WO2010029574A2 - An improved process for the preparation of gemcitabine and its intermediates using novel protecting groups and ion exchange resins - Google Patents

Abstract

An improved process for preparing gemcitabine or salts thereof, comprising the following steps, dissolved 2-Deoxy-2,2- difluoro-D-ribofuranose-3,5-dibenzoate in an organic solvent and allowed to cool to 0 to 10 degree centigrade temperature, stirred for a period of 1 to 2 hours to this triethylamine is added for a period of 2 to 10 minutes, an indigenous protecting compound added under constant stirring, solvents were distilled off, and to the residual volume IN HCL is added until to form a clear solution adjusted the PH of about 2, further to this an ethyl acetate was added followed by washing the organic phase using NaHCo3, water and brine solution, the organic phase was dried over MgSo4, concentrated the above reaction mixture under reduced pressure to obtain a crude 2-deoxy-2,2-difluoro-D- riboufuranose-3,5-dibenzoate-l-arylateas an oil phase, above product is dissolved in dry methanol under constant stirring using Ion exhange resin for a period of 15 to 25 hours at 30 to 45 degree centigrade temperature, which gives pure gemcitabine hydrochloride.

Description

Title: - An improved Process for the Preparation of Gemcitabine and its intermediates using Novel Protecting groups and Ion Exchange Resins,

Field of the invention :-

The present invention relates to the art of organic chemistry. Specifically it rlates to certain alpha., alpha, -difluoro-.beta — hydroxyl thiol esters and processes to make them using protecting groups and resin.

Back Ground of the invention :-

EP0306190 (expiry 2008) Patent claims a process for preparing an enantiomeric mixture of erythro and threo lactones using a dioxolan in presence of a strong acid followed by azeotropic distillation of water. EP0576229 (expiry 2013)This patent protects a process for preparing an alpha-anomer enriched 2-deoxy-2',2'-difluoro- Dribofuranosylsulfonates. EP0576232 (expiry 2013) Patent claims a process for separating 2-deoxy-2\2'- difluoro-D-ribofuranosylalkyl sulfonate anomers.

EP0577302 (expiry 2013)This process provides an alpha- anomer enriched ribofuranosyl sulfonate from a beta- anomer ribofuranosyl sulfonate. EP0577303 (expiry 2013) This process provides a process for preparing a beta- anomer enriched nucleoside from an alphaanomer enriched carbohydrate based on a SN2-Displacement reaction.

6. EP0640614 (expiry 2014) Patent claims a stereoselective process for preparing a beta-enriched carbohydrate. The schematic synthesis does not reflect whether alpha or beta anomers or an enantiomeric mixture are used or a SN1 or SN2 displacement is favoured. EP0655454 (expiry 2014) We cannot evaluate this patent which claims a process for preparing useful intermediates for carbohydrates synthesis. EP0677527 (expiry 2015)This patent claims a process for preparing intermediates in carbohydrate synthesis. EP0688782 (expiry 2008)This patent claims a process for selectively isolating (erythro)-2',2'- difluorolactones from an enantiomeric mixture of erythro and threo lactones. EP0688783 (expiry 2008) Patent comprises a process for producing a 2'-deoxy-2',2^I- difluoronucleoside in about a 1 :1 alpha to beta ratio. EP0712860 expiry 2015) Patent claims a process to purify and isolate a beta-anomer enriched nucleoside in presence of a nucleobase and acetonitrile. EP0735043 (expiry 2013) It is not clear whether the patent also claims the introduction of fluor in the carbohydrate.

Gemcitabine HCI. Marketed by EIi Lilly is a nucleotide analogue that exhibits antitumur activity and belongs to a general group of chemotherapy drugs known as antimetabolites. Germa=ctabine orevents cells from producing DNA and RNA by interfering with the synthesis of nucleic acids, thus stopping the growth of cancer cells and causing them to die.

Gemcitabine is a synthetic glucoside analog of cyctosine, which is chemically described as 4-amino-1-(2-deoxy-2,2- difluroro-[beta]-D-ribofuranosyl)-pyrimidin-2)(1 H)-one or 2(1 H)- one or T-deoxy-2',2'-difluorocytidine ([beta] isomer). Gemcitabine HCI has the following structure.

Gemsar is supplied in vials as the hydrochloride salt in sterile form, only for intravenous use, containing either 200 mg or 1 g of gemcitabine HCL (as free base ) formulated with mannitol (200 mg or 1 g, respectively ) and sodium acetate (12.5mg r 62.5 mg, respectivel ) as a sterile lyophilized powder , Hydrochloride acid and /or sodium hydroxide may have been added for PH adjustment.

