FIELD OF THE INVENTION
The present invention relates to a novel and stereoselective synthetic process for the preparation of optically active cis-nucleoside derivatives of Formula I,

wherein R3 represents H , F, CI, CMH alkyl.
including but not limited to stereoselective preparation of Lamivudine, Emtricitabine
and related compounds.
BACKGROUND OF INVENTION
Cis-Nucleoside derivatives Lamivudine (3TC) and Emtricitabine (FTC) are useful in the treatment of retroviral infections caused by Human immuno deficiency virus (HIV), Hepatitis B virus (HBV) and Human T-Lymotropic virus (HTLV).
Lamivudine (3TC) is presently marketed by GlaxoSmithkline and is available as "EPIVIR" and is disclosed first in US 5,047,407. Emtricitabine is developed by Emory University and marketed by Gilead Sciences Inc., in the name of EMTRIVA and TRUVUDA and is first disclosed in US 5,814,639.
The oxathiolane derivatives, such as Lamivudine (3TC) of Formula-II and Emtricitabine (FTC) of Formula-Ill,



have two chiral centres and it can have four stereo isomers namely (2R,5S), (2S,5S), (2R,5R) and (2S,5R). The Pharmaceutically more active and less cytotoxic isomer is (-)-cis-isomer and has an absolute configuration (2R,5S) in both Lamivudine and Emtricitabine.
US 5,047,407 and US 6,903,224 B2 disclose the preparation of Lamivudine as a raccmic mixture i.e. (±) 3TC. The process comprises, reacting silylated cytosine with 2-benzoyloxymethyI-5-ethoxy-l,3-oxathiolane in presence of trimethylsilyl triflate (TMSTf) for three days under reflux to yield glycosidated product. Glycosidatcd product was isolated after chromatography as a mixture of cis and trans isomers (1:1). The isomeric mixture obtained after chromatography was acetylated and the acetylated mass was again subjected to chromatography for separating cis and trans isomers. Thereafter the diprotected cis or trans derivative was deprotected in basic medium (methanolic ammonia) to yield racemic Lamivudine. As evident, this is not the industrially desired condition where the glycosidation reaction is carried out for three days at reflux temperature, the reaction results in isomeric mixture (1:1) and the reaction product requires extra acetylation step for isomeric separation, involves repeated chromatographic separation and as a result of all these gives a very low overall yield of the racemic product.
US 6,831,174 and US 6,175,008 disclose a process to prepare racemic Lamivudine wherein the glycosidation reaction of silylated pyrimidine bases such as uracil / cytosine derivative and the 1,3-oxathiolane moiety, was carried out in the presence of Lewis acids such as TiCU, ZnCh, TMSI, TMSTf, SnCU or a mixture of ZnCl2 and TiCl4. Glycosidation using all these Lewis acids has resulted in the formation of mixture of cis and trans isomer in different proportions with almost 50:50 in few cases. Further, an additional step of acylation was required to separate the cis and trans isomer. The mixture or individual isomer yield was between 31-60%. TMSI had the best selectivity among all the Lewis acids. In US 6,831,174, it is stated that

when halogen containing silyl Lewis acid such as trimethyl silyl iodide is used, it converts the existing leaving group into iodo and results in better selectivity for the cis-isomcr.
US 6,153,751 and US 5,248,776 disclose a process to prepare ^-isomer of 1,3-oxathiolane pyrimidine nucleoside as racemic mixture by reacting 5-(0-protected)-2-(protectcd hydroxymethyl)-l,3-°xathiolane derivative with silylated pyrimidine base in the presence of SnCU. Also, the enantiomerically enriched 3TC and its analogs have been prepared by using the chirally pure 5-(0-protected)-2-(protected oxymethyl)-l,3-oxathiolane. The chirally pure intermediate was prepared either by enzymatic resolution, where 50% of the undesired isomer is lost resulting in lower overall efficiency of the synthesis or by low yielding multi step tedious synthesis starting from chiral sugar e.g. i-gulose. In addition to the difficulty in handling the highly air and moisture sensitive, corrosive, fuming SnCU, use of SnCl4 in nucleoside synthesis are known to pose problems due to formation of undesirable emulsions during the work-up of the reaction mixture. Some times it generates inseparable complex mixtures of a and fi isomers, which requires commercially unviable, repeated column separations. Further, it may also result in the formation of a number of stable 5-complexes between the SnCU and the basic silylated hcterocyclcs, which are difficult to remove and thus results in lower yield.
Most of the above described approach of synthesis results in racemic mixtures. In US 6,180,639, US 5,728,575, US 7,160,999, US 6,703,396 the optical active isomer of (-)-3TC or (-)-FTC has been obtained from the racemic mixture either by preparative HPLC using chiral column or by enzymatic separation involving the enantioselective hydrolysis of 5'-nucIeoside or by the enzyme assisted selective dcamination reaction.
US 6,051,709 and US 5,696,254 describes a stereocontrolled synthesis of the desired cis-nucleoside analogue starting from optically pure intermediate. As per the process disclosed in US 5,696,254, the glycosidation reaction is carried out in the presence of a silylated Lewis acid R5R6R.7R.gSi eg. iodotrimethyl silane. The process is as shown below:


As per ihe process disclosed in US 6,051,709 the glycosidation is effected without the Lewis acid catalyst, however the leaving group is limited to halo, cyano or R9SO2. The process is as shown below:


There are number of disadvantages associated with the use of silylated Lewis acid as they are highly reactive, moisture sensitive and unstable compounds. They are very expensive and have significant toxic effects. In US 6,051,709 for preparing intermediate A, halogenatiflg agent was used, which is selected from oxalyl halide, thionylhalide, phosphoroushalide, phosphorousoxyhalide. These are corrosive, hazardous and moisture sensitive compounds. In addition, hydrogen halide byproduct generated during the preparation of the halo leaving group may also result in destruction / decomposition of acid sensitive 1,3-oxathiolane moiety.
US 6,939,965 discloses the glycosidation of silylated 5-fIuorocytosinc with oxathiolane having a protected hydroxyl methyl group at second position of the oxathiolane ring. The glycosidation reaction is carried out using a Lewis acid, TiCl](0/Pr) to give the desired (2R,5S) isomer in excess and it is further purified by fractional crystallization.

