Description:FIELD OF THE INVENTION
[1] The present invention generally relates to an improved process for the preparation of an anticancer drug. More particularly, the present invention relates to an improved process for the preparation of a nucleoside analog anticancer drug. Specifically, the present invention relates to an improved process for the preparation of Gemcitabine Hydrochloride. The present invention also discloses process for the preparation of intermediates.

BACKGROUND OF THE INVENTION
[2] Gemcitabine, is a synthetic pyrimidine nucleoside analog which is used in chemotherapy medication to treat cancers. Gemcitabine is one of the most prescribed anticancer drugs worldwide. This nucleoside analog exerts its antiproliferative action after tumoral conversion into active triphosphorylated nucleotides interfering with DNA synthesis and targeting ribonucleotide reductase. Gemcitabine is mostly used in its hydrochloride salt form.
[3] Gemcitabine (2-deoxy-2’,2’-difluorocytidine) is disclosed in US 4,526,988, which discloses a synthetic method comprising the preparation of 2-deoxy-D-erythro-2,2-difluoro-ribofuranose protected at the hydroxyls at the 3- and 5- positions with a suitable protective group (OBZ) by reduction of 2-deoxy-D-erythro-2,2-difluoro-ribofuranos-1-ulose protected at the hydroxyls at the 3- and 5- positions. Such synthetic process requires remarkable energy necessary to maintain the low temperatures (between -80 and -60°C) during the reduction reaction of the carbonyl group with diisobutylaluminum hydride to avoid the ring opening at the oxygen atom.
[4] Further, in existing processes for the synthesis of Gemcitabine, both the erythro and threo isomers forms. However, high selectivity of the isomers remains a point of concern which significantly impact the yield of the process. Moreover, diisobutylaluminum hydride (DIBAL) is the most preferred reducing agent among other metal hydrides.
[5] The synthesis of high purity Gemcitabine Hydrochloride, according to the regulatory requirements contemplated by the Official Pharmacopoeias, requires purification of the beta anomer by separation of the alpha anomer, which is difficult and often involves tedious chromatographic techniques. Such chromatographic techniques requires either strong cation exchange resin or large volume of organic solvents; which all make the process costly and less economic significant. In some processes, purification of intermediate is also required.
[6] The processes known to synthesize Gemcitabine (US 4,965,374, WO 2006/095359, WO 2005/095430 and WO 2007/049294) involve the use of high volumes of solvent (70 to 100 volumes per gram of product) for purification of Gemcitabine. Use of high volumes of solvent is neither ideal for the yield nor environmentally friendly.
[7] Thus, the known processes to synthesize Gemcitabine are tedious, require sophisticated and costly facilities, result in lesser yield, low selectivity and are not environmentally friendly. There is still required an industrial process for the preparation of Gemcitabine wherein the process is simple, requires less facilities, inexpensive, highly selective in terms of desired isomers, having economic significance and more sustainable.
[8] The present invention discloses an improved process for the preparation of Gemcitabine offering a simple process, which is highly selective in terms of desired isomer results into higher yield, needs lesser facility for managing desired reaction condition, devoid of costly purification technologies and requires lesser organic solvents which reduce the burden on the environment.

SUMMARY OF THE INVENTION
[9] The present invention provides an improved process for the preparation of Gemcitabine Hydrochloride, compound of formula (I).

Formula (I),
[10] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I);

Formula (I),
the process comprising steps of:
(i) reducing a compound of formula (IA)

Formula (IA)
to provide a compound of formula (IB)

Formula (IB)
(ii) reacting compound of formula (IB) with methane sulfonyl chloride to provide a compound of formula (IC)

Formula (IC);
(iii) reacting the compound of formula (IC) and cytosine to provide a compound of formula (ID)

