process for preparation of the 9-[(R)-2-[[(S)-[[(S)-l- (isopropoxycarbonyl)ethyl]amino] phenoxyphosphinyl]methoxy]propyl]adenine fumarate salt was first disclosed in the patent publication WO2002008241 which involves the reaction of monophenyl PMPA (wherein PMPA is 9-[2-(phosphonomethoxy)propyl]adenine) with thionyl chloride and further reacting with (L)-a aminobutyric acid ethyl ester. In this publication, separation of diastereomers is carried out by chromatography techniques which are batch elution chromatography, simulated bed chromatography and C18 Reverse phase high performance liquid chromatography.

WO2013052094 describes process of selective crystallization of 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino}phenoxyphosphinyl)methoxy]propyl}adenine using crystallization-induced dynamic resolution using seeds of 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino}phenoxyphosphinyl)methoxy]propyl}adenine.

WO2013025788 describes 9-[(R)-2-[[(S)-[[(S)-l- (isopropoxycarbonyl)ethyl]amino] phenoxyphosphinyl]methoxy]propyl]adenine hemifumarate; pharmaceutical compositions comprising the same and antiviral therapy using 9-[(R)-2-[[(S)-[[(S)-l- (isopropoxycarbonyl)ethyl]amino] phenoxyphosphinyl]methoxy]propyl]adenine hemifumarate.

Use of chromatographic techniques for the separation of diastereomers at the industrial scale requires a remarkable starting investment. Building the set-up for chromatography is an expensive process. Further the separation of diastereomers by chromatography is a tedious and time-consuming process. The diastereoisomers may be alternatively separated by resolving agents or through salt formation. Also large volume of solvent is required for these separation techniques which not only increases the overall cost but also environment unfriendly.

Thus there is need in the art to providea process for preparation of (9-[(R)-2-[[(S)-[[(S)-l- (isopropoxycarbonyl)ethyl] amino]phenoxyphosphinyl]methoxy]propyl]adenine and its salts with high diastereomeric purity and higher yield at the same time the process should be environment friendly.

Object of the Invention:

The main object of the present invention is to provide a simple, efficient and industrially feasible process for the preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine and its salts with high purity and high yield.

Another object of the present invention is to provide a stereoselctive process to obtain 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine and its salts with high diastereomeric purity.

Yet another object of the present invention is to provide novel intermediate useful in preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine with high diastereomeric purity.

Detailed Description of the Invention:

The invention will now be described in detail in connection with various embodiments, so that various aspects thereof may be more fully understood and appreciated.

The present invention provides a stereoselective process for the preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine and its salts with high diastereomeric purity that is cost-effective, environment friendly and industrially suitable process in comparison with the up-to-now known methods.

In the attempt to prepare a simple and efficient method for preparation of 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino}phenoxyphosphinyl)methoxy]propyl}adenine, the present inventors have found that use of silylating agent has very high success rate that provides higher diastereomeric excess and high yield.

According to a first aspect of the present invention, there is provided a process for the preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine and its salts which comprises,
a. reacting monophenyl PMPA (III) with silylating agent in presence of halogenating agent in a solvent;
b. treating step a with silylated L-alanine isopropyl ester (II-a) in presence of base;

Wherein Ak is an alkyl chain of C1-C4 carbon atoms.

c. isolating 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine (IV);
d. optionally isolating 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine salt; and
e. optionally purifying the product of step c and/or d.

Monophenyl PMPA (III), the starting material used in the invention may be prepared using known methods, for example as disclosed in WO2002008241A1.

The silylating agent used in the process of present invention may be selected from alkylsilane preferably trialkylhalosilane which may be selected from the group consisting of triethylsilylchloride, trimethylsilylchloride, triisoprpylsilylchloride and t-butyldimethylsilylchloride. Most preferably, the silylating agent is trimethylsilylchloride.

The silylating agent may be used in molar ratio between 0.1 moles to 0.5 mole, more preferably 0.3 moles, most preferably in a catalytic amounts.