WO 2004/085432 Al discloses a process to prepare emtricitabine, by condensing 5-fluorocytosine with activated 1,3-oxathiolane in the presence of a Lewis acid to give

an intermediate compound. This intermediate compound was dissolved in a solvent and treated with organic or mineral acids selected from oxalic acid, succinic acid, maleic acid, methanesulphonic acid, 4-chIorobenzenesulphonic acid, hydrochloric acid, to give an intermediate salt. Thereafter, the intermediate salt was desalified in situ and reduced using a reducing agent to give emtricitabine.
Therefore, there is currently a need for a general and economically attractive, efficient stereoselective synthesis of biologically active cis-nucleoside analogue such as I.amivudine, Emtricitabine where enantiomerically enriched /Msomer could be obtained using inexpensive reagents / catalysts in good yield with minimum formation of the undesired a-isomer,
!n the course of our investigation on the development of stereoselective glycosidation reaction for the efficient synthesis of 3TC, FTC and its analogues surprisingly it was discovered that triphenylmethyl perchlorate (trityl perchlorate) is an effective reagent for the glycosidation reaction. This reagent has been previously used for the synthesis of C-nucleoside / C-ribofuranoside {Chemistry Letters 1984; 907, 1529), where it was used during reaction of sugars with alcohols for the activation of acyloxy group on the anomeric centre of 1-O-acyl sugars. However, it has not been explored for the synthesis of 3TC, FTC and their analogues having a 1,3-oxathiolane moiety. The term analogues is meant to refer to nucleosides that are formed from pyrimidine bases substituted at the 5lh position that are coupled to substituted 1,3-oxathiolane moiety.
OBJECTIVE
The main objective of the present invention is to provide an improved stereoselective process for preparing 1,3-oxathiolane nucleoside and its intermediate, in cis-configuration, which over comes the disadvantages of prior art processes.
Yet another objective of the present invention is to provide an improved process to obtain optically pure intermediates useful in preparation of (-)-3TC and (-)-FTC and

their analogues by making use of safe reagent / catalyst in glycosidation reaction and isolation of desired stereo isomer from the diastereomeric mixture.
Yet another objective of the present invention is to provide an improved process to prepare 3TC and (-)-FTC and their analogues, which is simple, industrially applicable and economically viable.
SUMMARY OF THE INVENTION
The present invention relates to an improved process to prepare a cis-nucleoside derivative of Formula I,


wherein R2 represents H or COR4; R4 represents H, C^ alkyl; R3 represents H , F,
Cl,C,.,6alkyl.
in the presence of trityl glycosidation agent to give protected nucleoside of major [2R, 5S] compound of Formula VI and minor [2R, 5R] compound of Formula VII;


wherein Ri, and R3 are same as defined above.
d) isolating the [2R, 5S] compound of Formula VIII;
e) reducing the [2R, 5S] cis nucleoside compound of Formula VIII to give
compound of Formula I; and
0 isolating the cis-nucleoside derivative.
In another embodiment of the present invention, the deprotected cis-nucleoside compound of Formula VIII is isolated as a salt, of compound of Formula XVII and compound of Formula XVIII,

{2R, 5RJ
wherein Ri, and R3 are same as defined above,
and further converted in to Lamivudine / Emtricitabine.
In another embodiment of the present invention, Lamivudine / Emtricitabine may be efficiently isolated from an aqueous solution as water insoluble succinic acid salt or as cinnamate salt, and further converted in to Lamivudine / Emtricitabine.
DETAILED DESCRIPTION OF THE INVENTION
The 5-0-acyI-l,3-oxathiolane derivative of Formula IV is condensed with a silylated pyrimidinc base or JV-protected silylated pyrimidine base of Formula V, which includes cytosine or JV-alkanoyl-cytosine, 5-fluorocytosine or W-alkanoyl-S-fluorocytosine, in the presence of efficient trityl glycosidation agent selected from trityl perchlorate of compound of Formula X,


lo provide the corresponding nucleoside with high ^-selectivity. It will be understood that if the coupling step is carried out using a racemic 1,3-oxathiolane derivative, a racemic mixture of cis-nucleoside analog will be obtained. However, it is preferred that coupling is affected using an optically pure 1,3-oxathiolane compound of Formula IV, thereby producing the desired cis-nucleoside analog in high optically purity.
The coupling reaction of silylated base with oxathiolane derivative is carried out at 10-80°C, preferably at 40-60°C in a solvent selected from halogenated hydrocarbons such as methylene chloride, ethylene chloride; hydrocarbon such as toluene; a nitrile such as acetonitrile; an ether such as tetrahydrofuran; 1,2-dimethoxy ethane (DME) or mixtures thereof. The coupling reaction is preferably carried out in methylene chloride or toluene as a solvent, more preferably in a mixture of toluene and methylene chloride. The trityl glycosidation agent can be used in 0.3-1.3 equivalent moles based on the oxathiolane compound. However, better stereoselectivity (^-isomer > 80%) and lower reaction time was achieved when equivalent mole is used. When Jowcr quantity of trityl glycosidation agent (~0.3 m. eq.) is used it lakes longer time to complete the reaction.
When pyrimidine bases with protected amino group such as 4-N-acyl eg. 4-Ar-acetyl, 4-JV-propionyl etc., 4-AL(Ar,A'-diniethylamino methylene) were silylated and used in the coupling, the rate of reaction was better, with minimum side product formation and higher //-selectivity was obtained. Among the 4-A^-acyl protected pyrimidine bases 4-/Vr-propionyl protected base was most preferred as in the glycosidation reaction with 4-;V-acetyl cytosine formation of a major side product (-18% by HPLC analysis, Formula XII) was observed, which decreases the yield of the

