WO2023067623A1

WO2023067623A1 - A short and cost effective synthetic route towards anti viral eidd-1931 and its hydrate polymorphs

Info

Publication number: WO2023067623A1
Application number: PCT/IN2022/050929
Authority: WO
Inventors: Jubi JOHN; Sunil VARUGHESE; Radhakrishnan KOKKUVAYIL VASU; Ajayaghosh AYYAPPANPILLAI; Jagadeesh Krishnan; Divya INDIRA SOMAN; Abhijith BALAN; Akhil KRISHNAN RADHAKRISHNAPILLAI; Priyadarshini THOPPE SIVAKUMAR
Original assignee: Council Of Scientific And Industrial Research An Indian Registered Body Incorporated Under The Regn. Of Soc. Act (Act Xxi Of 1860)
Priority date: 2021-10-20
Filing date: 2022-10-18
Publication date: 2023-04-27

Abstract

EIDD 1931 is a broad-spectrum antiviral showing promising activities against disease causing RNA viruses such as influenza, SARS, MERS and COVID-19. The present invention intends to disclose the development of a process towards this antiviral via a short synthetic route starting from D-ribofuranose 1,2,3,5-tetraacetate. The present invention also decreases the usage of purification processes such as column chromatography and makes the process a green alternative. The present invention provides a cost effective and industrially viable process for preparing EIDD-1931 which involves use of cheap raw materials.

Description

A SHORT AND COST-EFFECTIVE SYNTHETIC ROUTE TOWARDS ANTI VIRAL EIDD-1931 AND ITS HYDRATE POLYMORPHS

FIELD OF THE INVENTION

[0001] The present invention relates to a short and cost-effective process for the synthesis of a broad- spectrum antiviral EIDD 1931 of Formula I. Particularly, the present invention relates to a process for the synthesis of a broad-spectrum antiviral drug EIDD 1931, identification, isolation, and characterization of its two hydrated polymorphic crystal forms.

Formula I

BACKGROUND AND PRIOR ART OF THE INVENTION

[0002] The most severe pandemic in history is the ‘Spanish flu’ or ‘1918 influenza pandemic’ caused by an H1N1 virus which infected 500 million people claiming lives of at least 50 million. More than 30 million people have lost their lives since 1980’s after HIV infection was characterized as a severe condition. Every year 2-3 new viruses are identified which causes diseases in human beings. Out of these newly identified viruses, RNA viruses mainly of zoonotic origin pose a huge threat to mankind such as Nipah, Ebola and MERS with 60%, 50% and 36% mortality rates respectively. The recent outbreak of corona virus disease (COVID-19) has affected 217 countries causing infection in approximately 160 million individuals claiming more than 3.4 million lives in a short time. The alarming fact is that there are no drugs available which can cure most of the above-mentioned viral infections.

[0003] EIDD 1931 is an experimental antiviral originally developed by Emory University which was proven to be active against a range of RNA viruses such as chikungunya virus, Venezuelan equine encephalitis virus (VEEV), respiratory syncytial virus (RSV), hepatitis C virus, norovirus, influenza A and B viruses, and Ebola virus. The researchers at Emory recently have shown that this molecule was active against MERS-CoV by inhibiting the RNA dependent RNA polymerase. This observation has triggered the research on checking whether this molecule would be active against CO VID- 19. They also claim that the prodrug form EIDD 2801 (Molnupiravir) could be taken orally which would be an added advantage compared to Remdesivir which is being administered intravenously. (Reference may be made to: (a) Painter et al. (WO2019113462A1); (b) Agostini M. L.; et al. J. Virol., 2019, 93, e01348-19; (c) Urakova N.; et al. J. Virol., 2017, 92, e01965-17; (d) Ehteshami M.; et al. Antimicrob. Agents Chemother., 2017, 61, e02395-16; (e) Costantini V. P.; et al. Antivir. Ther., 2012, 17, 981-991; (f) Yoon J-J.; et al. Antimicrob. Agents Chemother., 2018, 62,1427; (g) Reynard O.; et al. Viruses, 2015, 7, 6233-6240; (h) Stuyver L. J.; et al. Antimicrob. Agents Chemother. 2003, 47, 244-254.) This drug is currently in phase 2 clinical trials in USA and UK.