The halogenating agent may be selected from the group consisting of thionyl chloride, oxalyl chloride, phosphorous trichloride, phosphorous tribromide.

The solvent may be a polar aprotic solvent selected from the group consisting of ethyl acetate, methyl acetate, propyl acetate, isopropyl acetate, diethyl ether, diisopropyl ether, dimethyl formamide, tetrahydrofuran, dichloromethane, acetone, methyl ethyl ketone, methyl ter/-butylether, toluene, or acetonitrile, or a mixture thereof.

The base may be an organic base, selected from the group consisting of dimethylamine, triethyl amine, 4-(Dimethylamino)pyridine, N-Ethyldiisopropylamine, Tetramethylammonium hydroxide.

The addition of silylated L-alanine isopropyl ester (II) is carried out in pH range between 7-10, more preferably between pH 7-8.

The reaction is carried out at a temperature of 15-50°C preferably 20-30°C most preferably 25- 30°C.

In another aspect of the invention, isolation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine is carried out directly by general methods of isolation for example solvent extraction, filtration etc.

In another aspect of the invention, isolation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine salt is carried out directly from step (b) to (d).

In yet another aspect of the invention, 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine may be isolated or may be used in the form of slurry to prepare 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine salt.
In yet another aspect of the invention, isolation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine salt is carried out by treating 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine with an acid in a solvent.

The acid may be a suitable acid selected from the group consisting of but not limiting to fumaric acid, lactic acid, malic acid, succinic acid, malonic acid, oxalic acid and the like. Preferably, the acid used is fumaric acid.

In another aspect of the invention, purification is done by known methods for example crystallization, sublimation, distillation and chromatography.

In another aspect of the invention, the solvent for purification or/ isolation may be selected from organic solvent or water or mixture thereof. The solvents may be selected from methanol, ethanol, isopropanol, n-butanol, acetone and acetonitrile, cyclopentane, hexane, cyclohexane, toluene, 1,4-dioxane ,diethyl ether, dichloromethane, water.

In another aspect the monophenyl PMPA (III) is reacted with silylating agent in presence of halogenating agent and a solvent which is further treated with silylated L-alanine isopropyl ester (II) in presence of a base as depicted in scheme 1b to obtain 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl) ethyl]amino} phenoxy phosphinyl)methoxy] propyl}adenine (IV).

The 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino} phenoxy phosphinyl)methoxy] propyl}adenine thus obtained by the process herein above is at least about 90% or about 95% or about 97%, or about 99% diastereomerically pure.

The process described herein above provides 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl] amino} phenoxy phosphinyl)methoxy] propyl}adenine in at least 85% yield.

The 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino} phenoxy phosphinyl)methoxy] propyl}adenine obtained from the present invention may be converted to any acid addition salt by treatment with respective acids in a solvent.

The 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino} phenoxy phosphinyl)methoxy] propyl}adenine salt obtained by the process herein above is at least about 90% diastereomerically pure.

In another aspect, the present invention provides new silylated intermediate namely silylated L-alanine isopropyl ester II. The silylated L-alanine isopropyl ester II may be used as such or may be used in the form of slurry.

The present invention described herein above is a stereoselective process that uses silylated intermediates namely silylated L-alanine isopropyl ester (IIa) resulting in 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine or its salts having higher diastereomeric excess and high yield.

The present invention described herein above is method for preparation of 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino}phenoxyphosphinyl)methoxy]propyl}adenine, that uses silylating agent catalyzing the reaction, providing very high success rate.

The present invention described herein above is easier to perform, requires less time, uses cost effective and less toxic reagents than currently available methods to obtain 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine and its salts with high yield and diastereomeric purity.

The present invention described herein above provides a process that uses less amount of solvents to obtain (9-[(R)-2-[[(S)-[[(S)-l- (isopropoxycarbonyl)ethyl] amino]phenoxyphosphinyl] methoxy] propyl]adenine and its salts with high diastereomeric purity and higher yield leading to an environment friendly process.