reaction. Similar side reactions were not observed when instead of jV-acetyl cytosine, JV-propionyl or JV-butanoyl substituted cytosine was used for coupling reactions, this also results in the better yield of the coupled product.
The ratio of p:a isomer during the coupling in methylene chloride and toluene mixture was -80:20. However, when only acetonitrile or acetonitrile-toluene mixture was used as a solvent, the formation of desired /^-isomer was comparatively lower (fl:a isomer -71:29). The pure cis ,Y-alkanoyl nucleoside compound of Formula VI could be separated from the reaction mass (containing a diastereomeric mixture, 80% of y? i.e. 2R, 55, 20% of a, i.e. 2R, 5R) directly or a crude having the trans isomer up to 3% is isolated and then subjected to crystallization from a mixture of solvents preferably ethyl acetate and hexanes to get pure cis isomer having trans isomer less than 1%. Further, the isolated JV-alkanoyl nucleoside derivative was subjected to deacylation by hydrolysis using an acid preferably in trifluoroacctic acid, methane sulfonic acid or p-toluenesulfonic acid in an alcoholic solvent preferably absolute ethanol. The deacylated nucleoside in the form of acid addition salt could be isolated as such from the reaction mass or by adding an anti solvent such as hydrocarbons selected from toluene, hexanes, cyclohexane or an ether solvent selected from diisopropylether or an ester solvent selected from ethylacetate, isopropylacetate. The deacylated nucleoside could also be isolated as base from reaction mass by adding an organic base such as triethyiamine.
In order to minimize the loss in two isolations (isolation of iV-alkanoyl nucleoside derivative and isolation of nucleoside derivative), the reaction product having diastereomeric mixture is further subjected to acidic hydrolysis in alcoholic solvent eg. in ethanol with methane sulfonic acid, trifluoroacetic acid or p-toluenesulfonic acid. Hydrolysis is completed in -5-6 h at ambient temperature when methane sulfonic acid is used. However, in case of trifluoroacetic acid warming of reaction mass to ~45°C is required. An anti solvent selected from hydrocarbons selected from toluene, hexanes, cyclohexane or an ether solvent selected from diisopropylether or an ester solvent selected from ethylacetate, isopropylacetate is added to precipitate the pure ^-isomer as acid addition salt. Contrary to the literature reports where an additional acylation step was carried out to separate the cis and

trans isomers, pure cis isomer (having less than 2% trans isomer) could be recovered directly as methane sulfonic acid, p-toluenesulfonic acid or trifluoroacetic acid salt from a reaction mass containing a mixture of two isomers. Therefore, this presents an important advantage over the prior art processes and is an aspect of the instant invention where a and p isomers have been separated easily from the mixture as an acid addition salt. The acid addition salt thus obtained is neutralized with an organic base, preferably iriethylamine in a mixture of solvents preferably a mixture of ethyl acetate, hexanes and water. It was also noted that neither during the coupling using the trityl pcrchlorate catalyst nor during the acidic hydrolysis for acyl deprotection, any reacemization has taken place and the stereointegrity is maintained during the process. Lamivudine coupled ester / Emtricitabine coupled ester compound of Formula VIII prepared as per the above process is having a chromatographic purity of above 99% and chiral purity -100% and the diastereomer (2R, 5R) of compound of Formula IX content is < 0.3%.
The compound of Formula VIII is reduced using sodium borohydride to get 3TC or FTC or its analogue. The reduction reaction is normally carried out in aqueous alcoholic solvent, mixture of alcoholic solvent with water or aqueous tetrahydrofuran, thus resulting in a solution of Lamivudine / Emtricitabine, along with the reaction byproducts [eg. l(-)menthol] and the inorganics in aqueous alcohol. In view of the high solubility of Lamivudine / Emtricitabine in aqueous alcohol, isolation of product from such a solution is difficult by conventional methods. This difficulty has been overcome in US 6,051,709 by preparing specifically the salicylate salt, which are sparingly soluble in water and allows Lamivudine / Emtricitabine to get precipitated from water as salicylate salt. Although, the preparation of salicylate salt allows the isolation of product from water, preparation of salicylate salt has a serious problem as the salicylic acid is almost insoluble in water (0.20 g in 100 ml water, Merck Index Entry no. 8332) and therefore the precipitated salt is also accompanied with the unreacted salicylic acid and thus contaminates the product.
We have found that Lamivudine / Emtricitabine may be efficiently isolated from an aqueous solution as water insoluble succinic acid salt or as cinnamate salt. Succinic acid having good solubility in water (1 g dissolves in 1 ml of boiling water or 13 ml

of cold water, Merck Index Entry no. 8869) any succinic acid left unreacted during salt preparation gets eliminated in mother liquor during the filtration of the succinate salt and do not contaminate the product and thus overcomes the major draw back of prior art process and produces significant advantage over salicylate salt preparation. The succinate salt isolated was found to be equimolar salt having a mole of water attached with it. The preparation of water insoluble cinnamate salt affords similar advantage, as any unreacted cinnamic acid can be easily be washed out using hydrocarbon solvents such as toluene, which are usually used in the process to remove the process byproduct l(-)menthoI. Another advantage with cinnamate salt is realized when the cinnamate salt of Lamivudine is subsequently converted to the free base in organic solvent using organic tertiary base eg. triethylamine. Because of the solubility of cinnamate salt of organic base in most of the organic solvents, removal of the byproduct in isolation of Lamivudine free from any contaminant becomes very easy. Lamivudine was found to crystallize as dicinnamate salt i.e. for each mole of Lamivudine two moles of cinnamic acid is attached. Contrary to the salicylate / succinate salt the Lamivudine dicinnamate salt always crystallized as anhydrous product (with MC <0.3% w/w) from water solution. Attempt to prepare mono cinnamate salt of Lamivudine from water did not succeed. Infact, when equimolar quantity of cinnamic acid was added only 50% of product had crystallized as dicinnamate salt and the rest of the 50% had gone in aqueous mother liquor as free base.
The succinate or dicinnamate salt may be prepared by treating aqueous solution as such obtained from reduction reaction containing Lamivudine / Emtricitabine with succinic acid / cinnamic acid. A water miscible cosolvent such as methanol, efhanol, dioxane, tetrahydrofuran or mixture of these solvents could be also added during salt preparation. These salts of Lamivudine / Emtricitabine are subsequently converted into free bases in organic solvent by treatment with suitable organic bases selected from tertiary amines. Lamivudine from these salts was obtained as either Lamivudine polymorphic Form-I or Lamivudine polymorphic Form-II based on the solvents and reaction conditions used for such conversion.