Formula (ID);
(iv) reacting the compound of formula (ID) and diethylamine to provide Gemcitabine, base form of compound of formula (I); and
(v) reacting the Gemcitabine, base form of compound of formula (I) and hydrochloric acid to obtain Gemcitabine Hydrochloride, compound of formula (I).
[11] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein sodium borohydride is used as reducing agent in step (i).
[12] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein reaction of step (i) is carried out at -5 to 25 o C.
[13] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (i) is selected from the group comprising ethyl acetate, tetrahydrofuran and mixture thereof.
[14] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein zinc chloride (ZnCl2) is used as a catalyst in step (i).
[15] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein methylene chloride is used as solvent in reaction of step (ii).
[16] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein triethylamine is used as catalyst in reaction of step (ii).
[17] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein reaction of step (iii) comprises steps of:
(i) dissolving Cytosine and ammonium sulfate in hexamethyldisilane;
(ii) heating the mixture to get a clear solution;
(iii) adding ethylene glycol dimethyl ether and compound of formula (IC) to the reaction mixture; and

Formula (IC);
(iv) allowing to react compound of formula (IC) and cytosine at 60 to 120 o C.
[18] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein ethylene glycol dimethyl ether is used as solvent in reaction of step (iii).
[19] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iii) is carried out at 60 to 120 o C.
[20] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iv) is carried out at 50 to 70 o C.
[21] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (iv) is selected from the group comprising methanol, ethanol, isopropyl alcohol, water and mixture thereof.
[22] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein Gemcitabine Hydrochloride is prepared by reacting Gemcitabine base with hydrochloric acid in presence of disodium EDTA.
[23] In one aspect, the present invention provides a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction molar yield is more than 90%.
[24] In another aspect, the present invention provides a process for the preparation of compound of formula (IC):

Formula (IC);
the process comprising steps of:
(i) reducing a compound of formula (IA)

Formula (IA)
to provide a compound of formula (IB); and

Formula (IB)
(ii) reacting compound of formula (IB) with methane sulfonyl chloride to provide a compound of formula (IC);
wherein sodium borohydride is used as reducing agent in step (i); wherein reaction of step (i) is carried out at -5 to 25 o C;
wherein solvent in step (i) is selected from the group comprising ethyl acetate, tetrahydrofuran and mixture thereof;
wherein zinc chloride (ZnCl2) is used as a catalyst in step (i);
wherein methylene chloride is used as solvent in reaction of step (ii); and
wherein triethylamine is used as catalyst in reaction of step (ii).
[25] In another aspect, the present invention provides a process for the preparation of compound of formula (ID):

Formula (ID);
(i) dissolving cytosine and ammonium sulfate in hexamethyldisilane;
(ii) heating the mixture to get a clear solution;
(iii) adding ethylene glycol dimethyl ether and compound of formula (IC) to the reaction mixture; and
(iv) allowing to react compound of formula (IC) and Cytosine at suitable temperature.
[26] In one aspect, the present invention provides a process for the preparation of compound of formula (IC), wherein the solution of cytosine and ammonium sulfate in hexamethyldisilane is heated at 115 to 125 o C.
[27] In one aspect, the present invention provides a process for the preparation of compound of formula (IC), wherein the reaction between formula (IC) and cytosine is carried out at 100 to 110 o C.
[28] In another aspect, the present invention provides a process for the preparation of substantially pure Gemcitabine Hydrochloride, compound of formula (I) comprising steps of:
(i) dissolving the Gemcitabine Hydrochloride, compound of formula (I) in aqueous solution of disodium EDTA and aqueous ammonia;
(ii) filtering the solution obtained in step (i);
(iii) washing the solution obtained in step (ii) with water at suitable temperature,
(iv) isolating crystalline form of Gemcitabine Hydrochloride, compound of formula (I) by adding concentrated hydrochloric acid in the solution obtained in step (iii); and
(v) washing the crystalline fraction of Gemcitabine Hydrochloride, compound of formula (I) obtained in step (iv) with acetone to obtain substantially pure Gemcitabine Hydrochloride, compound of formula (I).
[29] In one aspect, the present invention provides a process for the preparation of substantially pure Gemcitabine Hydrochloride, compound of formula (I), wherein the water in the step (iii) is added at temperature about 50 to 100 o C.
[30] In one aspect, the present invention provides a process for the preparation of substantially pure Gemcitabine Hydrochloride, compound of formula (I), wherein substantial pure Gemcitabine Hydrochloride, compound of formula (I) is having HPLC purity equal to or greater than 99.5%.
[31] In one aspect, the present invention provides an improved process for the preparation of Gemcitabine Hydrochloride, compound of formula (I); wherein the invention shows improved selectivity of desired isomer of Gemcitabine Hydrochloride, compound of formula (I).
[32] In one aspect, the present invention provides an improved process for the preparation of Gemcitabine Hydrochloride, compound of formula (I); wherein the improved process is free from chromatographic techniques which require strong cation exchange resin and a large volume of organic solvents for the purification of Gemcitabine Hydrochloride, compound of formula (I) and intermediates. Thus, the present invention makes the process technically sound and economic significant.
[33] In one aspect, the present invention provides an improved process for the preparation of Gemcitabine Hydrochloride, compound of formula (I); wherein the process requires less volume of organic solvents which makes the invented process environmentally friendly as compared to existing processes.
[34] In one aspect, the present invention provides an improved process for the preparation of Gemcitabine Hydrochloride, compound of formula (I); wherein the process is best suited to industrial production of Gemcitabine Hydrochloride, compound of formula (I); in terms of technological advancement (ease, being simpler, requirement of lesser facilities) and economic significance (cheaper reagents, isomeric selectivity and yield).
[35] In one aspect, the present invention provides an improved process for the preparation of Gemcitabine Hydrochloride, compound of formula (I); wherein the process is simpler, requires lesser facilities, cost effective, more selective in terms of isomer resulting in higher yield, economically significant and environmentally friendly.

