Compositions comprising 2 '-deoxy-2', 2 '-difluorotetrahydrouridine

Abstract

The present invention relates to a composition comprising 2 '-deoxy-2', 2 '-difluorotetrahydrouridine. The present invention relates to a composition comprising Compound 1 or a salt thereof wherein the composition comprises less than 7 measurable impurities wherein there is no impurity at a level greater than 1.0 wt% of the composition wherein Compound 1 has a purity of at least 99% by weight of the composition, and wherein the composition comprises impurities at a level of at least 0.25 wt% of the composition. # imgabs0 #

Description

[0001] This application is a divisional application of the patent application with the application number 202080070847.0, the invention name of "High-Purity 2'-Deoxy-2',2'-Difluorotetrahydrouridine and Its Manufacturing Method", and the application date of October 7, 2020.

[0002] Priority Claim

[0003] This application claims the benefit of U.S. Provisional Application Serial No. 62 / 912,317, filed October 8, 2019, the entire content of which is incorporated herein by reference. Technical Field

[0004] The present invention relates to a method for synthesizing 2'-deoxy-2',2'-difluorotetrahydrouridine with increased purity and a uniform particle size distribution. In particular, the method of the present invention includes providing synthetic reaction intermediates and crystallization and separation procedures for the final compound in a high-purity form. Background Art

[0005] Several important chemotherapeutic compounds are analogs of the nucleotide cytidine, including decitabine, gemcitabine, 5-azacytidine, ara-C, tiazofurin, 5-fluoro-2'-deoxycytidine, and cytchlor. As analogs of cytidine, these compounds are degraded by the enzyme cytidine deaminase (CDA), which degrades the compounds into inactive metabolites. The presence of CDA limits the effectiveness of cytidine analogs, requiring higher and / or more frequent doses of the analogs to achieve therapeutic benefits.

[0006] One way to overcome this problem is to co-administer a CDA inhibitor with the cytidine analog, thereby blocking the degradation of the analog. One class of CDA inhibitors is the 2'-deoxy-2',2'-difluorotetrahydrouridine compounds. U.S. Patent No. 8,268,800 (incorporated herein by reference in its entirety) discloses such compounds, including Compound 1:

[0007]

[0008] There is a need for a more efficient method for producing CDA inhibitors, such as 2'-deoxy-2',2'-difluorotetrahydrouridine, for use in methods for treating cancer and other disorders. Summary of the Invention

[0009] The present invention relates to the development of a more efficient method for synthesizing 2'-deoxy-2',2'-difluorotetrahydrouridine compounds and intermediates involved in the synthesis. Previous synthetic methods in the art have been inconvenient due to the use of inefficient transfer hydrogenation processes and the use of preparative HPLC to separate the final compounds. The inventors of the present invention have developed a synthetic method with improved efficiency, which enables the purification of the final compound by precipitation or crystallization, such as crystallization-induced diastereoselective transformation (CIDT), which converts a mixture of epimers into the desired compound and thus results in an increased yield of the desired epimer (see WO 2015 / 066162). The present invention improves the method by reducing impurities, improving yields, shortening reaction times, and / or otherwise improving the conditions for industrial-scale production.

[0010] Another aspect of the present invention relates to a method for producing compound 1 (named (4R)-1-[(2R,4R,5R)-3,3-difluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl]-4-hydroxytetrahydropyrimidin-2(1H)-one)) or a salt thereof:

[0011]

[0012] comprising the following steps:

[0013] (a) Hydrogenating a starting compound of formula IV:

[0014]

[0015] wherein R is a hydroxy protecting group, to produce a compound of formula IIa:

[0016]

[0017] (b) Reducing the compound of formula IIa to produce a compound of formula IIIa:

[0018]

[0019] (c) Deprotecting the compound of formula IIIa to produce compound 2:

[0020] and

[0021] (d) Precipitating or crystallizing compound 2 in the presence of a catalyst to produce compound 1:

[0022]

[0023] or a salt thereof;

[0024] wherein the method comprises one or more of the following, in any combination:

[0025] (i) wherein the hydrogenation step (a) is carried out using a palladium catalyst in a hydrogen atmosphere;

[0026] (ii) wherein the reduction step (b) is carried out at a temperature of about -12 °C to about -5 °C;

[0027] (iii) wherein the deprotection step (c) is carried out in the presence of an organic base; and / or

[0028] (iv) wherein the work-up of the deprotected compound from the deprotection step (c) is carried out under non-aqueous conditions. In some embodiments, the present invention further comprises a step of recrystallizing the final product, and the recrystallization is carried out at a pH of about 6.0 to about 7.4 and is carried out by dissolving the final product at a temperature of about 50 °C to about 55 °C to produce a solution and then cooling the solution to about 5 °C.

[0029] Another aspect of the present invention relates to compound 1 of high purity (e.g., compound 1 having a purity of at least about 80%, such as at least about 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9%):

[0030]

[0031] or a salt thereof. High purity compound 1 can be produced by the method of the present invention. Other aspects of the present invention relate to compound 1 comprising a lower number of impurities and / or a lower level of individual and total impurities compared to compound 1 produced by existing methods. Detailed Description

[0032] The present invention will now be described hereinafter with reference to the accompanying examples, which illustrate embodiments of the present invention. However, the present invention may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.