In a further or alternative aspect of the present invention, the Lamivudine / Emtricitabine are isolated from aqueous solution as water insoluble sulfoxide. The aqueous solution of Lamivudine along with the inorganic impurities obtained from the reduction reaction was washed with a water immiscible solvent such as toluene / methylene chloride / cyclohexane / hexanes etc., to remove the byproduct menthol. Thereafter, the aqueous solution is treated with an oxidizing agent selected from hydrogen peroxide. It was surprisingly found that the Lamivudine sulfoxide precipitates from water leaving behind the inorganic impurities. Therefore, affords an alternative and efficient method for the isolation of Lamivudine from such media and removal of inorganic impurities. The sulfoxide thus obtained is treated with phosphorous pentasulfide (P4S10) in organic solvent to carry out deoxygenation and to obtain Lamivudine. The deoxygenation reaction is carried out under mild reaction condition in organic solvents selected from methylene chloride, toluene, acetone, pyridine, carbon disulphide or mixture of these solvents where the reaction byproducts are soluble and Lamivudine is completely insoluble. The deoxygenation reaction is carried out at RT or at ~45°C for 10-16 h.
The Lamivudine is isolated as crystalline polymorphic Form I or crystalline polymorphic Form II.
The ^-protected pyrimidine base of Formula V is prepared by known literature procedures eg., A'-acyl cytosine such as jV-propionyl cytosine.
The iV-propionyl cytosine is prepared by reacting cytosine with propionic anhydride in toluene in presence of pyridine base and catalytic amount of dimethylamino pyridine (DMAP). 4-A'-(A^Ar-dtrnethylamino methylene)eytosine was prepared by reaction of cytosine with JV-dimethylformamide dimethylacetal (DMF-dimethylacetal).
Silylated pyrimidine bases of Formula V were prepared as per the literature procedure, by reacting pyrimidine base or TV-protected pyrimidine base with a silylating agent such as hexamethyl disilazane (HMDS) with a drop of methane

sulfonic acid or a pinch of ammonium sulfate in toluene solvent. The toluene solution containing silylated product was used as such for the coupling reaction.
The 1,3-oxathiolane derivative of Formula IV was prepared by literature procedure eg. l(-)menthyl glyoxalate hydrate was prepared by reaction of l(-)menthoI with glyoxalic acid as described in US 5,489,705. l(-)menthol glyoxalate hydrate is treated with l,4-dithiane-2,5-diol as per literature procedure described in US 6,051,709 to give (2y?,5i?)-5-hydroxy-l,3-oxathioiane-2-carboxyIic acid l(-)menthyl ester. The 5-hydroxy compound is reacted with an acid anhydride / acid chloride as per the known methods (eg.: T.W. Greene "Protective groups in organic synthesis, John Wiley & Sons, New York) to get the 5-O-acyl oxathiolane derivative.
A small sample of the major side product ((- 18% by HPLC analysis) formed during the coupling of 1,3-oxathiolane and N-acetyl cytosine) was separated by column chromatography and characterized by mass and 'H & 13C NMR. The lH NMR spectrum of the above compound shows a clean singlet at 6 3.98 for the two protons of COCH2 group and the remaining protons were in the expected region. The mass spectra shows a molecular ion peak at m/z 665 with a m/z 243 (trityl fragment). Based on the characterization data, the structure of the side product was established as compound of Formula XII.

Although, the electron withdrawing nature of amide carbonyl is minimum, probably as the -NH group is attached to an aromatic nucleus, the methyl group becomes acidic enough that a proton can be picked to generate CH2". The anion generated can easily react with the stable trityl carbo-cation leading to the formation of compound of Formula XII. This non-polar impurity gets completely eliminated during the isolation of the compound of Formula VI or the corresponding dcacetylaled product compound of Formula VIII without effecting the quality of the product. Another significant observation was that during the deacetylation of the

compound of Formula VI by acidic hydrolysis using methane sulfonic acid / trifluoroacetic acid or ^j-toluenesulfonic acid in alcohol to get product of Formula VIII, the -COCH2C(C6H5)3 did not get hydrolyzed.



The invention is illustrated with the following examples, which are provided by way of illustration only and should not be construed to limit the scope of the invention.
EXAMPLE 1
PREPARATION OF N-ACETYLCYTOSINE
To a suspension of cytosine (200 g, 1.80 mol) in toluene (600 ml) at RT (25-30°C), pyridine (216 g, 2.73 mol) and DMAP (1.0 g) was added. Acetic anhydride (217 g, 2.13 mol) was diluted with toluene (350 ml) and added slowly in -60 min at 25-45°C. After addition, the reaction mass was heated to 50-55°C and continued stirring for 6 h to complete the reaction (checked by TLC). Reaction mass was