DETAILED DESCRIPTION OF THE INVENTION
Definitions
[36] The term "reaction conditions" is intended to refer to the physical and/or environmental conditions under which a chemical reaction proceeds. Examples of reaction conditions include, but are not limited to, one or more of following: reaction temperature, solvent, pH, pressure, reaction time, mole ratio of reactants, the presence of an acid base, or catalyst, radiation, etc. Reaction conditions may be named after the particular chemical reaction in which the conditions are employed, such as, coupling conditions, hydrogenation conditions, acylation conditions, reduction conditions, etc. Reaction conditions for most reactions are generally known to those skilled in the art or can be readily obtained from the literature. Exemplary reaction conditions sufficient for performing the chemical transformations provided herein can be found throughout, and in particular, the examples below. It is also contemplated that the reaction conditions can include reagents in addition to those listed in the specific reaction.
[37] The term "reducing agent" refers to the addition of hydrogen gas to a molecule. Exemplary reducing agents include hydrogen gas (H2) and hydride reagents such as borohydrides, lithium aluminium hydride, diisobutylaluminium hydride (DIBAL-H) and lithium triethylborohydride. Reducing agent particularly refers to sodium borohydride.
[38] The term "catalyst" refers to a chemical substance that enables a chemical reaction to proceed at a usually faster rate or under different conditions (such as at a lower temperature) than otherwise possible.
[39] The term "substantially pure" refers to a HPLC purity equal to or greater than 99.5%.
[40] In addition, abbreviations as used herein have respective meanings as follows:
h=hour(s)
mL= milliliter
equiv.= equivalent
g=gram
°C=degree centigrade
HCl=hydrochloric acid
IR=infrared
NMR=Nuclear Magnetic Resonance
ESI-MS= Electrospray Ionization Mass Spectrometry
HMDS=Hexamethyldisilane
GMC=Gemcitabine
EDTA= Ethylenediaminetetraacetic acid
DIBAL-H= Diisobutylaluminium hydride
[41] The present invention discloses an improved process for the preparation of Gemcitabine Hydrochloride, compound of formula (I).

Formula (I),
[42] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I);

Formula (I),
the process comprising steps of:
(i) reducing a compound of formula (IA)

Formula (IA)
to provide a compound of formula (IB)

Formula (IB)
(ii) reacting compound of formula (IB) with methane sulfonyl chloride to provide a compound of formula (IC)

Formula (IC);
(iii) reacting the compound of formula (IC) and cytosine to provide a compound of formula (ID)