US patent No. 4808614, describes a process for synthetically producing gemcitabine using D-glceraldehde ketals is reacted with bromodifluoroacetic acid ethyl ester in the presence of activated Zinc, to obtain ethyl 2,2-difluoro - 3- hydroxyl-3-(2,2-dimethldioxolan-4-yl) -propionate 3 as mixture of 3-Rand 3-S isomers. The 3-R to 3-S isomer ratio is about 3:1. the 3-Risomer has the stereochemistry required for producing the desired erythro (3-R) ribose structure , and can be separated from the 3-S isomer . the resulting product is cyclized by treatment with an acidic ion exchange resin, such as Dowex 5OW -X12, to produce 2-deoxy-2,2-difluoro-D-erythro-pentanoic acid-[gamma]-lactone 4. the hydroxyl groups of the lactone are protedted woth fert-butyldimethylsilyl.

TBDMS protecting groups to obtain the proteted lactone 3,5-bis-(ter-butyldimethylsilyloxy)-2-desoxy-2,2-difluoro-1- oxoriboseδ, and the prodct is redeiced to obtain 3,5-bis-tert- butyldimethylsilyl)-2-desoxy-2,2-difluororibose6.

The position 1 of the carbohydrate is acvated by the introduction of leaving group, e.g. methanesulfonyloxy(mesylate), formed by reacting compound 6 with methanesulfonyl chloride to obtain 3,5-bis-ter- butyldimethysilyloxy)-1-methanesulfonyloxy-2-desoxy- 2,2difluororibose 7. the base ring is coupled to the carbohydrate by reacting compound 7 with N,)O-bis-(trimethylsilyl)-cystosine 8 in the presence of a reaction inititor, such as trfluoromethanesulfonyloxy trimethylsilane ( trimethylsilyl triflate). Removal of the orotecting groups and chromatographic purification affords gemcitabine free base.

US Patent NO. 4526988 which teaches about erythro isomer can be isolated in crystalline form from a mixture of erythro and threo isomers,

The 3-hydroxy group of compound 3 is esterified with a benzoyl protecting group by reaction with benzoyl chloride, benzoyl bromide, benzoyl cyanide, benzoyl azide, etc, in presence of a tertiary amine or a catalyst such as 4- ( dimethylamino) pyridine or 4-pyrrolidine, to obtain ethyl 2, 2-difluoro-3- benzoyloxy-3- (2,2-dimethyldioxolan-4-yl)-propionate 9. The isoalkyllidene pretecting group of 9 is selectively removed , e.g. by using a strong acid such as concentrated sulfuric acid in ethanol, to produce ethyl-2,2-difluoro-3-benzoyloxy-4,5- dihydroxypentanoate 9A. the product is cylcised to lactone 10 and converted to the dibenxoate ester to produce the lactone 2 - deoxy-2,2-difluropentofuranos-1-ulose-3,5-dibenzoate 11 as a mixture of erythro and threo isomers. The 374 patent describes isolating at least a portion of the erytliro isomer from the mixture by selective precipitation. See also , chou et al systhesis, 565- 570, compound 11 is then reduced to obtain a mixture of alph, and beta anomers of 2-desoxy-2,2-difluoropentofuranose- debenzoate 12,which is activated with methanesulfonyl chloride to obtain an anomeric mixture of mesylates, 2-deoxy-2,2- difluoro-D-ribofuranosyl-3,5-di-O-benzoyl-1-O-[beta]- methanesulfonate 13, and coupled with N,O-bis(trimethylsilyl)- cytosine 8 to obtain the debenzoate ester of the si Iy I -protected nucleoside 14 as mixture of the [alpha]of the [alpha]-and [beta]- anomers ( about a 1 :1[alpha]/beta anomer ratio). Removal of the esters and silyl protecting group- provides a mixture of the [beta]-[anomer (gemcitabine) and the [alpha]-anomer (about1 :1[alpha]/[beta] anomer ratio). The 374 patent describes selectively isolating the [beta]-anomer ( gemcitabine) by forming a salt of the anomeric mixture ,e.g. the hydrochloride or hydeobromide salt, and selectively precipitating to obtain 2'- deoxy2',2'-difluorocytidine as the salt in 1 :4[alpha]/[beta] ratio. The 374 patent also describes selectively precipitating the [beta]-anomer in free base form in slightly basic aqueous solution. one such process involves dissolving the 1:1[alpha]/[beta] anomeric mixture in hot acidic water (PH adjusted to 2,5-5.0 ) and , once the mixture is substantially dissolved, increasing the PH to 7 - 9 and allowin the solution to cool, to produce srystals , which are isolated by filtration.