The process of the present invention also provides new silylated intermediate for preparation of 9-{(R)-2-[((S)-{[(S)-l - (isopropoxycarbonyl)ethyl]amino}phenoxyphosphinyl) methoxy]propyl} adenine.

According to a second aspect of the present invention, there is provided a silylated L-alanine isopropyl ester compound of formula (IIa)

Wherein, Ak is alkyl chain of C1-C4 carbon atoms

In an aspect of the invention, the silylated L-alanine isopropyl ester is compound of formula (II)

In an aspect of the invention, silylated L-alanine isopropyl ester (IIa) is prepared by treating L-alanine isopropyl ester or its salt (I) with silylating agent in a solvent.

L-alanine isopropyl ester or its salt (I) may be prepared by the methods known in the art.

The silylating agent used in the process of present invention is an alkylsilane preferably trialkylhalosilane may be selected from the group consisting of triethylsilylchloride, trimethylsilylchloride, triisoprpylsilylchloride, t-butyldimethylsilylchloride most preferably trimethylsilylchloride.

The silylating agent may be used in molar ratio between 0.1 moles to 0.5 moles, more preferably 0.3 moles, most preferably in a catalytic amount.

The solvent may be selected from organic solvent or mixture thereof. The solvents may be selected from methanol, ethanol, isopropanol, n-butanol, acetone and acetonitrile, cyclopentane, hexane, cyclohexane, toluene, 1,4-dioxane ,diethyl ether, dichloromethane, water.

The reaction is carried out at a temperature of 15-50°C preferably 20-30°C most preferably 25-30°C.

The process of the present invention is depicted below in following schemes 1a and 1b.

SCHEME 1a:

SCHEME 1b:

The following examples, which include preferred embodiments, will serve to illustrate the practice of this invention, it being understood that the particulars shown are by way of example and for purpose of illustrative discussion of preferred embodiments of the invention.
Examples
Reference example:

Preparation of (9-[(R)-2-[[(S)-[[(S)-l-(isopropoxycarbonyl)ethyl]amino] phenoxy phosphinyl]methoxy]propyl]adenine (IV) (without silylation)

100 gm (0.27 moles) of monophenyl PMPA (III) was treated with 600 ml toluene and 2 ml dimethyl formamide followed by addition of 30 ml (0.4 moles) thionyl chloride. The reaction mass was heated to 70-75°C, concentrated at 110-115°C and cooled. In another flask 138.2 gm L-alanine isopropyl ester hydrochloride was treated with 1.2 lit dichloromethane at 25-30°C and 146 ml (1mole) of triethyl amine. The L-alanine isopropyl ester obtained was added to the reaction mass and cooled. The reaction mass was cooled to -15 to -20°C. 200ml of sodium hydrogen phosphate and dichloromethane was added to the reaction mass and layers were separated. The organic layer was collected and concentrated to obtain oil. HPLC purity- about 60%

Example 1
Preparation of silylated L-alanine isopropyl ester (II)

138.2gm (0.83moles) of L-alanine isopropyl ester hydrochloride was treated 690 ml acetonitrile at 25-30°C. 98.89gm (1.1 moles) of trimethylsilyl chloride was added to the reaction mass at 25-30°C and stirred. Filtered and dried. Solid obtained is further characterized by NMR.
1H NMR (DMSO) ?? 1.19(d,6H), 4.93 (m, IH), 3.56(q, 1H), 1.27 (d, 3H), 1.5 (s, 1H), 0.08 (s, 9H)
Dry weight- 193gm

Example 2
Preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl]amino]phenoxy
phosphinyl]methoxy]propyl]adenine(IV)
100 gm (0.27 moles) of monophenyl PMPA (III) was treated with 600 ml toluene and 2 ml dimethyl formamide followed by addition of 30 ml (0.4 moles) thionyl chloride. The reaction mass was heated to 70-75°C, concentrated at 110-115°C and cooled. 9.12 gm (0.08 moles) of trimethylsilyl chloride and 500ml of dichloromethane was added.