[0033] Unless otherwise defined, 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 invention belongs. The terms used in the description of the present invention herein are for the purpose of describing particular embodiments only and are not intended to limit the present invention.

[0034] Unless the context otherwise requires, the various features of the present invention described herein are specifically intended to be used in any combination. In addition, the present invention also contemplates that in some embodiments of the present invention, any feature or combination of features set forth herein may be excluded or omitted. For illustration, if the specification states that a composition comprises components A, B, and C, it is specifically intended that any one or combination of A, B, or C may be omitted and waived, either singly or in any combination.

[0035] For the purposes of this document, if there is any ambiguity between the written chemical name and the drawn chemical structure, the drawn chemical structure will be controlling.

[0036] Definitions

[0037] As used herein, "a", "an", or "the" can mean one or more than one. For example, a compound can mean a single compound or multiple compounds.

[0038] Also as used herein, "and / or" refers to and encompasses any and all possible combinations of one or more of the associated listed items, as well as the absence of a combination when interpreted as an alternative ("or").

[0039] As used herein, the term "about", when referring to a measurable value such as an amount of a dose (e.g., an amount of a compound), means to encompass variations of ±10%, ±5%, ±1%, ±0.5%, or even ±0.1% of the specified amount.

[0040] As used herein, the terms "comprise", "comprises", and "comprising" specify the presence of the stated features, integers, steps, operations, elements, and / or components, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and / or groups thereof.

[0041] As used herein, the transitional phrase "consisting essentially of" means that the scope of a claim should be interpreted to cover the specified materials or steps recited in the claim, and those that do not materially affect one or more of the basic and novel features of the claimed invention. Thus, when used in a claim or the description of the present invention, the term "consisting essentially of" is not intended to be interpreted as equivalent to "comprising".

[0042] As used herein, the terms "increase", "increases", "increased", "increasing", and like terms denote an elevation of at least about 25%, 50%, 75%, 100%, 150%, 200%, 300%, 400%, 500%, or more.

[0043] As used herein, the terms "reduce", "reduces", "reduced", "reduction" and like terms mean a decrease of at least about 5%, 10%, 15%, 20%, 25%, 35%, 50%, 75%, 80%, 85%, 90%, 95%, 97% or more. In a particular embodiment, the reduction results in no or substantially no (i.e., a negligible amount, e.g., less than about 10% or even 5%) detectable activity or amount.

[0044] The term "its salts" includes pharmaceutically acceptable salts. The term "pharmaceutically acceptable salts" shall mean non-toxic salts of the compounds used in the present invention, which are generally prepared by reacting a free acid with a suitable organic or inorganic base or a free base with a suitable organic or inorganic acid. Examples of such salts include, but are not limited to, acetate, benzenesulfonate, benzoate, bicarbonate, bisulfate, bitartrate, borate, bromide, calcium, calcium edetate, camsylate, carbonate, chloride, clavulanate, citrate, dihydrochloride, edetate, edisylate, estolate, esylate, fumarate, gluceptate, gluconate, glutamate, glycollylarsanilate, hexylresorcinate, hydrabamine, hydrobromide, hydrochloride, hydroxynapthoate, iodide, isothionate, lactate, lactobionate, laurate, malate, maleate, mandelate, mesylate, methyl bromide, methyl nitrate, methyl sulfate, mucate, naphthalenesulfonate, nitrate, oleate, oxalate, pamoate, palmitate, pantothenate, phosphate / diphosphate, polygalacturonate, potassium, salicylate, sodium, stearate, subacetate, succinate, tannate, tartrate, teoclate, tosylate, triethiodide, and valerate.

[0045] As used herein, the term "Bronsted-Lowry base" refers to a substance having the ability to accept a proton.