[0004] WO2019113462A1 discloses the preparation of EIDD 1931 (broad spectrum antiviral) from an advanced intermediate uridine as a three-step process. The first step of the synthesis involved the protection of the 2’, 3’ and 5’ hydroxyls of the ribose ring with TBDMS group. In the second step, the installation of hydroxylamine moiety was carried out. The final step furnished EIDD 1931 after TBDMS -deprotection with EuN.HF. Very recently a route to synthesize EIDD 1931 was reported starting from another advanced intermediate cytidine (Vasudevan N.; et al. Chem. Commun., 2020, 56, 13363-13364 and doi.org/10.26434/chemrxiv.12818327.vl). The two processes mentioned above required the use of advanced intermediates as starting materials, expensive reagents/enzymes and also the intermediates were purified by extensive column chromatography throughout.

[0005] Though there have been synthetic routes to produce the antiviral EIDD 1931, there is an unmet need for a novel synthesis route for the production of drug in higher yields starting from an economical and viable source. The present situation demands for a process which also reduces the usage of column chromatography for the purification of intermediates obtained.

ABBREVIATIONS USED

HMDS - hexamethyldisilazane

BSTFA - N,O-(bistrimethylsilyl)acetamide

TMSOTf - Trimethylsilyl trifluoromethanesulfonate

CH3CN - Acetonitrile

SOCh - Thionylchloride

CHCI3 - Chloroform

NH2OH.HCI - Hydroxylamine hydrochloride

EtsN - Triethylamine

MeOH - Methanol

KHSO4 - Potassium bisulphate

DCM - Dichloromethane

DCE - 1 ,2-Dichloroethane

TLC - Thin layer chromatography

NMR - Nuclear Magnetic Resonance

HRMS - High Resolution Mass Spectroscopy

SXRD - Single crystal X-ray diffraction

PXRD - Powder X-ray Diffraction

DSC - Differential Scanning Calormetry

TG - Thermogravimetry

OBJECTIVES OF THE INVENTION

[0006] Main objective of the present invention is to provide a short, industrially viable and cost-effective process for the synthesis of broad spectrum antiviral EIDD 1931. [0007] Another object of the present invention is to provide a process for the synthesis of a broad-spectrum antiviral drug EIDD 1931 from D-Ribofuranose 1 ,2,3,5- tetraacetate.

[0008] Yet another object of the present invention is to provide industrially viable and cost-effective process which utilizes cheap raw materials that are available in plenty in comparison to the prior art.

[0009] Another object of the present invention is to provide two hydrate polymorphs of EIDD 1931 and establish their differing thermal stability behavior.

SUMMARY OF THE INVENTION

[0010] In an aspect of the present invention, there is provided a process for the synthesis of anti-viral EIDD-1931 of Formula I and its hydrated polymorphic forms

Formula I comprising the steps of: i. glycosylation of uracil (1.2 equivalent) with compound of Formula 1 (1.0 equivalent) in the presence of A,O-(bistrimethylsilyl)acetamide (BSTFA, 2.0 equivalent) and trimethylsilyl trifluoromethanesulfonate (TMSOTf, 1.2 equivalent) in acetonitrile to obtain triacetylated uridine of Formula 2;

Formula 1 Formula 2 ii. treating compound of Formula 2 (1.0 equivalent) as obtained in step (i) with thionyl chloride (SOCh, 12.0 equivalent) in the presence of dimethylformamide (DMF, 0.75 equivalent) in chloroform to obtain an intermediate compound of Formula 3 followed by treating with hydroxylamine hydrochloride (NH2OH.HCI, 5.0 equivalent) in the presence of EtaN (trimethylamine, 6.0 equivalent) in methanol (MeOH) to obtain antiviral EIDD 1931 of Formula I; and

Formula 3 Formula I iii. treating crude compound of Formula 2 (1.0 equivalent) as obtained in step (i) with hexamethyldisilazane (HMDS, 7.0 equivalent), Imidazole (0.5 equivalent), potassium hydrogen sulphate (KHSO4, 2.1 equivalent) and hydroxylamine hydrochloride (NH2OH.HCI, 1.1 equivalent) at temperature in the range of 80 to 85 °C for a period in the range of 36 to 48 hours to obtain triacetyl -/V-hydroxy-cytidine of Formula 4 followed by treating with excess of ammonia in methanol at room temperature in the range of 25 to 30°C to obtain antiviral EIDD- 1931 of Formula I.