In another flask 138.2 gm L-alanine isopropyl ester hydrochloride was treated with1.2 lit dichloromethane at 25-30°C. 9.12 gm (0.08 moles) of trimethylsilyl chloride was added to the reaction mass at 25-30°C and stirred. 146 ml of triethyl amine was added to the reaction mass and stirred at 0-5°C to obtain a solution.

The reaction mass was added to this obtained solution and cooled. The reaction mass was cooled to -15 to -20°C. The pH of the reaction mass was maintained between 7 to 8. 200ml of sodium hydrogen phosphate and dichloromethane was added and layers were separated. The organic layer was collected and concentrated to obtain a residue. The residue was treated with 100 ml isopropyl acetate, filtered and dried.

Dry weight- 120gm, diastereomeric purity- 97.5%

Example 3
Preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl]amino]phenoxy
phosphinyl]methoxy]propyl]adenine (IV)
100 gm (0.27 moles) of monophenyl PMPA (III) was treated with 600 ml toluene and 2 ml dimethyl formamide followed by addition of 33.4 ml (0.4 moles) oxalyl chloride. The reaction mass was heated to 70-75°C, concentrated at 110-115°C and cooled. 9.12 gm (0.08 moles) of trimethylsilyl chloride and 500ml of dichloromethane was added.
In another flask 138.2 gm L-alanine isopropyl ester hydrochloride was treated with1.2 lit dichloromethane at 25-30°C. 9.12 gm (0.08 moles) of trimethylsilyl chloride was added to the reaction mass at 25-30°C and stirred. 146 ml of triethyl amine was added to the reaction mass and stirred at 0-5°C to obtain a solution.

The reaction mass was added to this obtained solution and cooled. The reaction mass was cooled to -15 to -20°C. The pH of the reaction mass was maintained between 7 to 8. 200ml of sodium hydrogen phosphate and dichloromethane was added and layers were separated. The organic layer was collected and concentrated to obtain a residue. The residue was treated with 100 ml isopropyl acetate, filtered and dried.

Dry weight- 118gm, diastereomeric purity- 97.5%

Example 4
Preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl]amino]phenoxy
phosphinyl]methoxy]propyl]adenine(IV)
100 gm (0.27 moles) of monophenyl PMPA (III) was treated with 600 ml toluene and 2 ml dimethyl formamide followed by addition of 30 ml (0.4 moles) thionyl chloride. The reaction mass was heated to 70-75°C, concentrated at 110-115°C and cooled. 12.4 gm (0.08 moles) of triethylsilyl chloride and 500ml of dichloromethane was added.

In another flask 138.2 gm L-alanine isopropyl ester hydrochloride was treated with1.2 lit dichloromethane at 25-30°C. 12.4 gm (0.08 moles) of triethylsilyl chloride was added to the reaction mass at 25-30°C and stirred. 146 ml of triethyl amine was added to the reaction mass and stirred at 0-5°C to obtain a solution.

The reaction mass was added to this obtained solution and cooled. The reaction mass was cooled to -15 to -20°C. The pH of the reaction mass was maintained between 7 to 8. 200ml of sodium hydrogen phosphate and dichloromethane was added and layers were separated. The organic layer was collected and concentrated to obtain a residue. The residue was treated with 100 ml isopropyl acetate, filtered and dried.