[0046] As used herein, the term "hydroxy protecting group" can be any suitable hydroxy protecting group, i.e., a labile chemical moiety known in the art for protecting a hydroxy group from unwanted reactions during synthetic procedures. After one or more of said synthetic procedures, the protecting group (blocking group) as described herein can be selectively removed. See, for example, A. Isidro-Llobet et al., Amino Acid-Protecting Groups, Chem. Rev. 109:2455–2504 (2009) and T. Greene and P. Wuts, Protective Groups in Organic Synthesis (3d Ed. 1999). In some embodiments, the hydroxy protecting group is an acid-stable hydroxy protecting group. Examples of hydroxy protecting groups include, but are not limited to, alkyl, cycloalkyl, arylalkyl, aryl, ether, ester, cyclic ether, cyclic ester, acetal, cyclic acetal, ketal, and cyclic ketal groups, etc., which can be removed under acidic or basic conditions, thereby removing the protecting group and replacing it with a hydrogen atom. Specific hydroxy protecting groups include, but are not limited to, methyl, ethyl, acetate, ethyl acetate, propionate, ethylene glycol, propylene glycol, 4-methoxybenzyl, benzyl, trityl, trimethylsilyl, tetrahydropyranyl, and benzoyl. Other hydroxy protecting groups include, but are not limited to, methoxymethyl (MOM), methylthiomethyl (MTM), tert-butylthiomethyl, (phenyldimethylsilyl)methoxymethyl (SMOM), benzyloxymethyl (BOM), p-methoxybenzyloxymethyl (PMBM), (4-methoxyphenoxy)methyl (p-AOM), guaiacolmethyl (GUM), tert-butoxymethyl, 4-pentenoxymethyl (POM), silyloxymethyl, 2-methoxyethoxymethyl (MEM), 2,2,2-trichloroethoxymethyl, bis(2-chloroethoxy)methyl, 2-(trimethylsilyl)ethoxymethyl (SEMOR), tetrahydropyranyl (THP), 3-bromotetrahydropyranyl, tetrahydrothiopyranyl, 1-methoxycyclohexyl, 4-methoxytetrahydropyranyl (MTHP), 4-methoxytetrahydrothiopyranyl, 4-methoxytetrahydrothiopyranyl S,S-dioxide, 1-[(2-chloro-4-methyl)phenyl]-4-methoxypiperidin-4-yl (CTMP), 1,4-dioxan-2-yl, tetrahydrofuranyl, tetrahydrothiofuranyl, 2,3,3a,4,5,6,7,7a-octahydro-7,8,8-trimethyl-4,7-methano-benzofuran-2-yl, 1-ethoxyethyl, 1-(2-chloroethoxy)ethyl, 1-methyl-1-methoxyethyl, 1-methyl-1-benzyloxyethyl, 1-methyl-1-benzyloxy-2-fluoroethyl, 2,2,2-trichloroethyl, 2-trimethylsilylethyl, 2-(phenylseleno)ethyl, tert-butyl, allyl, p-chlorophenyl, p-methoxyphenyl, 2,4-dinitrophenyl, benzyl, p-methoxybenzyl, 3,4-dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl, p-halobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl, p-phenylbenzyl, 2-pyridylmethyl, 4-pyridylmethyl, N-oxid 3-methyl-2-picolyl (3-methyl-2-picolyl N-oxido), diphenylmethyl, p,p'-dinitrodiphenylmethyl, 5-dibenzosuberyl, triphenylmethyl, α-naphthyldiphenylmethyl, p-methoxyphenyldiphenylmethyl, bis(p-methoxyphenyl)phenylmethyl, tris(p-methoxyphenyl)methyl, 4-(4'-bromobenzoylmethyloxyphenyl)diphenylmethyl, 4,4',4”-tris(4,5-dichlorophthalimidophenyl)methyl, 4,4',4”-tris(acetoacetyloxyphenyl)methyl, 4,4',4”-tris(benzoyloxyphenyl)methyl, 3-(imidazol-1-yl)bis(4',4”-dimethoxyphenyl)methyl, 1,1-bis(4-methoxyphenyl)-1'-pyrenylmethyl, 9-anthryl, 9-(9-phenyl)xanthenyl, 9-(9-phenyl-10-oxo)anthryl, 1,3-benzodithiolan-2-yl, S,S-dioxidobenzisothiazolyl, trimethylsilyl (TMS), triethylsilyl (TES), triisopropylsilyl (TIPS), dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS), dimethylethylsilyl, tert-butyldimethylsilyl (TBDMS), tert-butyldiphenylsilyl (TBDPS), tribenzylsilyl, tri-p-xylylsilyl, triphenylsilyl, diphenylmethylsilyl (DPMS), tert-butylmethoxyphenylsilyl (TBMPS), formate, benzoylformate, acetate, chloroacetate, dichloroacetate, trichloroacetate, trifluoroacetate, methoxyacetate, triphenylmethoxyacetate, phenoxyacetate, p-chlorophenoxyacetate, 3-phenylpropionate, 4-oxopentanoate (levulinate), 4,4-(ethylenedithio)pentanoate (acetoacetyldithioacetal), pivalate, adamantoate, crotonate, 4-methoxycrotonate, benzoate, p-phenylbenzoate, 2,4,6-trimethylbenzoate (mesitoate), alkyl methyl carbonate, 9-fluorenylmethyl carbonate (Fmoc), alkyl ethyl carbonate, alkyl 2,2,2-trichloroethyl (Troc), 2-(trimethylsilyl)ethyl carbonate (TMSEC), 2-(phenylsulfonyl)ethyl carbonate (Psec); 2-(triphenylphosphine)ethyl carbonate (Peoc), alkyl isobutyl carbonate, alkyl vinyl carbonate, alkyl allyl carbonate, alkyl p-nitrophenyl carbonate, alkyl benzyl carbonate, alkyl p-methoxybenzyl carbonate, alkyl 3,4-dimethoxybenzyl carbonate, alkyl o-nitrobenzyl carbonate, alkyl p-nitrobenzyl carbonate, alkyl S-benzyl thiocarbonate, 4-ethoxy-1-naphthyl carbonate, methyl dithiocarbonate, 2-iodobenzoate, 4-azidobutyrate, 4-nitro-4-methylvalerate, o-(dibromomethyl)benzoate, 2-formylbenzenesulfonate, 2-(methylthiomethoxy)ethyl, 4-(methylthiomethoxy)butyrate, 2-(methylthiomethoxymethyl)benzoate, 2,6-dichloro-4-methylphenoxyacetate, 2,6-dichloro-4-(1,1,3,3-tetramethylbutyl)phenoxyacetate, 2,4-bis(1,1-dimethylpropyl)phenoxyacetate, chlorodiphenylacetate, isobutyrate, monosuccinate, (E)-2-methyl-2-butenoate, o-(methoxycarbonyl)benzoate, α-naphthoate, nitrate, alkyl N,N,N',N'-tetramethylphosphoryldiamide, alkyl N-phenylcarbamate, borate, dimethylthiophosphinyl, alkyl 2,4-dinitrophenylsulfinate, sulfate, methanesulfonate (mesylate), benzylsulfonate and toluenesulfonate (Ts). Methods for protecting and deprotecting hydroxy groups are well known and can be found, for example, in Protective Groups in Organic Synthesis (T. Green and P. Wuts; 3rd Edition; John Wiley and Sons, 1999).