Formula (ID);
(iv) reacting the compound of formula (ID) and diethylamine to provide Gemcitabine, base form of compound of formula (I); and
(v) reacting the Gemcitabine, base form of compound of formula (I) and hydrochloric acid to obtain Gemcitabine Hydrochloride, compound of formula (I).
[43] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein sodium borohydride is used as reducing agent in step (i) to reduce a compound of formula (IA).
[44] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein reaction of step (i) is carried out at 0 to 25 o C.
[45] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein reaction of step (i) is carried out at 0 to 10 o C.
[46] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein reaction of step (i) is carried out for around 3 hours.
[47] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (i) is selected from the group comprising ethyl acetate, tetrahydrofuran and mixture thereof.
[48] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (i) is a mixture of ethyl acetate and tetrahydrofuran.
[49] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein zinc chloride (ZnCl2) is used as a catalyst in step (i).
[50] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein methylene chloride is used as solvent in reaction of step (ii).
[51] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein triethylamine is used as catalyst in reaction of step (ii).
[52] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein reaction of step (iii) comprises steps of:
(i) dissolving Cytosine and ammonium sulfate in hexamethyldisilane;
(ii) heating the mixture to get a clear solution;
(iii) adding ethylene glycol dimethyl ether and compound of formula (IC) to the reaction mixture; and

Formula (IC);
(iv) allowing to react compound of formula (IC) and cytosine at 60 to 120 o C.
[53] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein ethylene glycol dimethyl ether is used as solvent in reaction of step (iii).
[54] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iii) is carried out at 60 to 120 o C.
[55] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iii) is carried out at 100 to 110 o C.
[56] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iii) is carried out for about 8 to 15 hours; preferably from 10 to 12 hours; more preferably for 12 hours.
[57] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iv) is carried out at 50 to 70 o C.
[58] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iv) is carried out at 60 to 65 o C.
[59] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction of step (iv) is carried out for about 8 to 16 hours; preferably from 10 to 14 hours; more preferably for 12 to 14 hours.
[60] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (iv) is selected from the group comprising methanol, ethanol, isopropyl alcohol, water and mixture thereof.
[61] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (iv) is methanol or a mixture of methanol and water.
[62] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (iv) is ethanol or a mixture of ethanol and water.
[63] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (iv) is isopropyl alcohol or a mixture of isopropyl alcohol and water.
[64] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the solvent in step (iv) is a mixture of isopropyl alcohol and water.
[65] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein Gemcitabine Hydrochloride is prepared by reacting Gemcitabine base with hydrochloric acid in presence of disodium EDTA.
[66] In one embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction molar yield is more than 90%.
[67] In one preferred embodiment, the present invention discloses a process for the preparation of Gemcitabine Hydrochloride, compound of formula (I), wherein the reaction molar yield is more than 91%, more than 92%, more than 93%, more than 94% or more than 95%.
[68] In one embodiment, the present invention discloses a compound of formula (IC):

Formula (IC).
[69] In another embodiment, the present invention discloses a process for the preparation of compound of formula (IC):

Formula (IC);
the process comprising steps of:
(i) reducing a compound of formula (IA)

Formula (IA)
to provide a compound of formula (IB); and

Formula (IB)
(ii) reacting compound of formula (IB) with methane sulfonyl chloride to provide a compound of formula (IC);
wherein sodium borohydride is used as reducing agent in step (i); wherein reaction of step (i) is carried out at -5 to 25 o C;
wherein solvent in step (i) is selected from the group comprising ethyl acetate, tetrahydrofuran and mixture thereof;
wherein zinc chloride (ZnCl2) is used as a catalyst in step (i);
wherein methylene chloride is used as solvent in reaction of step (ii); and
wherein triethylamine is used as catalyst in reaction of step (ii).
[70] In one embodiment, the present invention discloses a compound of formula (ID):

Formula (ID).
[71] In another embodiment, the present invention discloses a process for the preparation of compound of formula (ID):