cooled to 25-28°C and product was filtered and washed with toluene (350 ml).
Further, product was washed with water (2 x 300 ml). Product was dried at 60-65°C
under reduced pressure to give title compound.
Yield: 254 g
'H NMR(DMSO-4): 5 2.08 (s, 3H, CH3), 7.08-7.10, (d, 1H, CH-cytosine), 7.79-
7.81(4, 1H, CH-cytosine), 10.75(broad singlet, 1H, NH), 11.50(broad singlet, 1H,
Nil).
EXAMPLE 2
PREPARATION OF N-PROPIONYLCYTOSINE
To a suspension of cytosine (200 g, 1.80 mol) in toluene (700 ml) at RT (25-30°C), pyridine (222 g, 2.81 mol) and DMAP (1.0 g) was added. The suspension was heated to ~45DC and propionic anhydride (273 g, 2.10 mol) was diluted with toluene (500 ml) and added slowly over 2 h at 45-55°C. After completion of addition, stirring was continued at 55-58°C for 9 h to complete the reaction (checked by TLC). Reaction mass was cooled to 25-28°C and product was filtered and washed with toluene (350 ml). Further, product was washed with water (2 x 300 ml). The wet product was stirred for 90 min at 38-40°C in water (1500 ml) containing triethylamine (6 g). Product was filtered and washed with water (2 x 300 ml). Product was dried at 50-55°C under reduced pressure to give title compound. Yield: 272.3 g.
'H NMR(DMSO-d6): 5 0.99-1.04, (t, 3H, CHj), 2.35-2.43, (q, 2H, CH2), 7.11-7.13(4, 1H, CH-cytosine), 7.79-7.82(4, 1H, CH-cytosine), 10.71(broad singlet, 1H, NH), 11.49 (broad singlet, 1H, NH).
EXAMPLE 3
PREPARATION OF (l'R^'S^'RJ-MENTHYL-S-ACETOXY-M-OXATHIOLANE-2R-CARBOXYLATE
To aprecooled suspension of (l'J?,2'S,5'i?)-menthyl-5Ji-hydroxy-l>3-oxathiolane-2/?-
carboxylate (Hydroxy compound, 300 g, 1.04 mol) in diisopropyl ether (1050 ml) at 0-5°C, pyridine (111.8 g, 1.41 mol) and 4-dimethylaminopyridine (3 g, 0.02 moles) was added under nitrogen. Acetic anhydride (125.6 g, 1.23 mol) was diluted with

diisopropyl ether (250 ml) and added to reaction mass at 0-8°C in -2 h. Stirring of reaction mass was continued at 3-8°C for 6 h to complete the reaction. After completion of reaction (checked by TLC), the reaction mass was heated to RT (20-30°C) and diluted with diisopropyl ether (250 ml). Thereafter, reaction mass was washed with warm water (3 x 500 ml, ~40°C) to ensure complete removal of pyridine. Organic layer was concentrated under reduced pressure at less than 45°C to obtain a residue. To the residue hexanes (420 ml) was added and the contents were heated to ~50°C and maintained for 15-20 min at same temperature. Thereafter, it was cooled to RT and stirred for -30 min. Further, diefhylefher (25 ml) was added to the slurry at ~25°C and stirred for 30 min. The product slurry was cooled to 4-6°C and stirred for 2 h at the same temperature. Product was filtered and washed with prechilled hexanes (150 ml, 0-5°C). Product was dried at ~45°C under reduced pressure for 6 h to give title compound. Yield: 259.5 g
'H NMR(DMSO-ds): 6 6.69(4 1H, H-5), 5.78(5, 1H, H-2), 4.60-4.66(m, 1H, CH-O-CO of menthyl), 3.31-3.37(m, 2H, H-2) 2.04(5, 3H, O-COCH3), {(1.87-1.91, 1.62-1.66, 1.39-1.43, 2H+2H+2H = 6H of menthyl), (1.02-1.08, 2H of menthyl), (0.86-0.91 & 0,71-0.74 7H+3H=10H of menthyl)}.
EXAMPLE 4
(2R,5S)-5-(CYTOSIN-l-YL)-l,3-OXATHIOLANE-2R-CARBOXYLIC ACID (1 'R, 2'S, 5'R)-MENTHYL ESTER (LAMIVUDINE COUPLED ESTER)
Hexamethyl disilazane (65 g, 0.40 mol) was added to a suspension of N-acetylcytosine (61.2 g, 0.40 mol) in toluene (305 ml) at room temperature. Mefhanesulfonic acid (0.3 g) was added to the above suspension and the contents were heated to reflux (108-113°C). Reflux was continued for 3 h to obtain a clear solution and to complete the silylation reaction. Thereafter, -80 ml toluene was distilled from the reaction mass at atmospheric pressure. The contents were cooled to 30-35°C under nitrogen atmosphere and methylene chloride (405 ml) was added. Further, (l'R, 2'S, 5'R)-menthyl-5-acetoxy-l,3-oxathiolane-2R-carboxylate (134.7 g, 0.41 mot) was added and the resulting solution was cooled to 10-15°C. Trityl pcrchlorate (137 g, 0.40 mol) was added at 10-30°C. The resulting solution was