[0047] The bond identified by is present or absent.

[0048] The bond identified by is a bond that includes a mixture of stereochemical isomers.

[0049] As used herein, the term "g / g" refers to grams per gram of substrate. The substrate is defined as 1 equivalent in each step, and all other aspects in that step are defined relative to the substrate.

[0050] The term "enantiomer" refers to stereoisomers of a compound that are mirror images of each other and are non-superimposable. In this application, unless otherwise mentioned or indicated, the chemical designation of a compound represents a mixture of all possible stereochemical isomeric forms.

[0051] The term "diastereomer" refers to stereoisomers of a compound (and thus not enantiomers) that have different configurations at one or more stereocenters but are not mirror images of each other.

[0052] The term "epimer" refers to two diastereomers that differ from each other at only one stereocenter.

[0053] The term "alkyl" denotes a straight-chain or branched hydrocarbon chain containing 1 to 12 carbon atoms, such as 1 to 6 or 1 to 4 carbon atoms. Examples of alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, and the like.

[0054] The term "aryl" refers to an aromatic 5- to 8-membered monocyclic or 8- to 12-membered bicyclic system. The term also includes aromatic bicyclic systems in which one, two, or three ring carbons in one ring have had hydrogen atoms added (e.g., partially saturated rings). Examples of aryl groups include phenyl, naphthyl, and the like.

[0055] The term "acyl" denotes an alkyl or aryl group attached to a carbonyl group. Examples of acyl groups include formyl, acetyl, propionyl, acryloyl, benzoyl, and the like.

[0056] As used herein, the term "benzoyl" refers to the acyl group of benzoic acid (attached through the carbonyl carbon) and has the following structure.

[0057]

[0058] Compound

[0059] One aspect of the present invention relates to Compound 1 (or a salt thereof) produced by the method of the present invention, particularly the free base form of Compound 1 having high purity (e.g., epimeric purity or low amounts of impurities, solvents, reaction by-products, and / or degradation products). Another aspect of the present invention relates to Compound 1 having a purity of at least about 80% by weight, such as at least about 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% or higher. In some embodiments, Compound 1 comprises less than about 20% by weight, such as less than about 15%, 10%, 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2% or 0.1% of impurities, solvents, reaction by-products, and / or degradation products, such as the impurities shown in Tables 1 and 4 below. In some embodiments, Compound 1 comprises less than about 20% by weight, such as less than about 15%, 10%, 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2% or 0.1% of its corresponding enantiomers and / or epimers of Compound 1. Another aspect of the present invention relates to Compound 1 having a molar ratio of the desired epimer (Compound 1) to another epimer (Compound 6) of at least about 60:40 by weight, such as at least about 70:30, 80:20, 90:10, 95:5, 96:4, 97:3, 98:2, 99:1 or higher, such as an epimeric purity of at least about 60%, such as at least about 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or more, such as 99.5% or 99.9%.

[0060] In some embodiments, Compound 1 (or a salt thereof) produced by the method of the present invention comprises less than 20 different measurable impurities, such as less than 15 impurities, such as less than 15, 14, 13, 12, 11, 10, 9, 8, 7 or 6 impurities, such as 5 or fewer impurities. As used herein, a measurable impurity is an impurity that can be detected by methods routinely used in the art for testing chemical purity (e.g., HPLC or mass spectrometry). A measurable impurity is an impurity present in an amount greater than 0.03 wt%.

[0061] In some embodiments, Compound 1 (or a salt thereof) produced by the method of the present invention comprises less than about 4.0% by weight of epimeric Compound 6, such as less than about 3.5%, 3.0%, 2.5%, 2.0%, 1.9%, 1.8%, 1.7%, 1.6%, 1.5%, 1.4%, 1.3%, 1.2%, 1.1% or 1.0%.

[0062] In some embodiments, Compound 1 (or a salt thereof) produced by the method of the present invention comprises less than about 2.5% by weight of total impurities excluding epimeric Compound 6, such as less than about 2.0%, 1.9%, 1.8%, 1.7%, 1.6%, 1.5%, 1.4%, 1.3%, 1.2%, 1.1%, 1.0%, 0.9%, 0.8%, 0.7%, 0.6% or 0.5%.

[0063] In some embodiments, Compound 1 (or a salt thereof) produced by the method of the present invention comprises less than about 1.0% by weight of any single impurity, including or excluding epimeric Compound 6, for example, less than about 1.0%, 0.95%, 0.9%, 0.85%, 0.8%, 0.75%, 0.7%, 0.65%, 0.6%, 0.55%, 0.5%, 0.45%, 0.4%, 0.35%, 0.3%, 0.25%, 0.2%, 0.15% or 0.1%.

[0064] Certain compounds described herein contain one or more chiral centers or may otherwise be capable of existing as a variety of stereoisomers. The scope of the present invention includes pure stereoisomers as well as mixtures of stereoisomers, such as purified enantiomers / diastereomers / epimers, enantiomer / diastereomer / epimer-enriched mixtures, or racemates. In some embodiments, the compound has a stereochemical purity of at least about 80%, such as at least about 85%, 90%, 95%, 96%, 97%, 98%, 99% or higher.