Formula (ID);
(i) dissolving cytosine and ammonium sulfate in hexamethyldisilane;
(ii) heating the mixture to get a clear solution;
(iii) adding ethylene glycol dimethyl ether and compound of formula (IC) to the reaction mixture; and
(iv) allowing to react compound of formula (IC) and Cytosine at suitable temperature.
[72] In one embodiment, the present invention discloses a process for the preparation of compound of formula (IC), wherein the solution of cytosine and ammonium sulfate in hexamethyldisilane is heated at 100 to 140 o C.
[73] In one embodiment, the present invention discloses a process for the preparation of compound of formula (IC), wherein the solution of cytosine and ammonium sulfate in hexamethyldisilane is heated at 115 to 125 o C.
[74] In one embodiment, the present invention discloses a process for the preparation of compound of formula (IC), wherein the reaction between formula (IC) and cytosine is carried out at 100 to 110 o C.
[75] In one embodiment, the present invention discloses a process for the preparation of compound of formula (IC), wherein the reaction between formula (IC) and cytosine carried out for about 8 to 15 hours; preferably from 10 to 12 hours; more preferably for 12 hours.
[76] In another embodiment, the present invention discloses a process for the preparation of substantially pure Gemcitabine Hydrochloride, compound of formula (I) comprising steps of:
(i) dissolving the Gemcitabine Hydrochloride, compound of formula (I) in aqueous solution of disodium EDTA and aqueous ammonia;
(ii) filtering the solution obtained in step (i);
(iii) washing the solution obtained in step (ii) with water at suitable temperature;
(iv) isolating crystalline form of Gemcitabine Hydrochloride, compound of formula (I) by adding concentrated hydrochloric acid in the solution obtained in step (iii); and
(v) washing the crystalline fraction of Gemcitabine Hydrochloride, compound of formula (I) obtained in step (iv) with acetone to obtain substantially pure Gemcitabine Hydrochloride, compound of formula (I).
[77] In one embodiment, the present invention discloses a process for the preparation of substantially pure Gemcitabine Hydrochloride, compound of formula (I), wherein the water in the step (iii) is added at temperature about 50 to 100 o C.
[78] In one embodiment, the present invention discloses a process for the preparation of substantially pure Gemcitabine Hydrochloride, compound of formula (I), wherein substantial pure Gemcitabine Hydrochloride, compound of formula (I) is having HPLC purity equal to or greater than 99.5%.

EXAMPLE:
[79] The invention can be better understood with the help of the below example. Compound of formula (I) could be synthesized through the reaction claimed in the present patent application. A detailed protocol is provided herein below which makes a person skilled in the art to synthesize the compound using the claimed invention.
Step-1: Synthesis of ((2R,3R)-3-(Benzoyloxy)-4,4-difluoro-5-hydroxytetrahydrofuran-2-yl)methyl benzoate (IB):

[80] 2-Deoxy-2,2-difluoro-D-erythro-pentafuranos-1-ulose-3,5-dibenzoate (IA) (100 g, 0.266 mole, 1.0 equivalent) was dissolved in the mixture of ethyl acetate (450 mL) and tetrahydrofuran 150 mL). Zinc Chloride (18.13 g, 0.133 mole, 0.5 equivalent) and tert-butanol (20.0 g, 0.269 mole, 1.0 equivalent) were added, followed by addition of sodium borohydride (6.037 g, 0.159 mole, 0.6 equivalent) at 0 to 5 o C and reaction mass was stirred at 0 to 10 o C for 3 hours. After completion of reaction, reaction mass was quenched with 10% aqueous hydrochloric acid solution (1000 mL) and organic layer was washed with brine (1000 mL) and dried with sodium sulfate (5 g). Reaction mass was filtered through Hyflo® bed followed by 0.25-micron filter and washed with ethyl acetate and distilled out completely under vacuum. The residue left behind is ((2R,3R)-3-(benzoyloxy)-4,4-difluoro-5-hydroxytetrahydrofuran-2-yl)methyl benzoate (IB).

Step-2: Synthesis of 2-Deoxy-2,2-difluoro-D-erythro-pentofuranose-3,5-dibenzoate-1-methanesulfonate (IC)