stirred for 60-70 min at 20-30°C and thereafter heated to reflux (43-51°C). Reflux was continued and progress of reaction was followed by qualitative HPLC analysis, till N-acetylcytosine left unreacted was < 6% (it took -12 h to achieve this). After completion of reaction, the reaction mass was diluted with methylene chloride (150 ml) and the reaction mass was cooled to 20-30°C. The reaction mass was poured into 1.2 Lt of 7.0% w/v aqueous sodium bicarbonate solution under stirring at 20-30°C. Organic layer was separated and concentrated under reduced pressure at ~50°C to get a residue containing the N-acetyl coupled ester along with its diastereomer ifi:a = 79:21, by HPLC analysis).
The above residue was dissolved in ethanol (400 ml) and methane sulfonic acid (68 g) was added at 20-30°C. The solution was stirred at 20-30°C, where a precipitation was observed after -90 min of stirring. The progress of reaction was followed by qualitative HPLC analysis and it was continued till the starting material N-acety] coupled ester was < 1% (it took -4 h). The reaction mass was diluted by slow addition of diisopropyl ether (700 ml) and further stirring was continued at 20-30QC for 2 h to complete the crystallization. The product was filtered and washed with a mixture of ethanol (50 ml) and diisopropyl ether (90 ml). Further, it was washed with diisopropyl ether (80 ml) and sucked thoroughly under nitrogen to remove most of the mother liquor. The Lamivudine coupled ester methane sulfonic acid salt (119 g, wet) was obtained.
Small sample from above wet product was dried and analyzed by 'H NMR. DMSO-4 : 5 2.32 (S, 3H, methane sulfonic), 3.38(dtf, 1H, C-4), 3.65(: -174.4 (c - 1 in CHC1;, on anhydrous basis).
EXAMPLE - 7
(2R,5S)-5-(N-4-PROPIONYLCYTOSIN-l-YL)-l,3-OXATHIOLANE-2R-CARBOXYLIC ACID (l'R, 2'S, 5'R)-MENTHYL ESTER (N-
PROPIONYLLAMIVUDINE COUPLED ESTER)
JJexamethy] disilazane (35.4 g, 0.22 mol) was added to a suspension of N-propionylcytosine (33.4 g, 0.20 mol) in toluene (180 ml) at room temperature. Methanesulfonic acid (0.2 g) was added to the above suspension and the contents were heated to reflux (108-113°C). Reflux was continued for 4 h to obtain a clear solution and to complete the silylation reaction. Thereafter, -50 ml toluene was distilled from the reaction mass at atmospheric pressure. The contents were cooled to ~40°C under nitrogen atmosphere, then a mixture of toluene (60 ml) and methylene chloride (180 ml) was added. Further, (l'R, 2'S, 5'R)-menthyl-5-acetoxy-l,3-oxathiolane-2R-carboxylate (70 g, 0.21 mol) was added and the resulting solution was cooled to 10-15°C. Trityl perchlorate (70 g, 0.20 mol) was added at 10-30°C. The resulting solution was stirred for 60-70 min at 20-30°C and thereafter heated to 53-56°C. Stirring at 55-56°C was continued and progress of reaction was followed by qualitative HPLC analysis, till N-propionylcytosine left unreacted was < 8% (it took -15 h to achieve this). After completion of reaction, the reaction mass was diluted with methylene chloride (100 ml) and the reaction mass was cooled to 20-30°C. The reaction mass was poured into 800 ml of 6.0% w/v aqueous sodium bicarbonate solution under stirring at 20-30°C. The organic layer was separated and washed with water (200 ml). The washed organic layer was concentrated under

reduced pressure at ~50°C to get a residue. A mixture of cyclohexane (270 ml) and ethyl acetate (85 ml) was added to the residue and the contents were refluxed for 30-40 min. The product slurry was cooled to ~18°C and stirred for about 30 min at this temperature. Product was filtered and washed with chilled ethyl acetate (50 ml). Product was dried under reduced pressure at ~45°C to obtain iV-Propionyllamivudine coupled ester. Yield: 46.8 g SOR I«]2SD: -158.4 (c = 1 in CHC13, on anhydrous basis).
EXAMPLE - 8
4-AMINO-l-[(2R,5S)-2-(HYDROXYMETHYL-l,3-OXATHIOLAN-5-YL)-(HI)-PYRIMIDIN-2-ONE DICINNAMATE (LAMIVUDINE DICINNAMATE)
Dipotassium hydrogen phosphate (129 g) was dissolved in DM water (190 ml) at 23-35°C. Ethanol (1000 ml) was added to the above solution and cooled to 18-25°C. (2R,5S)-5-(cytosin-l-yl)-l,3-oxathiolane-2R-carboxyIic acid (l'R, 2'S, 5'R)menthyl ester (Lamivudine coupled ester; 145 g, 0.38 moles) was added to the above biphasic mixture followed by methanol (75 ml) at 18-25°C. Sodium borohydride (32 g, 0.85 moles) was dissolved in precooled sodium hydroxide solution (0.02% w/v, 190 ml). This sodium borohydride solution was added to the reaction mass over a period of 90 min at 18-25°C. Stirring was continued at 18-25°C for -3 h to complete the reaction. Reaction mass was settled for 1 h and the aqueous (buffer layer) was separated. The aqueous layer was reexiracfed with ethanol (100 ml). The organic layers were combined and pH was adjusted to -5.8 with dilute hydrochloric acid (-10% w/w, -20 ml) to decompose the unreacted sodium borohydride. Thereafter, pH was adjusted back to 7.5-7.8 using 10% w/v aqueous sodium hydroxide solution (22 ml). Most of the ethanol - methanol mixture was removed by distillation at atmospheric pressure (at ~83°C). Further, any residual ethanol was distilled under reduced pressure at ~55°C. The resulting oily residue was dissolved in water (600 ml) and washed with toluene (160 ml) to remove the byproduct (l)-menthol. The aqueous solution was treated with charcoal (5 g) and filtered through a hyflo bed and the bed was washed with water (50 ml) to obtain a clear filtrate (-760 ml).

Cinnamic acid (107 g, 0.72 moles) was added to the clear filtrate and the suspension
was heated at 40-45°C for 30 min. Thereafter, cooled the product suspension to 20-
25°C and stirred for 3 h to complete the crystallization of the product. Product was
filtered and washed with DM water (100 ml) followed by toluene (70 ml). The
product was dried under reduced pressure at A0-45°C to obtain Lamivudine
dicinnamatc.
Yield: 172 g
■ll NMR(DMSO-d6): 8 3.05(44, 1H), 3.39(44, 1H), 3.74(5, 2H, 5-17(f, 1H), 5.40(5,
br, 1H), 5.72(4, 1H), 6.21((, 1H), 6.53(4,./ = 16 Hz, 2H cinnamic), 7.25(4, br, 211),
7.42(m, 6H), 7.58(4; = 16 Hz, 2H cinnamic), 7.68(m, 4H), 7.82(4, 1H), 12.42(s, br,
2H).
EXAMPLE - 9
4-AMINO-l-[(2R,5S)-2-(HYDROXYMETHYL-l,3-OXATHIOLAN-5-YL)-(lH)-PYRIMIDIN-2-ONE SUCCINATE MONOHYDRATE {LAMIVUDINE SUCCINATE MONOHYDRATE)
Reduction of Lamivudine coupled ester (145 g) was carried out as described in
Example-8 and by similar work-up an aqueous solution (-760 ml) containing
Lamivudine was obtained. Succinic acid (43.7 g, 0.37 mol) was added to the
solution and the contents were heated to 35-40°C for 25-30 min. Thereafter, the
product suspension was cooled to 5-8°C and stirred for 2 h at the same temperature.
Product was filtered and washed with prechilled water (100 ml). The product was
dried at 40-45°C under reduced pressure to obtain Lamivudine succinate
monohydrate.
Yield: 124.6 g
Moisture content: 5.31% w/w
Succinic acid content (by titrimetry): 32.42% w/w
'II NMR(DMSO-d6): 5 2.42(j, 4H, succinic), 3.03(44, 1H), 3.41(44, 1H), 3.72(m,
2H), 5.l7(f, 1H), 5.3 1(J, br, 1H), 5.73(4, 1H), 6.20((, 1H), 7.25(4, br, 2H), 7.82(4,
HI).