Formula 4

[0011] In another aspect of the present invention, there is provided a process for the selective formation of Form-I of EIDD 1931, comprising the steps of: i. dissolving EIDD 1931 in an organic solvent followed by slow evaporation for crystallization and isolating Form I of EIDD 1931.

[0012] In yet another embodiment of the present invention, an organic solvent used is selected from the group consisting of methanol, ethanol, actonitrile, tetrahydrofuran or 1,4-dioxane. [0013] In one another aspect of the present invention there is provided a process for preparation of Form- II of EIDD 1931 comprising the steps of: a) dissolving EIDD 1931 in an organic alcohol; b) diffusing the vapour of ether into the alcoholic solution; and c) isolating the concomitantly formed crystal forms (Form-I and Form II), based on the morphology.

[0014] In yet another embodiment of the present invention, organic solvent used is methanol.

[0015] In yet another embodiment of the present invention, Crystalline forms (Form I and II) of the monohydrated EIDD 1931 were characterized by X-ray diffraction and thermal analysis.

[0016] These and other features, aspects, and advantages of the present subject matter will be better understood with reference to the following description and appended claims. This summary is provided to introduce a selection of concepts in a simplified form. This summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used to limit the scope of the claimed subject matter.

BRIEF DESCRIPTION OF THE FIGURES

[0017] Figure 1 illustrates the glycosylation of uridine with //-D-ribofuranose 1, 2,3,5- tetraacetate, in accordance with an embodiment of the present disclosure.

[0018] Figure 2 illustrates the one -pot synthesis of EIDD 1931 from triacetyl uridine by installation of a hydroxylamine moiety and deprotection of acetyl groups, in accordance with an embodiment of the present disclosure.

[0019] Figure 3 illustrates an alternate one -pot synthesis of EIDD 1931 from triacetyl uridine by installation of a hydroxylamine moiety and deprotection of acetyl groups, in accordance with an embodiment of the present disclosure.

[0020] Figure 4 illustrates (a) and (b) the synthon variations, tautomerism and conformational differences in the hydrate polymorphic forms Form I and II respectively, (c) overlap image showing the difference in the torsional shifts, in accordance with an embodiment of the present disclosure.

[0021] Figure 5 illustrates the powder diffraction plots of the crystal forms, the distinct peak position and relative intensity highlights the structural differences in the polymorphs, in accordance with an embodiment of the present disclosure.

[0022] Figure 6 (a) and (b) illustrate the differential scanning calorimetric (DSC) thermogram of the polymorphs Form I and II respectively, in accordance with an embodiment of the present disclosure.

[0023] Figure 7 (a) and (b) illustrates the thermogravimetric TG plots of the crystal forms Form I and II respectively, in accordance with an embodiment of the present disclosure.

DETAILED DESCRIPTION OF THE INVENTION

[0024] Those skilled in the art will be aware that the present disclosure is subject to variations and modifications other than those specifically described. It is to be understood that the present disclosure includes all such variations and modifications. The disclosure also includes all such steps, features, compositions, and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations of any or more of such steps or features.

Definitions

[0025] For convenience, before further description of the present disclosure, certain terms employed in the specification, and examples are delineated here. These definitions should be read in the light of the remainder of the disclosure and understood as by a person of skill in the art. The terms used herein have the meanings recognized and known to those of skill in the art, however, for convenience and completeness, particular terms and their meanings are set forth below.

[0026] The articles “a”, “an” and “the” are used to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. [0027] The terms “comprise” and “comprising” are used in the inclusive, open sense, meaning that additional elements may be included. It is not intended to be construed as “consists of only”.

[0028] Throughout this specification, unless the context requires otherwise the word “comprise”, and variations such as “comprises” and “comprising”, will be understood to imply the inclusion of a stated element or step or group of element or steps but not the exclusion of any other element or step or group of element or steps.