[81] Residue (((2R,3R)-3-(benzoyloxy)-4,4-difluoro-5-hydroxytetrahydrofuran-2-yl)methyl benzoate (IB)) obtained from step 1 was dissolved in methylene chloride (100 mL) followed by addition of triethylamine (40.37 g, 0.399 mole, 1.5 equivalent) and methane sulfonyl chloride (60.94 g, 0.532 mole, 2.0 equivalent) and stirred at -65 o C for 1 hour. After completion of the reaction, reaction mass was quenched with water and organic layer was washed with saturated sodium bicarbonate solution and brine. Organic layer was distilled out completely under vacuum and isolated in methanol to get the desired compound 2-Deoxy-2,2-difluoro-D-erythro-pentofuranose-3,5-dibenzoate-1-methanesulfonate (IC) as white solid (110 g).
Yield: 90%.
IR: Vmax (cm-1): 1740, 1718, 1653, 1601, 1584, 1558, 1490, 1450, 1441, 1414, 1375, 1358, 1340, 1333, 1313, 1262, 1235, 1206, 1173, 1144, 1132, 1094, 1071, 1053, 1043, 1022, 985, 936, 891, 871, 853, 826, 765, 685.
1H NMR (CDCl3, 400 MHz) δ 8.09-8.03 (m, 4H, Ar-H), 7.67-7.56 (m, 2H, Ar-H), 7.52-7.41 (m, 4H, Ar-H), 6.14 (d, 1H, CH), 5.58 (dd, 1H, CH), 4.85 (q, 1H, CH), 4.80-4.62 (m, 2H, CH2), 3.17 (s, 3H, CH3).
ESI m/z: 457.0 [M+H] +.

Step-3: Synthesis of 2',2'-Difluoro-2'-deoxycytidine-3',5'-dibenzoate (ID):

[82] Hexamethyldisilane (HMDS) (707.0 g, 4.3806 mole, 20 equivalent), cytosine (243.3g, 2.189 mole, 10.0 equivalent) and ammonium sulfate (0.433 g, 0.0032 mole, 0.015 equivalent) were added and heated at 115- 125 o C to get clear solution. After that reaction mass was cooled and Hexamethyldisilane was distilled out completely under vacuum and stripped out with ethylene glycol dimethyl ether (300 mL). Fresh ethylene glycol dimethyl ether (300 mL) and compound of formula (IC) obtained in step 2 (100.0 g, 0.2 mole, 1.0 equivalent) were added to the reaction mixture and heated at 100-110 o C for 12 hours. After completion of the reaction, reaction mass was cooled and poured into water (800 mL) and washed with dilute hydrochloric acid. Reaction mass was filtered, dried and isolated in methanol (800 mL) to get the desired compound 2',2'-Difluoro-2'-deoxycytidine-3',5'-dibenzoate (ID) as white solid. 70 g.
Yield: 67.83 %.
IR: Vmax (cm-1): 1687, 1599, 1534, 1448, 1383, 1341, 1316, 1306, 1260, 1216, 1195, 1167, 1132, 1111, 1085, 1068, 1052, 1036, 1027, 918, 816, 784, 760, 745, 689, 674.
1H NMR (DMSO-d6, 400 MHz) δ 9.82 (s, 1H, NH2), 8.78 (s, 1H, NH2), 8.06-8.04 (m, 2H, Ar-H), 8.01 (d, 1H, Ar-H), 7.96-7.94 (m, 2H, Ar-H), 7.74 (t, 1H, Ar-H), 7.66 (t, 1H, Ar-H), 7.58 (t, 2H, Ar-H), 7.49 (t, 2H, Ar-H), 6.43 (t, 1H, Ar-H), 6.20 (d, 1H, CH), 5.81 (q, 1H, CH), 4.89-4.84 (m, 1H, CH), 4.78-4.69 (m, 2H, CH2).
ESI m/z: 472.03 [M+H]+.
Step-4: Synthesis of Gemcitabine Hydrochloride (GMC·HCl), Compound of formula (I):

[83] 2',2'-Difluoro-2'-deoxycytidine-3',5'-dibenzoate (ID) (100.0 g, 0.2 mole, 1.0 equivalent) was dissolved in methanol (1400 mL) followed by slow addition of diethylamine (41.9 g, 0.57 mole, 2.7 equivalent) and heated at 60-65 o C for 12-14 hours. After completion of the reaction, activated carbon (1 g) was added and reaction mass was filtered through Hyflo® bed and washed with hot methanol (200 mL). Filtrate was cooled and disodium EDTA (0.025 g) and concentrated hydrochloric acid was added and stirred at 0 to 5 o C for 6 to 8 hours to get Gemcitabine Hydrochloride.
1H NMR (DMSO-d6, 400 MHz) δ 10.03 (s, 1H, NH2), 8.90 (s, 1H, NH2), 8.16 (d, 1H, Ar-H), 6.26 (s, 1H, Ar-H), 6.10-6.06 (m, 1H, CH), 4.24-4.16 (m, 1H, CH), 3.93-3.90 (m, 1H, CH), 3.80 (d, 1H, CH2), 3.65 (dd, 1H, CH2).