Processes for separating anomeric mixture of alkylsulfonate intermediates also have been described US patent No., s 5256797 and 4526988 describes processes for separating anomers of 2-deoxy-2,2-difluoro-D-ribofuranosyl-1- alkylsulfonates, and US patent No. 5256798 describes a process for obtaining [alpha]-anomers-enriched ribofuranosyl sulfonates. Others intermediates that mau useful for preparing gemcitabine have benn disclosed . for instance, US patent No. 5480992 describes anomeric mixture of 2,2-diluoroibosyl azide and corresponding amine intermediates that can be prepared, e.g by reacting a 2-deoxy-2,2-nucleophile, such as lithium azide.to obtain the azide, reduction of the azide produces the corresponding amine, which can be synthetically converted into a nucleoside, see also us patent No. 5541345, 5594155.

2006 / 070985 and WO 2006/071090, which describes about for preparation for the gamcitabine have been reported that the stereoselective preparation of 2'-deoxy-2',T-difluorocytidine the process comprises activating the 2'-deoxy-2'2'-difluoro-D- ribofuranosyl-3,5-diester 12 A with diphenyl chlorophosphate followed by isomer separation, then obtaining the corresponding 1-bromo-ribofuranose intermediate, which is coupled withN, O-bis(trimethylsilyl)-cytosine 8, then deprotected to obtain gemcitabine.However, this process requires an additional step as compared to the process outlined in scheme 2, which makes it less attractive for industrial implementation. There are ineherent problems associated with the production of gemcitabine, particularly for processes that requires the production and separation of isomers, which tend to afford poor yields on a commercial scale.Accordingly there is a need for improved methods pf preparing gemcitabine and intermediates thereof, which facilitate the production of gemcitabine, particularly on commercial scale. The present invention provides such methods and intermediates, as will be apparent from the description of the invention provided herein.

Therefore the present invention relates to an improved process for the prepation of Gemcitabine , in which the isolation of pure gemcitabine using the catalyst and the ion resine has not been reported in the prior art]

\ Objects of the present invention:-

The important object of the present invention is that the above said prior art is suffer from anyone or more of the following limitations e.g. that the utilization of expensive hydroxyl protective group like tert- butyldimethylsilyl group and reducing agents like diisobutyl aluminium hydride, lithium aluminum hydride, which , moreover ar hazardous , requiring special handling care,

Furhte object of the present invention is that the utilization of multiple protection and deprotection steps not only increasing the cost and risk of manufacture in the prior art.

Further important object of the present invention is that the prior art which utilization of high boiling solvents and elavted reaction temperatures necessitating high energy consumption,

Summery of the invention:-

The present invention provides for the preparation of an improved process of preparing gemcitabine or a salt thereof, which comprising the following steps, removing at least a substantial portion of the [alpha] anomer from an anomeric mixture of N-l-pretected-2'deoxy-2%2'- difluorocytidine-3%5'-diester to produce a product the resultant product which is enriched in the [beta] anomer followed by removing the 3'- ester, the 5'-ester and the N-protecting group; and optinally converting the product into a salt.

The preparation of the phosphate salt as the protecting group and the use of ion exchange resin for the isolation and the preparation of the hydrochloride salt conveniently is the major focus of this invention. The protected 2-deoxy-2,2-difluoro-D-erythro-pentofuranos-l-ulose-3,5- diesters of the present invention are particularly useful for the commercial scale production of gemcitabine. These D-erythro derivatives are crystalline materials, and can be purified by separation from a mixture of the D-erythro and L-threo isomers, e.g. by precipitation. The intermediates and methods of the present invention facilitate the pure synthesis of gemcitabine, and are easier to use and produce higher than processes reported in the prior art.