[0029] The term “including” is used to mean “including but not limited to”. “Including” and “including but not limited to” are used interchangeably.

[0030] The term “at least one" is used to mean one or more and thus includes individual components as well as mixtures/combinations.

[0031] For the purpose of the present invention, the term “TLC” is used to refer to the analytical technique thin layer chromatography which is used to separate non-volatile mixtures. It is performed on an analytical scale as a means of monitoring the progress of a reaction, or on the preparative scale to purify small amounts of a compound. The technique is based on the different affinities of various components in a mixture with the adsorbant stationary phase such as coated over an inert substrate like glass.

[0032] The term “NMR” or “Nuclear Magnetic Resonance” is used to refer to the spectroscopic technique to observe the magnetic field around an atomic nuclei. The sample is subjected to radio waves in magnetic field to produce nuclear magnetic resonance due to the nuclear excitation in sample.

[0033] The terms “HRMS” and “High Resolution Mass Spectroscopy” are used to refer to the spectroscopic technique to analyse the molecules in trace amounts in the vaporised samples to achieve accurate measurements of masses and isotope identification.

[0034] For the purpose of the present disclosure, the term “SXRD” is used to refer to the single crystal X-ray diffraction method is a non-destructive analytical technique wherein the x-ray is irradiated on a crysalline solid sample to analyse the internal lattice, unit cell dimentions. [0035] The term PXRD or Powder X-ray Diffraction is used to refer to the technique used for the structural characterization of a sample wherein x-ray is irradiated on the powdered samples to analyse the purity and crystallinity of the samples.

[0036] The terms “DSC” and “Differential Scanning Calorimetry” are used to refer to the thermoanalytical technique wherein the difference in the amount of heat required to increase the temperature of sample and reference is measured as a function of temperature.

[0037] The terms “TG” and “Thermogravimetry” are used to refer to the thermoanalytical technique in which the mass of a sample is measured over time as the temperature changes to analyse the thermal stability of the compound.

[0038] Ratios, concentrations, amounts, and other numerical data may be presented herein in a range format. It is to be understood that such range format is used merely for convenience and brevity and should be interpreted flexibly to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub-range is explicitly recited.

[0039] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the disclosure, the preferred methods, and materials are now described. All publications mentioned herein are incorporated herein by reference.

[0040] The present disclosure is not to be limited in scope by the specific embodiments described herein, which are intended for the purposes of exemplification only. Functionally equivalent products, compositions, Formulations, and methods are clearly within the scope of the disclosure, as described herein.

[0041] As discussed in the background, the existing processes for the synthesis of the antiviral EIDD1931 required the use of advanced intermediates as starting materials, expensive reagents/enzymes and also the intermediates were to be purified by extensive column chromatography throughout.

[0042] Thus, there is an unmet need for a novel synthesis route for the production of drug in higher yields starting from an economical and viable source though there have been synthetic routes to produce the antiviral EIDD 1931. Accordingly the present disclosure provides a process for preparing antiviral EIDD 1931 which is economically viable and also reduces the usage of column chromatography for the purification of intermediates obtained. The present invention discloses a novel process towards EIDD 1931 synthesis and its polymorphic forms. Upon extensive investigations related to the synthetic routes towards the broad spectrum antiviral EIDD 1931, the present invention describes that the molecule could be made in fewer number of chemical transformations starting from cheap and basic starting materials.

[0043] The present invention provides an industrially viable and cost-effective process which utilizes cheap raw materials that are available in plenty in comparison to the prior art. The present invention also decreases the usage of purification processes such as column chromatography after individual steps in comparison to the prior art which makes the process a green alternative.

[0044] The present invention intends to offer a process which was finalized after extensive optimizations of individual chemical transformations. This has led to high yielding reactions that has increased the overall yield of the process.

[0045] In an embodiment of the present invention, there is disclosed a short and cost- effective process for the synthesis of broad- spectrum antiviral EIDD 1931 from /TD- ribofuranose 1 , 2, 3 , 5 -tetraacetate .