[0065] In certain cases, the compounds of the present invention may also exist as tautomers, such as amide / iminol tautomers. Although only one delocalized resonance structure may be described, all such forms are contemplated within the scope of the present invention.

[0066] As described above, the compounds disclosed herein can be prepared in the form of their pharmaceutically acceptable salts. Pharmaceutically acceptable salts are salts that retain the desired biological activity of the parent compound and do not impart undesired toxicological effects. Examples of such salts are (a) acid addition salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid, etc.; and salts formed with organic acids such as, for example, acetic acid, oxalic acid, tartaric acid, succinic acid, maleic acid, fumaric acid, gluconic acid, citric acid, malic acid, ascorbic acid, benzoic acid, tannic acid, palmitic acid, alginic acid, polyglutamic acid, naphthalenesulfonic acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalenedisulfonic acid, polygalacturonic acid, etc.; (b) salts formed from elemental anions such as chlorine, bromine, and iodine, and (c) salts derived from bases such as ammonium salts, alkali metal salts such as sodium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, and salts with organic bases such as dicyclohexylamine and N-methyl-D-glucamine. In one embodiment, the compounds disclosed herein are prepared in the form of the free base.

[0067] It should also be understood that the compositions herein include the compounds and combinations with stoichiometric or non-stoichiometric amounts of water (as in hydrates) or other components (as in solvates).

[0068] Synthetic methods

[0069] Another aspect of the present invention relates to a method for producing Compound 1 or a salt thereof:

[0070]

[0071] comprising precipitating or crystallizing Compound 1 from a solution of Compound 2 in the presence of a catalyst:

[0072]

[0073] . In one embodiment, the method yields Compound 1.

[0074] This method utilizes crystallization-induced diastereoselective transformation (CIDT) to provide an enhancement in the production of the desired epimer (Compound 1). Any suitable catalyst can be used in this method. As used herein, the term "catalyst" with respect to the precipitation or crystallization step refers to a compound that promotes the equilibrium between Compound 6 and Compound 1 when present in a sub-stoichiometric amount relative to Compound 2. Without being limited by mechanism, it is believed that Compound 1 and its epimer Compound 6 are in equilibrium with the open aldehyde structure of the intermediate compound. The catalyst is believed to act by promoting the opening of Compound 6 to the aldehyde form, thereby increasing the conversion of one epimer to the other and balancing the amounts of Compound 1 and Compound 6 in solution, as Compound 1 preferentially precipitates or crystallizes out of solution when using an appropriate solvent. The catalyst is present in a catalytically effective amount. In some embodiments, the catalyst can be an acid, such as an inorganic acid, such as an organic acid, such as acetic acid or trifluoroacetic acid. In other embodiments, the catalyst can be a base, such as a Bronsted-Lowry base, such as a weak base (a base that is not completely ionized in an aqueous solution). In other embodiments, the catalyst can be diisopropylethylamine or ammonium hydroxide. In some embodiments, the base has a basicity of 10 or higher in a solvent. In some embodiments, the base has a pKa of 10 or greater in a solvent such as DMSO, as reported in, for example: Bordwell, Acc. Chem. Res. 21:456 1988); Crampton, J. Chem. Res. (S) 22 (1997); Kaliurand et al., J. Org. Chem. 65(19):6202 (2000); Kaljurand et al, J. Org. Chem. 70(3):1019 (2005). In some embodiments, the catalyst is a strong base. In some embodiments, the catalyst is a strong base, such as a sterically hindered strong base, such as a strong base that is a weak nucleophile. In some embodiments, the catalyst is 1,8-diazabicyclo(5.4.0)undec-7-ene (DBU). DBU can be present in any effective amount, such as from about 1 mol% to about 20 mol%, such as from about 2 mol% to about 15 mol%, such as from about 5 mol% to 10 mol%, such as about 5 mol%, or such as about 10 mol%, such as about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 mol%. In some examples, DBU can be present in an amount from 1 mol% to 20 mol%, such as from 2 mol% to 15 mol%, such as from 5 mol% to 10 mol%, such as 5 mol%, or such as 10 mol%, such as, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 mol%.

[0075] Any solvent or solvent combination that causes compound 1 to precipitate or crystallize preferentially over compound 6 can be used. In one embodiment, the solvent is a solvent in which compound 6 has a greater solubility than compound 1. In some embodiments, the solution used to form the solution of compound 2 comprises an organic solvent, consists essentially of an organic solvent, or consists of an organic solvent. In some embodiments, the solution comprises water or an aqueous solvent, consists essentially of water or an aqueous solvent, or consists of water or an aqueous solvent. In some embodiments, the solvent is a protic solvent. In some embodiments, the solvent is a water-miscible solvent. In a specific embodiment, the solution is acetonitrile, acetone, tetrahydrofuran, dimethyl sulfoxide, or methanol. In a specific embodiment, the solution is an aqueous solution of acetonitrile, an aqueous solution of acetone, an aqueous solution of tetrahydrofuran, an aqueous solution of dimethyl sulfoxide, or an aqueous solution of methanol. In a specific embodiment, the solution is an aqueous solution of acetonitrile.