Step-5: Purification of Gemcitabine Hydrochloride (GMC·HCl), Compound of formula (I):
[84] Reaction mass was filtered and washed with isopropyl alcohol (2 x 50 mL) and acetone (2 x 50 mL) to get Gemcitabine Hydrochloride. The wet Gemcitabine hydrochloride was dissolved in aqueous solution of disodium EDTA (0.025 g) and aqueous ammonia to get clear solution. Reaction mass was filtered through 0.25-micron filter and washed with hot water (60 mL). Concentrated hydrochloric acid (20 g) was added to adjust the pH of the solution. Reaction mass was filtered and washed with acetone and dried under vacuum to get the analytically pure Gemcitabine Hydrochloride (GMC·HCl) as white solid. 60 g.
Yield: 94.34%.
IR: Vmax (cm-1): 3386, 3249, 3078, 2946, 1622, 1535, 1429, 1394, 1351, 1328, 1282, 1255, 1226, 1198, 1142, 1123, 1111, 1031, 1001, 917, 883, 854, 813, 784, 764, 739, 718.
1H NMR (CDCl3, 400 MHz) δ 10.03 (s, 1H, NH2), 8.90 (s, 1H, OH), 8.15-8.17 (d, 1H, Ar-H), 6.44 (s, 1H, OH), 6.26 (d, 1H, Ar-H), 6.06-6.10 (m, 1H, CH), 4.16-4.24 (m, 1H, CH), 3.90-3.93 (m, 1H, CH), 3.78-3.81 (d, 1H, CH), 3.63-3.67 (dd, 1H, CH).
ESI m/z: 264.4 [M+H] +.
ADVANTAGES OF THE PRESENT INVENTION:
[85] The present invention uses sodium borohydride as reducing agent which doesn’t require extremely low temperature as reaction condition. Thus, the present invention reduces the energy requirement to carry out the process and makes it simple and cost effective.
[86] The present invention uses aprotic polar solvents, e.g. tetrahydrofuran and ethyl acetate at a temperature ranging from -70 to +45°C.
[87] The present invention doesn’t require tedious and time-consuming chromatographic techniques for purification of Gemcitabine hydrochloride. Thus, the present invention has economic significance as compared to other processes and makes it most suitable for industrial scale production.
[88] The present invention shows improved selectivity for desired erythro isomers; wherein in the existing processes both erythro and threo isomers are formed remaining the selectivity a major concern. Due to improved selectivity, the present invention provides higher yield. The present process shows 80:20 selectivity for intended isomer in step-2 (synthesis of compound of formula IC) and 1:0 selectivity for intended isomer in step-3 (synthesis of compound of formula ID). Therefore, the yield is significantly higher in terms of desired isomer.
[89] The present invention doesn’t need purification of intermediates. Such intermediates are forwarded in downstream process as such. Thus, the present invention makes the process simple due to lesser number of steps.
[90] The present invention results into high purity Gemcitabine hydrochloride without the need of special purification technique.
[91] The present invention does not use any harmful and carcinogenic solvent and reagents; which makes the process environment friendly and green process with improved yield, purity and selectivity.
, C , Claims:I/We Claim:
1. A process for the preparation of Gemcitabine Hydrochloride, compound of formula (I);

Formula (I),
the process comprising steps of:
(i) reducing a compound of formula (IA)

Formula (IA)
to provide a compound of formula (IB)

Formula (IB)
(ii) reacting compound of formula (IB) with methane sulfonyl chloride to provide a compound of formula (IC)

Formula (IC);
(iii) reacting the compound of formula (IC) and cytosine to provide a compound of formula (ID)