[0046] In another embodiment of the document there is provided a synthesis of EIDD 1931 starting from //-D-ribofuranose 1,2,3,5-tetraacetate. The first step involves the glycosylation of uridine with //-D-ribofuranose 1,2,3,5-tetraacetate to generate triacetyl -uridine 2 with a yield of 94% as shown in Figure 1.

[0047] In another embodiment of the present invention, there is provided a one-pot conversion of Formula 2 to EIDD 1931. The crude product 2 was taken directly to the next step. The second step involves the chlorination of amide carbonyl moiety of uracil in triacetyl -uridine with SOCh and DMF in CHCI3 to synthesize intermediate 3. In the same pot, the hydroxylamine installation and acetyl-group deprotection was carried out by treating 3 with NH2OH.HCI and EtsN to furnish EIDD 1931 with a yield of 84% which is depicted in Figure 2.

[0048] In another embodiment of the present invention there is provided an alternate one-pot conversion of 2 to EIDD 1931. This route involves the treatment of crude triacetyl-uridine 2 with HMDS, imidazole, KHSO4 and NH2OH.HCI to furnish the intermediate triacetyl-/V-hydroxy-cytidine 4. In the same pot, the acetyl-group deprotection was carried out by treating 4 with excess ammonia in MeOH to furnish EIDD 1931 with a yield of 94% (Figure 3). This route was devoid of purification by column chromatography. The present invention describes two synthetic routes towards EIDD 1931 with overall yields of 79% and 88% starting from />-D-ribofuranose 1 ,2,3,5- tetraacetate.

[0049] In the 2^nd route no column purification was done. All the other reaction mixtures were worked up or distilled and the crude was taken directly to the next step.

[0050] The present invention further identified, isolated and characterized the structural and thermal stability profile of two monohydrate crystal forms. The crystal forms occur concomitantly and have distinct morphology. Form-I exist as long blocks, while, Form- II has a sugar-like cubical shape.

[0051] In an embodiment of the present invention there is provided a process for the synthesis of an anti-viral EIDD-1931 of Formula I and its hydrated polymorphic forms

Formula I i) glycosylation of uracil (1.2 equivalent) with compound of Formula 1 (1.0 equivalent) in the presence of A,O-(bistrimethylsilyl)acetamide (BSTFA, 2.0 equivalent) and tnmethylsilyl tnfluoromethanesulfonate (TMSOTf, 1.2 equivalent) in acetonitrile to obtain triacetylated uridine of Formula 2;

Formula 1 Formula 2 ii) treating the compound of Formula 2 (1.0 equivalent) as obtained in step (i) with thionyl chloride (SOCh, 12.0 equivalent) in the presence of dimethylformamide (DMF, 0.75 equivalent) in chloroform to obtain an intermediate compound of Formula 3 followed by treating with hydroxylamine hydrochloride (NH2OH.HCI, 5.0 equivalent) in the presence of Et₃N (trimethylamine, 6.0 equivalent) in methanol (MeOH) to obtain the antiviral EIDD 1931 of Formula I; and

Formula 3 Formula I iii) treating the crude compound of Formula 2 (1.0 equivalent) as obtained in step (i) with hexamethyldisilazane (HMDS, 7.0 equivalent), Imidazole (0.5 equivalent), potassium hydrogen sulphate (KHSO4, 2.1 equivalent) and hydroxylamine hydrochloride (NH2OH.HCI, 1.1 equivalent) at temperature in a range of 80 to 85 °C for a period in a range of 36 to 48 hours to obtain triacetyl-/V-hydroxy-cytidine of Formula 4 followed by treating with excess of ammonia in methanol at room temperature in a range of 25 to 30°C to obtain the antiviral EIDD- 1931 of Formula I.

[0052] In an embodiment of the present invention there is provided a process for the selective formation of Form-I of EIDD 1931, comprising the steps of dissolving EIDD 1931 in an organic solvent followed by slow evaporation for crystallization and isolating Form I of EIDD 1931. In another embodiment, wherein the organic solvent is selected from the group consisting of methanol, ethanol, actonitrile, tetrahydrofuran, and 1,4-dioxane.