[0076] The precipitation or crystallization can be carried out for a sufficient length of time to form a suitable amount of compound 1, for example, about 0.5 days to 14 days, for example, about 1 - 4 days, for example, about 2 - 3 days, for example, about 3 - 10 days, for example, about 4 - 6 days. The precipitation or crystallization can be carried out at any suitable temperature, for example, at about room temperature, optionally followed by a temperature of about 0°C to about 10°C. After the precipitation or crystallization is complete, the precipitate can be collected, for example, by filtration, and washed, for example, with an aqueous solution of acetonitrile and / or acetonitrile, optionally cooled to a temperature of about 0°C to about 10°C. The precipitate can then be dried, for example, under vacuum, for example, at a temperature below about 45°C. The progress of the reaction can be monitored, for example, by sampling the supernatant of the reaction mixture and determining the ratio of compound 1 to compound 6. Completion of the reaction is indicated by the presence of a 50:50 mixture of compound 1 and compound 6 in the supernatant. If this ratio is not reached, additional catalyst can be added and the reaction continued until completion.

[0077] After precipitation or crystallization, compound 1 is optionally further purified by recrystallization or slurrying, for example, from an aqueous solution of acetonitrile, optionally with the addition of an acid, such as trifluoroacetic acid. For example, the precipitate can be resuspended in a water:acetonitrile ratio of about 1:2 to about 1:10 (v / v), heated to about 35 - 45°C, and then cooled to about 0°C. The resulting precipitate can be washed in a water:acetonitrile ratio of about 1:2 to about 1:10 (v / v), and then washed in acetonitrile, optionally cooled to a temperature of about 0°C to about 10°C. In certain embodiments, compound 1 is optionally further purified by other methods known in the art, such as HPLC.

[0078] In some embodiments, Compound 1 is further purified by recrystallization from acetone and water, and the pH is adjusted to about 6.0 to about 7.4 (e.g., about 6, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7, 7.1, 7.2, 7.3, or 7.4 or any value or range therein), for example, using an aqueous formic acid solution alone or in combination with an aqueous DBU solution. In some embodiments, the mixture is further heated, for example, to a temperature of about 50 °C to about 55 °C, for example, at a rate of 0.5 to 1.5 °C / min, until complete dissolution is observed, filtered, washed with a mixture of acetone and water, and then cooled, for example, to a temperature of about 33 °C to about 43 °C, for example, at a rate of 0.1 to 0.5 °C / min, then seeded and further cooled, for example, to a temperature of about -10 °C to about 0 °C, for example, at a rate of 0.1 to 0.5 °C / min, and allowed to stand, and then further preparation known in the art is carried out (e.g., filtration, washing, and drying). In some embodiments, the recrystallization mixture is further heated to about 53 °C, clarified and filtered, and then cooled to a temperature of about 35 °C, seeded, cooled to a temperature of about -5 °C, and allowed to stand (e.g., aged) for about 12 to about 16 hours (e.g., about 12, 13, 14, 15, or 16 hours or any value or range therein). The inventors of the present invention have found that specific recrystallization parameters as disclosed herein, such as higher temperatures and / or specific pH ranges, produce higher purity products, improve the particle size distribution, and avoid dimer impurities. In some embodiments, the method produces a compound with a more uniform particle size distribution, such as a unimodal particle size distribution, for example, a particle size distribution centered at about 100 - 200 μm (e.g., about 130 μm). In some embodiments, at least 50% of the particles have a size of 20 μm to 300 μm, for example, at least 60%, 70%, 80%, or 90%.

[0079] After precipitation or crystallization, the molar ratio of Compound 1 (i.e., the desired epimer) to the epimer of Compound 1 (e.g., Compound 6) can be at least about 60:40, for example, at least about 70:30, 80:20, 90:10, 95:5, or 98:2 or higher. After a second purification step (e.g., recrystallization or slurrying), the molar ratio of the desired epimer (Compound 1) to the other epimer (Compound 6) can be at least about 80:20, for example, at least about 90:10, 95:5, or 98:2 or higher.

[0080] Another aspect of the present invention relates to a method for producing Compound 1 or a salt thereof:

[0081]

[0082] comprising the following steps:

[0083] (a) Hydrogenate the starting compound of formula IV:

[0084]

[0085] wherein R is a hydroxyl protecting group,

[0086] to produce the compound of formula IIa:

[0087]

[0088] (b) Reduce the compound of formula IIa to produce the compound of formula IIIa:

[0089]

[0090] (c) Deprotect the compound of formula IIIa to produce compound 2:

[0091] and

[0092] (d) Precipitate or crystallize compound 2 in the presence of a catalyst to produce compound 1:

[0093]

[0094] or a salt thereof;

[0095] wherein the method comprises one or more of the following, in any combination:

[0096] (i) wherein the hydrogenation step (a) is carried out using a palladium catalyst in a hydrogen atmosphere;

[0097] (ii) wherein the reduction step (b) is carried out at a temperature of about -12 °C to about -5 °C;

[0098] (iii) wherein the deprotection step (c) is carried out in the presence of an organic base; and / or

[0099] (iv) wherein the work-up of the deprotected compound from the deprotection step (c) is carried out under non-aqueous conditions. In some embodiments, the present invention further comprises a step of recrystallizing the final product, and the recrystallization is carried out at a pH of about 6.0 to about 7.4 and is carried out by dissolving the final product at a temperature of about 50 °C to about 55 °C to produce a solution and then cooling the solution to about 5 °C.