EXAMPLE -10
4-AMINO-l-((2R,5S)-2-(HYDROXYMETHYL)-3-OXO-13-OXATHIOLAN-5-YL]-(lII)-PYRIMIDIN-2-ONE (LAMIVUDINE SULFOXIDE)
Reduction of Lamivudine coupled ester (145 g) was carried out as described in Example 7 and by similar work-up an aqueous solution (-760 ml) containing Lamivudine was obtained. Hydrogen peroxide (27 g, -48% w/w, -0.38 mol) was added slowly in -60 min to get the clear solution at 22-30°C. The contents were stirred at 22-30°C for -6 h, where product crystallizes out. Thereafter, the product slurry was cooled to 5-8°C and stirred for 2 h at the same temperature. Product was filtered and washed with prechilled water (75 ml). The product was dried at 45-50°C under reduced pressure to obtain Lamivudine sulfoxide. Yield: 82.5 g
'II NMR(DMSO-d6): 5 3.06(44, 1H), 3.33(44, 1H), 3.72(m, 1H), 3.78(m, IH), 4.64(/, IH), 5.48(f, IH), 5.80(4 IH), 6.72(44, IH), 7.38(6™, 2H), 7.69(4 IH).
EXAMPLE-11
4-AMINO-l-[(2R,5S)-2-(HYDROXYMETHYL)-13-OXATHIOLAN-5-YLl-(1 H)-PYRIMJDIN-2-ONE (LAMIVUDINE)
Lamivudine sulfoxide (10 g, 41 mmol) was taken in pyridine (40 ml) and phosphorous pentasulfide (4.40 g, 20 mmoles) was added at 22-30°C under nitrogen atmosphere. Stirring was continued at 22-30°C for 6 h. Thereafter, the mass temperature was raised to ~40°C and further stirred for 4 h at 40-42°C to complete the reaction. Reaction mass was filtered and the clear filtrate was concentrated under reduced pressure to about half of its volume. The concentrate was diluted with toluene to precipitate the product. Crude product was filtered washed with hot toluene. Further, purification of crude product was carried out by crystallization from ethanol to yield the title compound. Yield: 6.3 g.

EXAMPLE - 12
4-AMINO-l-I(2R,5S)-2-(HYDROXYMETHYL-l,3-OXATHIOLAN-5-YL> (lII)-PYRIMIDIN-2-ONE (LAMIVUDINE FORM I)
Lamivudine dJcinnamate (100 g, 0.19 moles) was added to ethyl acetate (600 ml) containing water (7 ml) and the resulting slurry was stirred at 22-30°C for -15 min. Triethylamine (44.5 g, 0.44 moles) was diluted with ethyl acetate (80 ml) and added to the above slurry slowly over a period of 30 min at 22-30°C. The product slurry was further stirred at 22-3G°C for 4 hours. The product was filtered and washed with ethyl acetate (2 x 60 ml). Product was dried under reduced pressure at 40±2°C to obtain Lamivudine Form I. Yield: 39.6 g
EXAMPLE -13
4-AMINO-l-[(2R,5S)-2-(HYDROXYMETHYL-l,3-OXATHIOLAN-5-YL)-(lH)-PYRIMIDIN-2-ONE (LAMIVUDINE FORM II)
Lamivudine dicinnamate (100 g, 0.19 moles) was added to ethanol (350 ml) and the resulting slurry was stirred at 22-30°C for -15 min. Triethylamine (44.5 g, 0.44 moles) was diluted with ethanol (50 ml) and added to the above slurry slowly over a period of 30 min at 22-30°C. The product slurry was further heated to reflux (~80°C) and continued till a clear solution was obtained. The solution was cooled to ~55°C and seeded with Lamivudine Form-II (0.1 g) and further cooled to 15-18°C. Stirring was continued at 15-18°C for 2 h. The product was filtered and washed with precooled ethanol (50 ml). Product was dried under reduced pressure at 50-55°C to obtain Lamivudine Form II. Yield: 38.2 g