Dry weight- 123gm, diastereomeric purity- 98%

Example 5:
Preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl]amino]phenoxy
phosphinyl]methoxy]propyl]adenine fumarate
100 gm (0.27 moles) of monophenyl PMPA (III) was treated with 600 ml toluene and 2 ml dimethyl formamide followed by addition of 30 ml (0.4 moles) thionyl chloride. The reaction mass was heated to 70-75°C, concentrated at 110-115°C and cooled. 9.12 gm (0.08 moles) of trimethylsilyl chloride and 500ml of dichloromethane was added. In another flask 138.2 gm L-alanine isopropyl ester hydrochloride was treated with1.2 lit dichloromethane at 25-30°C. 9.12 gm (0.08 moles) of trimethylsilyl chloride was added to the reaction mass at 25-30°C and stirred. 146 ml of triethyl amine was added to the reaction mass and stirred at 0-5°C to obtain a solution. The reaction mass was added to this obtained solution and cooled. The reaction mass was cooled to -15 to -20°C. The pH of the reaction mass was maintained between 7 to 8. 200ml of sodium hydrogen phosphate and dichloromethane was added to the reaction mass and layers were separated. The organic layer was collected and concentrated to obtain a residue. The residue was treated with 100 ml isopropyl acetate, filtered and dried. The obtained 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl]amino]phenoxyphosphinyl]methoxy] propyl]adenine was treated with 500 ml acetone at 25-30°C. The reaction mass was heated to 50-55°C and filtered. 11.60 gm fumaric acid was added to filtrate. The reaction mass was cooled to 25-30°C, filtered and dried to obtain a solid.

Dry wt-130 gm, diastereomeric purity- 99.8%
,CLAIMS:We claim:

1. A stereo selective process for preparation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine or its salts comprising,
a. reacting monophenyl PMPA (III) with silylating agent in presence of halogenating agent in a solvent;

(III)
b. treating step a with silylated L-alanine isopropyl ester (IIa) in presence of a base;

(IIa)
Wherein Ak is an alkyl chain of C1-C4 carbon atoms.
c. isolating 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine;
d. optionally isolating 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine salt; and
e. optionally purifying the product of step c and/or d.

2. The process according to claim 1, wherein the silylating agent is selected from alkylsilane, preferably trialkylhalosilane.

3. The process according to claim 2, wherein the silylating agent is selected from the group consisting of triethylsilylchloride, trimethylsilylchloride, triisoprpylsilylchloride and t-butyldimethylsilylchloride.

4. The process according to claim 3, wherein the silylating agent is trimethylsilylchloride.

5. The process according to claim 1, wherein the halogenating agent is selected from the group consisting of thionyl chloride, oxalyl chloride, phosphorous trichloride and phosphorous tribromide.

6. The process according to claim 1, wherein the solvent is a polar aprotic solvent.

7. The process according to claim 6, wherein the polar aprotic solvent is selected from the group consisting of ethyl acetate, methyl acetate, propyl acetate, isopropyl acetate, diethyl ether, diisopropyl ether, dimethyl formamide, tetrahydrofuran, dichloromethane, acetone, methyl ethyl ketone, methyl ter/-butylether, toluene, or acetonitrile, or a mixture thereof.

8. The process according to claim 1, wherein the base is selected from organic bases.

9. The process according to claim 8, wherein the base is selected from the group consisting of dimethylamine, triethyl amine, 4-(Dimethylamino)pyridine, N-Ethyldiisopropylamine, and Tetramethylammonium hydroxide.

10. The process according to claim 1, wherein the isolation of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine as its acid addition salt comprises treatment of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine with an acid in a solvent.
11. The process according to claim 10, wherein the solvent is selected from organic solvent, water or mixture thereof and the acid is fumaric acid.

12. The process according to claim 11, wherein the organic solvent is selected from the group consisting of methanol, ethanol, isopropanol, n-butanol, acetone and acetonitrile, cyclopentane, hexane, cyclohexane, toluene, 1,4-dioxane ,diethyl ether and dichloromethane.

13. The process according to claim 1, wherein the purification of 9-[(R)-2-[[(S)-[[(S)-1-(Isopropoxycarbonyl)ethyl] amino] phenoxyphosphinyl] methoxy] propyl] adenine or its salt is conducted using the methods selected from crystallization, sublimation, distillation and chromatography.

14. A silylated L-alanine isopropyl ester compound of formula (IIa)

Wherein Ak is an alkyl chain of C1-C4 carbon atoms.

15. The silylated L-alanine isopropyl ester according to claim 14 is compound of formula (II)