[0053] In an embodiment of the present invention there is provided a process for preparation of Form- II of EIDD 1931, comprising the steps of: d) dissolving EIDD 1931 in an organic alcohol; e) diffusing the vapour of ether into the alcoholic solution; and f) isolating the concomitantly formed crystal forms (Form-I and Form II), based on the morphology

[0054] In another embodiment of the present invention, the organic solvent used is methanol.

[0055] In an embodiment of the present invention there is provided a process for the selective formation of Form-I of EIDD 1931, comprising the steps of dissolving EIDD 1931 in an organic solvent selected from the group consisting of methanol, ethanol, actonitrile, tetrahydrofuran, and 1,4-dioxane followed by slow evaporation for crystallization and isolating Form I of EIDD 1931.

[0056] In an embodiment of the present invention there is provided a process for the selective formation of Form-I of EIDD 1931, comprising the steps of dissolving EIDD 1931 in methanol followed by slow evaporation for crystallization and isolating Form I of EIDD 1931.

[0057] In yet another embodiment of the present invention, crystalline forms (Form I and II) of the monohydrated EIDD 1931 were characterized by X-ray diffraction and thermal analysis.

[0058] In yet another embodiment of the present invention, the polymorphic forms of EIDD, prepared by the vapour diffusion method, exhibit a rare combination of tautomeric, synthon and conformational polymorphism. The monohydrate crystal forms describe a notable difference in the role of lattice water in the structure formation, thereby leading to distinct thermal stability. The thermal studies and slurry experiments confirms that the two forms are monotropically related. The corresponding unit cell dimensions are:

Form-I: a = 5.164(6), b = 8.466(1), c = 13.440(2) A, ft = 92.62(2)°, V= 587.1(1), P2i Form-II: a = 19.679(7), b = 5.448(2), c = 11.332(4) A, ? = 105.4(1)°, V = 1171.2(7), C2

Form-I and Form-II differ in their primary synthons, conformations and also exist as tautomeric polymorphs as illustrated in Figure 4. The thermal profile depicted in Figure 5 and 6, and slurry experiments show that the crystal forms are monotropically related. [0059] Although the subject matter has been described in considerable detail with reference to certain examples and implementations thereof, other implementations are possible.

EXAMPLES

[0060] The disclosure will now be illustrated with working examples, which is intended to illustrate the working of disclosure and not intended to take restrictively to imply any limitations on the scope of the present disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice of the disclosed methods and compositions, the exemplary methods, devices, and materials are described herein. It is to be understood that this disclosure is not limited to the particular methods, and experimental conditions described, as such methods and conditions may apply.

EXAMPLE 1

Synthesis of triacetyl-uridine 2

[0061] In a solution of uracil (2.1 g, 18.86 mmol) in dry acetonitrile (100 ml) under argon atmosphere in a two neck RB flask equipped with a magnetic stirrer and condenser A,O-(bistrimethylsilyl) acetamide (9.27 ml, 34.56 mmol) was added. This was allowed to stir at 65 C for 45 minutes to result in the formation of a clear solution. Then the reaction was allowed to cool to room temperature to which a solution of /TD- ribofuranose 1,2,3,5-tetraacetate 1 (5g, 15.72 mmol) in acetonitrile (10 ml) was added followed by the addition of trimethylsilyl trifluoromethanesulfonate (3.40 ml, 18.85 mmol). Further, the reaction was stirred at 65 °C for 3 hours. After the completion of the reaction, as observed from TLC, the mixture was diluted with ethyl acetate, washed with water (2 x 100 ml), saturated NaHCCh (2 xlOO ml) solution and brine (1 x 40ml). The aqueous layers were back-extracted with ethyl acetate. The combined organic layer was dried and concentrated to give the desired compound as a white amorphous solid (5.47g, 94%).