[0100] The starting compound of formula IV can be obtained commercially, for example, from Aurora Fine Chemicals (San Diego, CA), or synthesized by known methods, such as those disclosed in Wheeler et al., J. Labeled Compounds Radiopharm. 29:583 (1991) and Chou et al., Synthesis 6:565 (1992), which are incorporated herein by reference in their entirety.

[0101] The hydrogenation of the compound of formula IV to produce the compound of production IIa in step (a) can be carried out by methods known in the art, for example, as disclosed in U.S. Patent No. 8,268,800 and / or Patent Publication WO 2015 / 066162, the disclosures of which are incorporated herein by reference in their entirety. For example, this step can be carried out under catalytic transfer hydrogenation conditions, for example, in the presence of palladium on carbon (Pd / C), for example, in the presence of about 5% Pd / C. The amount of the catalyst such as Pd / C used in the hydrogenation step (a) can be any catalytically effective amount, for example, a catalytic amount not exceeding 0.1 part by weight per 1 part by weight of the compound of formula IV. In some embodiments, the amount of Pd / C used in the hydrogenation step (a) can be, for example, about 0.025 to about 0.05 parts of Pd / C per 1 part by weight of the compound of formula IV. The hydrogenation can be carried out by heating the compound of formula IV to reflux, for example, with formic acid and hydrochloric acid or with hydrogen (for example, at a hydrogen pressure of about 2 to about 4 bar) and optionally with acetic acid) in a solvent such as an aqueous solution of ethyl acetate. The hydrogenation can be carried out at a temperature of about 0 °C to about 100 °C, for example, about 50 °C to about 80 °C, for example, about 63 °C to about 77 °C, for example, about 68 °C for about 0.5 - 48 hours, for example, about 5 to about 24 hours, about 10 to about 20 hours, about 15 to about 20 hours, or any value or range therein, for example, about 24 hours. The reagents used to effect the hydrogenation (such as palladium and carbon) can be added after bringing the reaction mixture to an elevated temperature (for example, about 50 °C to about 80 °C, or about 68 °C). After completion, the catalyst can be removed, for example, by filtration at, for example, about 60 °C to about 70 °C, and washed, for example, with ethyl acetate. After separation of the organic layer, it can be washed with, for example, an aqueous solution of potassium carbonate, an aqueous solution of sodium bicarbonate, and / or an aqueous solution of NaCl. The volume of the organic layer can be reduced (for example, by distillation), and the residue can be heated (for example, at about 70 °C) until dissolved and cooled (for example, to about 45 °C to about 55 °C), seeded with the product and stirred, for example, for about 1 h, for example, at about 45 °C to about 55 °C. The volume of the reaction mixture can be reduced (for example, by distillation), and methyl tert-butyl ether can be slowly added at a temperature of about 40 °C to about 50 °C, and then the reaction mixture can be slowly cooled to about 0 °C to about 10 °C. The resulting suspension can be stirred, for example, for about 2 to about 16 h, and then filtered. The filtrate can be washed with, for example, methyl tert-butyl ether and optionally cooled to about 0 °C to about 10 °C.

[0102] Alternatively, the filtrate from the hydrogenation reaction can be washed (for example, with acetic acid), heated (for example, to about 80 °C) to dissolve and water (for example, preheated to about 80 °C) can be added. After cooling, the precipitate can be collected by filtration, washed (for example, with water and ethanol) and dried.

[0103] The solvent used for the hydrogenation step (a) in this article can be any conventional solvent that does not cause adverse effects in the reaction. Non-limiting examples of such solvents for hydrogenation include water, alcohols (such as methanol, ethanol, isopropanol, n-butanol, trifluoroethanol, ethylene glycol), ethers (such as tetrahydrofuran, dioxane, diethyl ether, diglyme), esters (such as methyl acetate, ethyl acetate), other organic solvents, and / or a mixed solvent of two or more solvents as disclosed herein. The reaction can preferably be carried out in the presence of organic acids such as formic acid, acetic acid, oxalic acid, maleic acid, fumaric acid, malic acid, tartaric acid, citric acid, succinic acid, and / or benzoic acid. In some embodiments, the hydrogenation can be carried out in a mixed solvent comprising ethyl acetate, acetic acid, and water.

[0104] In some embodiments, the hydrogenation can be carried out at an increased pressure (relative to ambient pressure), for example, an increased pressure of about 0.1 to about 1 megapascal (MPa), such as about 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, 0.95, 0.96, 0.97, 0.98, 0.99, or 1 MPa or any value or range therein. In some embodiments, the hydrogenation step (a) can be carried out at ambient pressure (e.g., atmospheric pressure, e.g., normal pressure). In some embodiments, the hydrogenation step (a) can be carried out at an increased pressure of about 0.1 to about 0.85 MPa, or about 0.15 to about 0.73 MPa, or about 0.1 to about 0.5 MPa.

[0105] The reduction of the compound of formula IIa to the compound of formula IIIa in step (b) can be carried out by methods known in the art, for example, as disclosed in U.S. Patent No. 8,268,800 and / or Patent Publication WO 2015 / 066162. For example, the reduction can be carried out with a reducing agent such as sodium borohydride in an organic solvent such as a mixture of dichloromethane and ethanol. The reduction can be carried out at any suitable temperature, for example, about -5°C to about 10°C, such as about 0°C to about 5°C for about 0.5 to 3 hours, such as about 1.5 hours.