EXAMPLE -14
4-AMINO-l-[(2R,5S)-2-(HYDROXYMETHYL-l,3-OXATHIOLAN-5-YL)-(lH)-PYRIMIDIN-2-ONE (LAMIVUDINE FORM I)
Lamivudine succinate monohydrate (100 g, 0.27 moles) was added to a mixture of cthanol (210 ml) and water (24 ml) and the resulting slurry was stirred at 22-30X for -15 min. Triethylamine (57.6 g, 0.57 moles) was diluted with ethanol (80 ml) and added to the above slurry slowly over a period of 30 min at 22-30°C. The product slurry was further stirred at 22-30°C for 4 hours. Further, it was cooled to ~5°C and stirred at this temperature for 60 min. The product was filtered and washed with precooled ethanol (60 ml). Product was dried under reduced pressure at 40±2°C to obtain Lamivudine Form I. Yield: 49.8 g
EXAMPLE -15
4-AMINO-l-[(2R,5S)-2-(HYDROXYMETHYL-13-OXATHIOLAN-5-YL)-(HI)-PYRIMIDIN-2-ONE (LAMIVUDINE FORM I)
Lamivudine succinate monohydrate (100 g, 0.27 moles) was added to a mixture of methanol (230 ml) and water (30 ml) and the resulting slurry was stirred at 22-30°C for -15 min. Triethylamine (57.6 g, 0.57 moles) was diluted with methanol (40 ml) and added to the above slurry slowly over a period of 30 min at 22-30°C. The product slurry was further stirred at 22-30°C for 4 hours. Further, it was cooled to ~-5°C and stirred at this temperature for 60 min. The product was filtered and washed with a mixture of methanol (40 ml) and ethyl acetate (60 ml). Product was dried under reduced pressure at 40±2°C to obtain Lamivudine Form I. Yield: 51.6 g Methanol content: -1.3%

EXAMPLE-16
4-AMINO-l-[(2R,5S)-2-(HYDROXYMETHYL-l,3-OXATHIOLAN-5-YL)-(lH)-PYRIMIDIN-2-ONE (LAMIVUDINE FORM I)
Lamivudine succinate roonohydrate (51 g, 0.14 moles) was suspended in a mixture of isopropmol (250 ml) and water (7.5 ml) at 23-28°C. Triethylamine (29.6 g, 0.29 moles) was added to the above suspension at 23-28°C in about 20 min and the resulting slurry was stirred for 3 h at 23-28°C. The product slurry was cooled to 0-5°C and stirred for 60 min. The product was filtered and washed with precooled isopropanol (100 ml, 5°C). Dried the wet product under reduced pressure at ~45°C to obtain Lamivudine polymorph Form-I. Yield: 28.5 g
SOR|a|D ' (c = 1 in methanol on anhydrous basis): -142° Water content: 1.7 % w/w.

WE CLAIM
1) An improved process to prepare a cis-nucleoside derivative of Formula I,

wherein R3 represents H , F, CI, C1-6 alkyl. which comprises: a) reacting 1,3-oxathiolane compound of Formula IV,
Formula IV
wherein R represents acyl group selected from -COCH3, -COC2H5, -COCH2CI, -COCHiBr, -COC6H5, -COR5; R5 represents substituted phenyl group selected from 4-nitrophenyI, 4-chlorophenyl; R1 represents hydrogen, alkyl, aralkyl, alkenyl, aryl preferably an alkyl group / substituted alkyl group, more preferably a chiral auxiliary with one or more chiral centres such as d(+)menthol or l{-)menthol. with a pyrimidine base of Formula V,
Formula V
wherein R2 represents H or CORj; R4 represents H, C1-6 alkyl; R3 represents H , F,
C1, C1-,6alkyl.
in the presence of irityl glycosidation agent to give protected nucleoside of major [2R, 5S] compound of Formula VI and minor [2R, 5R] compound of Formula VII;


d) isolating the [2R, 5S] compound of Formula VIII;
e) reducing the [2R, 5S] cis nucleoside compound of Formula VHI to give a compound of Formula I; and
f) isolating the cis-nucleoside derivative.


3) The process according to claim 2, trityi glycosidation agent can be used in 0.3-
1.3 equivalent moles based on the 1,3-oxathiolane compound of Formula IV.
4) The process according to claim 1, the step (a) reaction is carried out at 10-
80°C, preferably at 40-60°C, in a solvent.
5) The process according to claim 4, the solvent is selected from halogenated
hydrocarbons such as methylene chloride, ethylene chloride; hydrocarbon such
as toluene; a nitrile such as acetonitrile; an ether such as tetrahydrofuran; 1,2-
dimethoxy ethane (DME) or mixtures thereof
6} The process according to claim I, the deprotection of the compound of Formula VI is carried out using an acid selected from trifluoroacetic acid, methane sulfonic acid, p-toluenesulfonic acid in an alcoholic solvent selected from absolute ethanol.
7) The process according to claim 6, the deprotected cis nucleoside compound of Formula VIII and compound of Formula IX in step (c) is isolated in the form of acid addition salt as such from the reaction mass or by adding an anti solvent selected from hydrocarbons selected from toluene, hexancs. cyclohexane or an ether solvent selected from diisopropylether or an ester


8) The process according to claims 1, step (d) the deprotected nucleoside of
compound of Formula VIII is isolated by adding an organic base selected from
iriethylamine in a mixture of solvents selected from a mixture of ethyl acetate,
hexanes and water.
9) The process according to claim 1, the reduction in step (e), is carried out using
sodium borohydride in aqueous alcoholic solvent, mixture of alcoholic solvent
with water or aqueous tetrahydrofuran.
10) The process according to claim 1, after the reduction the cis-nucleoside is
isolated as a succinic sah or cinnamate salt.
11) The process according to claim 1, the succinic sah or cinnamate salt of cis-
nucleoside is hydrolysed to give cis-nucleoside of compound of Formula I.

12) The process according to claim I, the cis-nucleoside is isolated from aqueous
solution as water insoluble sulfoxide, by adding an oxidizing agent selected
from hydrogen peroxide.
13) The process according to claim 13, the sulfoxide formed is treated with phosphorous pentasulfide (P4S10) in organic solvent selected from methylene chloride, toluene, acetone, pyridine, carbon disulphide or mixture of these solvents to carry out deoxygenation and to obtain cis-nucleoside.
14) The process according to claims 1, 10 and 12 the cis-nucleoside is selected from Lamivudine, Emtricitabine.



wherein R, represents hydrogen, alkyl, aralkyl, alkenyl, aryl preferably an alkyl group / substituled alkyJ group, more preferably a chiral auxiliary with one or more chiral centres such as d(+)menthol or l(-)menthol; X represents methanesulphonic acid, trifluoroacetic acid, p-to)uenesu]fonic acid.