Chemical Formula: C15H118N2O9, White solid; M.P: 106-108 °C, ¹H NMR (500 MHz, CDCI3) 5 8.69 (s, ^XH), 7.33 (d, J = 8.2 Hz, 1H), 5.97 (d, J = 4.4 Hz, 1H), 5.73 (d, J = 8.1 Hz, 1H), 5.28 - 5.26 (m, 2H), 4.31 - 4.26 (m, 3H), 2.08 (s, 3H), 2.07 (s, 3H), 2.04 (s, 3H) ppm. ¹³C NMR (126 MHz, CDCI3) 5 170.14, 169.66, 162.50, 150.03, 139.29, 103.41, 87.49, 79.96, 72.74, 70.19, 63.13, 20.80, 20.53, 20.44 ppm.

Example 2

Synthesis of EIDD 1931 (Route 1)

[0062] To a Schenck flask under argon, 5.0 g of triacetyl uridine (13.51 mmol) and 125 mL of dry CHCI3 was added. Freshly double distilled (over 20% quinolone and 40% linseed oil) thionyl chloride (11.76 ml, 162.10 mmol) followed by dry DMF (785 pL, 10.13 mmol) was added to this solution at room temperature. This mixture was then refluxed for 6 hours. After evaporation of the solvent under vacuum, the residue was co-evaporated with dry toluene (2 x 125 ml). This residue containing crude compound 3 was used for the next step without further purification.

[0063] The above residue was dissolved in 60 ml of dry methanol in an RB flask equipped with a magnetic stirrer and condenser under argon. To this solution, triethylamine (11.30 ml, 81.06 mmol) followed by hydroxylamine hydrochloride (4.69 g, 67.55 mmol) were added and allowed to reflux for 18 hours. After removal of the solvent under vacuum the crude was purified by flash chromatography on silica gel (EtOAc:MeOH, 95:5) to yield EIDD 1931 (2.94 g. 84%) as white solid.

Chemical Formula: C9H13N3O6, White solid; M.P: 113-115 °C, ¹H NMR (500 MHz, MeOD) 5 7.16 (d, J = 8.2 Hz, 1H), 5.86 (d, J = 5.6 Hz, 1H), 5.62 (d, J = 8.2 Hz, 1H), 4.15 (dt, J = 9.0, 5.2 Hz, 2H), 3.95 (d, J= 3.0 Hz, 1H), 3.79 (dd, J= 12.0, 2.2 Hz, 1H), 3.70 (dd, J = 12.1, 3.1 Hz, 1H).

¹³C NMR (125 MHz, CDCI3) 5 170.31, 169.78, 169.74, 149.39, 144.59, 129.75, 99.57, 86.52, 79.44, 77.29, 77.04, 76.78, 71.93, 70.38, 63.39, 20.81, 20.57, 20.47. HRMS (ESI) (M+H)⁺ : calculated for C15H19N3O9 is 386.1200; found 386.1218.

Example 3

Synthesis of EIDD 1931 (Route 2)

[0064] Imidazole (0.460 g, 6.76 mmol) in HMDS (20 ml, 95.4 mmol) under argon was stirred until a clear solution was formed (around 30 min.) at 85 °C in a 250 ml Schlenk flask. Then the solution was cooled to room temperature followed by addition of potassium hydrogen sulphate (4.60 g, 33.78 mmol), and was stirred at 85 °C for 30 minutes. The reaction mixture was then cooled to room temperature. Hydroxylamine hydrochloride (1.17 g, 16.89 mmol) and triacetyl uridine (5.00 g, 13.51 mmol) were introduced into it. The reaction mixture was allowed to stir for 48 hours at 85 °C, then cooled to ambient temperature, to which water (30.00 ml) was added, and the mixture was extracted with ethyl acetate (3x20 ml). The combined organic layer was washed with brine (1x20 ml) and dried over Na2SC>4 and concentrated under vacuum. The residue was used for the next reaction without further purification.

[0065] The above residue was dissolved in 20ml of ammonia in methanol and stirred for 12 h at room temperature. The crude product was recrystallized using methanol and dichloromethane mixture. The product was obtained as a white solid (3.5 g, 94%).

Example 4. Crystallization of EIDD 1931 [0066] Solid-forms of EIDD 1931 were screened by slow evaporation and vapor diffusion methods.