Detailed description of the invention:

The present invention relates to a process for preparing the gemcitabine or a salt thereof, comprising the following steps reacting 2-Deoxy-2,2- difluoro-D-ribofuranose-3,5-dibenzoate dissolved in a organic solvent and allowed to cool to 0 to IOdegree centigradeand stirred for a period of 1 to 2 hours to this triethylamine is added for a period of 2 to 10 minutes, further to this an indigenous protecting compound is added under constant stirring, more particularly a chlorotricresyl phosphoryl chloride is selected, after adding the protecting group the solvents were distilled off, and to the residual volume IN HCL is added until to form a clear solution adjusted the PH of about 2, further to this an ethyl acetate was added followed by washing the organic phase using NaHCo3, water and brine solution, the organic phase was dried over MgSo4 , concentrated under reduced pressure to obtain a crude 2-deoxy-2,2- difluoro-D-riboufuranose-3,5-dibenzoate-l-arylateas an oil phase. Which can be stated in step - 1, under step - 11 process the above product is dissolved in in dry methanol under constant stirring using Ion exhange resin for period of 15 to 25 hours at 30 degree centigrade to 45 degree centigrade temperature, which gives pure gemcitabine hydrochloride.

The above reaction mechanism is as follow,

2- Dαnty-2^- difluor o-D- 2- Deoxy-2^2-difluoro-3^-dib enzoate- rib ofuranose-3^- dib enzoate 1-arytate (pG)

Gemcitabine hydrochloride

Example:-

The following example demonstrates the preparation of crude 2-deoxy-2,2-difluoro-D- riboufuranose-3 ,5-dibenzoate-1-arylate.

To a 500 ml round flask was added crude 2-deoxy-2,2-difluoro-D- erythro- pentofuranos-l-ulose-3,5-dibenzoate ( 23.36 mmol), which was dissolved in dry THF (100 ml) and cooled to 0-5C. Triethylamine ( 12.1 mmol) was added over a period of 5 minutes, after which time the mixture was stirred for another 1 hour. Chlorotricresyl phosphoryl chloride (23.36 mol) was added with stirring. Most of the solvents were distilled off, and to the residual volume (about 30 ml) IN HCI was added (50 ml) thus forming a clear solution with phi value of about 2. Ethyl acetate (100 ml) was added and the organic phase was washed with 5% NaHCO3 (50 ml), water (50 ml), and brine (50 ml). The ethyl acetate phase was dried over MgSO4 and concentrated under reduced pressure to obtain crude 2-deoxy-2,2-difluoro-D- riboufuranose- 3,5-dibenzoate-1-arylate as an oil. Yield: 8.52g, (19.8 mmol, 84.8%, [alpha]/[beta] isomer ratio about 1/1).

In a 100 ml round flask, crude Step 1 ( 5.8mmol) was dissolved in dichloromethane (20 ml), and bis(trimethylsilyl)acetyl cytosine (0.7g, 6.9 mmol) was added. Triflate (0.79g, 6.9 mmol) was added drop wise while cooling to 0-5C. The mixture was stirred for another 1 hour, and washed with IN HCI (15 ml), 5% NaHCO3 (15 ml), and dried over MgSO4. The reaction mixture was stirred with an "Indion" Ion Exchange Resin to yield a product Step 2 Yield: 2.82 g, (5.55 mmol), 95.7%, purity 99.2%.

lna 100 ml flask, the Step 2 product was diisolved in dry methanol 250 ml and stirred with an "Indion" Ion Exchange Resin 30 gms at 40C for 18-20 hours to afford Gemcitabine hydrochloride. Yield: 5 g, 50.5% yield. Purity: 98%, ee 95

Claims

We Claim,

1. An improved process for preparing gemcitabine or salts thereof, comprising the following steps, dissolved 2-Deoxy-2,2-difluoro-D- ribofuranose-3,5-dibenzoate in an organic solvent and allowed to cool to 1 to 10 degree centigrade temperature, allowed to stirred for a period of 1 to 2 hours added triethylamine for a period of 2 to 20 minutes, indigenous protecting compound added under constant stirring, solvents were distilled off, and to the residual volume HCL is added until to form a clear solution adjusted the PH, further to this an ethyl acetate was added followed by washing the organic phase using NaHCo3, water and brine solution, the organic phase was dried over MgSo4 , concentrated the above reaction mixture under reduced pressure to obtain a crude 2-deoxy-2,2-difluoro-D-riboufuranose-3,5-dibenzoate-l- arylateas an oil phase, above product is dissolved in dry methanol under constant stirring using Ion exhange resin for a period of 15 to 25 hours at 30 to 45 degree centigrade temperature, which gives pure gemcitabine hydrochloride,

2. A process as claimed in claim 1, wherein the the indigenous protecting compound is chlorotricresyl phosphoryl chloride,

3. An improved process for preparing gemcitabine or salts thereof, such as herein described with reference to an example.