Formula (ID);
(iv) reacting the compound of formula (ID) and diethylamine to provide Gemcitabine, base form of compound of formula (I); and
(v) reacting the Gemcitabine, base form of compound of formula (I) and hydrochloric acid to obtain Gemcitabine Hydrochloride, compound of formula (I).
2. The process claimed in the claim 1, wherein sodium borohydride is used as reducing agent in step (i).
3. The process claimed in the claim 1, wherein reaction of step (i) is carried out at 0 to 25 o C.
4. The process claimed in the claim 1, wherein the solvent in step (i) is selected from the group comprising ethyl acetate, tetrahydrofuran and mixture thereof.
5. The process claimed in the claim 1, wherein zinc chloride (ZnCl2) is used as a catalyst in step (i).
6. The process claimed in the claim 1, wherein methylene chloride is used as solvent in reaction of step (ii).
7. The process claimed in the claim 1, wherein triethylamine is used as catalyst in reaction of step (ii).
8. The process claimed in the claim 1, wherein reaction of step (iii) comprises steps of:
(i) dissolving cytosine and ammonium sulfate in hexamethyldisilane;
(ii) heating the mixture to get a clear solution;
(iii) adding ethylene glycol dimethyl ether and compound of formula (IC) to the reaction mixture; and

Formula (IC);
(iv) allowing to react compound of formula (IC) and cytosine at 60 to 120 o C.
9. The process claimed in the claim 1, wherein ethylene glycol dimethyl ether is used as solvent in reaction of step (iii).
10. The process claimed in the claim 1, wherein the reaction of step (iii) is carried out at 60 to 120 o C.
11. The process claimed in the claim 1, wherein the reaction of step (iv) is carried out at 50 to 70 o C.
12. The process claimed in the claim 1, wherein the solvent in step (iv) is selected from the group comprising methanol, ethanol, isopropyl alcohol, water and mixture thereof.
13. The process claimed in the claim 1, wherein Gemcitabine Hydrochloride is prepared by reacting Gemcitabine base with hydrochloric acid in presence of disodium EDTA.
14. The process claimed in the claim 1, wherein the reaction molar yield is more than 90%.
15. A process for the preparation of compound of formula (IC):

Formula (IC);
the process comprising steps of:
(i) reducing a compound of formula (IA)

Formula (IA)
to provide a compound of formula (IB); and

Formula (IB)
(ii) reacting compound of formula (IB) with methane sulfonyl chloride to provide a compound of formula (IC);
wherein sodium borohydride is used as reducing agent in step (i); wherein reaction of step (i) is carried out at -5 to 25 o C;
wherein solvent in step (i) is selected from the group comprising ethyl acetate, tetrahydrofuran and mixture thereof;
wherein zinc chloride (ZnCl2) is used as a catalyst in step (i);
wherein methylene chloride is used as solvent in reaction of step (ii); and
wherein triethylamine is used as catalyst in reaction of step (ii).
16. A process for the preparation of compound of formula (ID):

Formula (ID);
(i) dissolving cytosine and ammonium sulfate in hexamethyldisilane;
(ii) heating the mixture to get a clear solution;
(iii) adding ethylene glycol dimethyl ether and compound of formula (IC) to the reaction mixture; and
(iv) allowing to react compound of formula (IC) and Cytosine at suitable temperature.
17. The process claimed in the claim 16, wherein the solution of cytosine and ammonium sulfate in hexamethyldisilane is heated at 115 to 125 o C.
18. The process claimed in the claim 16, wherein the reaction between formula (IC) and cytosine is carried out at 100 to 110 o C.
19. A process for the preparation of substantially pure Gemcitabine Hydrochloride, compound of formula (I) comprising steps of:
(i) dissolving the Gemcitabine Hydrochloride, compound of formula (I) in aqueous solution of disodium EDTA and aqueous ammonia;
(ii) filtering the solution obtained in step (i);
(iii) washing the solution obtained in step (ii) with water at suitable temperature,
(iv) isolating crystalline form of Gemcitabine Hydrochloride, compound of formula (I) by adding concentrated hydrochloric acid in the solution obtained in step (iii); and
(v) washing the crystalline fraction of Gemcitabine Hydrochloride, compound of formula (I) obtained in step (iv) with acetone to obtain substantially pure Gemcitabine Hydrochloride, compound of formula (I).
20. The process claimed in the claim 16, wherein the water in the step (iii) is added at temperature about 50 to 100 o C.
21. The process claimed in the claim 16, wherein substantial pure Gemcitabine Hydrochloride, compound of formula (I) is having HPLC purity equal to or greater than 99.5%.
Dated this 30th day of January 2025.
Jaya Makhija [IN/PA-3488]
Agent/Attorney for the Applicant