[0067] Slow evaporation of near saturated solution of EIDD 1931 in organic solvents such as methanol, ethanol, actonitrile, tetrahydrofuran or 1,4-dioxane yielded the crystals of Form I exclusively. In a typical experiment, 50 mg of the compound was dissolved in 2-5 mL of respective solvents by warming over a hotplate. The resultant clear solution was filtered and kept for slow evaporation. The crystals appeared within 2-5 days.

[0068] Vapor diffusion of diethyl ether into saturated solution of EIDD 1931 in methanol or ethanol (2 mL) at ambient temperature and pressure conditions yielded the crystals of the polymorphs (Form-I and Form II) concomitantly. Based on their morphological differences, crystals were identified and manually separated under a polarizing stage microscope. Each crystal forms were analyzed by X-ray diffraction and thermal (DSC and TG) methods. Slurry experiments were done in hydrocarbon solvent (hexane and cyclohexane) wherein the crystal forms were insoluble and was agitated for 24 hours before the diffraction studies. DSC and TG experiments were performed from room temperature to 120 °C under inert atmorphere.

ADVANTAGES OF THE INVENTION

[0069] The present invention provides a novel short, industrially viable and cost- effective process for the synthesis of an anti-viral EIDD 1931. The synthetic route utilizes cheap raw materials which are easily available. Synthesis of EIDD 1931 from glycoside intermediate avoids usage of intermediate purification. The present invention exhibits better yielding than prior art. The present disclosure provides a process which decreases the usage of purification processes such as column chromatography after each indivdual steps in comparison to the prior art. The two novel crystal forms have distinct structural characteristics and thermal stability profile.

Claims

I/We claim:

1. A process for the synthesis of an anti-viral EIDD-1931 of Formula I and its hydrated polymorphic forms

Formula I comprising the steps of: i. glycosylation of uracil (1.2 equivalent) with compound of Formula 1 (1.0 equivalent) in the presence of A,O-(bistrimethylsilyl)acetamide (BSTFA,

2.0 equivalent) and trimethylsilyl trifluoromethanesulfonate (TMSOTf, 1.2 equivalent) in acetonitrile to obtain triacetylated uridine of Formula 2;

Formula 1 Formula 2 ii. treating the compound of Formula 2 (1.0 equivalent) as obtained in step (i) with thionyl chloride (SOCh, 12.0 equivalent) in the presence of dimethylformamide (DMF, 0.75 equivalent) in chloroform to obtain an intermediate compound of Formula 3 followed by treating with hydroxylamine hydrochloride (NH2OH.HCI, 5.0 equivalent) in the presence of Et₃N (trimethylamine, 6.0 equivalent) in methanol (MeOH) to obtain the antiviral EIDD 1931 of Formula I; and

Formula 3 Formula I iii. treating the crude compound of Formula 2 (1.0 equivalent) as obtained in step (i) with hexamethyldisilazane (HMDS, 7.0 equivalent), Imidazole (0.5 equivalent), potassium hydrogen sulphate (KHSO4, 2.1 equivalent) and hydroxylamine hydrochloride (NH2OH.HCI, 1.1 equivalent) at temperature in a range of 80 to 85 °C for a period in a range of 36 to 48 hours to obtain triacetyl-/V-hydroxy-cytidine of Formula 4 followed by treating with excess of ammonia in methanol at room temperature in a range of 25 to 30°C to obtain the antiviral EIDD-1931 of Formula I.

Formula 4 A process for the selective formation of Form-I of EIDD 1931 as claimed in claim 1, comprising the steps of: i. dissolving EIDD 1931 in an organic solvent followed by slow evaporation for crystallization and isolating Form I of EIDD 1931. The process as claimed in claim 2, wherein the organic solvent is selected from the group consisting of methanol, ethanol, actonitrile, tetrahydrofuran, and 1,4- dioxane. A process for preparation of Form-II of EIDD 1931 as claimed in claim 1, comprising the steps of: a) dissolving EIDD 1931 in an organic alcohol; b) diffusing the vapour of ether into the alcoholic solution; and c) isolating the concomitantly formed crystal forms (Form-I and Form II), based on the morphology. The process as claimed in claim 2, wherein organic solvent used is methanol. Crystalline forms (Form I and II) of the monohydrated EIDD 1931 were characterized by X-ray diffraction and thermal analysis.