[0106] In some embodiments, the reduction can be carried out at a temperature of from about -12 °C to about -3 °C, such as about -12, -11, -10, -9, -8, -7, -6, -5, -4 or -3 °C or any value or range therebetween. In some embodiments, the reduction can be carried out at a temperature of from about -11 °C to about -3 °C, from about -12 °C to about -5 °C, from about -11 °C to about -5 °C or from about -10 to about -6 °C, or at a temperature of about -8 °C. The present invention is in part based on the unexpected finding that unusually cold conditions (e.g., a temperature of from about -11 °C to about -5 °C) minimize the formation of impurities such as but not limited to DPU and DCU and indirect CYU (which is produced by the conversion of DCU in subsequent steps) that are not well removed further in the production of Compound 1. The structures of exemplary impurities are shown in Table 1.

[0107] Table 1: Examples of impurity reduction (Bz = benzoyl)

[0108]

[0109]

[0110] The reduction can optionally be carried out in the presence of cerium(III) chloride. In one embodiment, the amount of cerium chloride is about 50 mol% (e.g., 50 mol%). In another embodiment, the amount of cerium chloride is about 20 mol% or about 10 mol% (e.g., 20 mol% or 10 mol%). After adding cerium(III) chloride, the reaction can be heated (e.g., to about 15 °C to about 25 °C) for example about 20 minutes before cooling.

[0111] After the reduction, the reaction can be quenched, for example, with acetone and the solution neutralized with an acid such as citric acid. The organic layer containing the compound of Formula IIIa can be separated and washed, for example, with water. The organic layer can be heated (e.g., to about 20 °C to about 30 °C) and the pH adjusted with an acid (e.g., citric acid), repeating the steps until the pH is stable. The organic phase can be washed (e.g., with sodium bicarbonate) at a temperature of from about 20 °C to about 35 °C, and then washed with water at a temperature of from about 0 °C to about 10 °C. The volume of the organic phase can be reduced (e.g., by distillation), methyl tert-butyl ether can be added, and the resulting precipitate collected, washed with methyl tert-butyl ether cooled to about 0 °C to about 10 °C, and dried.

[0112] The deprotection of the compound of formula IIIA to produce compound 2 in step (c) can be carried out by methods known in the art, such as those disclosed in U.S. Patent No. 8,268,800 and / or Patent Publication WO2015 / 066162. For example, the deprotection can be carried out in the presence of a weak base such as ammonium hydroxide in a solvent such as methanol. In some embodiments, the deprotection can be carried out in the presence of an organic base, such as one or more bases selected from the group consisting of DBU, trimethylamine, N,N-dimethyl-4-aminopyridine (DMAP), and 1,3-diazabicyclo[2.2.2]octane (DABCO). In some embodiments, the deprotection can be carried out in the presence of ammonia in a solvent such as methanol, for example, at a temperature of about 20 °C to about 30 °C. The amount of the organic base used for deprotection should not be limited as long as it is the necessary amount and does not cause any adverse effects, for example, does not cause side reactions. In some embodiments, the amount of the organic base used for deprotection can be about 0.01 to about 2.2 moles per mole of the compound of formula IIIa (e.g., about 0.01, 0.02, 0.03, 0.04, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.15, 2.16, 2.17, 2.18, 2.19, or 2.2 moles per mole of the compound of formula IIIa, or any value or range therein). For example, in some embodiments, the deprotection step (c) can be carried out in the presence of about 0.01 to about 2.2 moles of the organic base per mole of the compound of formula IIIa, or about 0.05 to about 2.0 moles of the organic base per mole of the compound of formula IIIa, or about 1.0 to about 1.9 moles of the organic base per mole of the compound of formula IIIa. The deprotection can be carried out for about 12 - 48 hours, for example, about 24 hours. After deprotection, the mixture can be concentrated, dissolved in an aqueous solvent such as water, and washed with an organic solvent such as ethyl acetate.

[0113] In some embodiments, after deprotection, the work-up can be carried out under non-aqueous conditions, for example, using an evaporation solvent exchange process. The inventors of the present invention have found that using a non-aqueous evaporation solvent exchange process forms a non-tacky solid and avoids hydrolysis instability and dimer formation. In some embodiments, the deprotection can be further followed by additional milling to remove impurities, such as genotoxic impurities, such as but not limited to benzamide. In some embodiments, the deprotection in the non-aqueous work-up can be carried out with isopropanol and / or acetonitrile.

[0114] In some embodiments, the non-aqueous workup can be carried out by reducing the volume of the reaction mixture (e.g., by distillation under reduced pressure), adding methanol and continuing to reduce the volume, adding isopropanol and acetonitrile and continuing to reduce the volume, and then adding acetonitrile and continuing to reduce the volume. The reaction can then be cooled, e.g., to about 0 °C to about 10 °C, e.g., over 1 - 2 hours, and held, e.g., for 1 - 6 hours. The solid product can be collected and washed with acetonitrile.

[0115] The precipitation or crystallization of compound 2 to produce compound 1 in step (d) can be carried out as described above.

[0116] In some embodiments, the precipitation or crystallization of compound 2 to produce compound 1 in step (d) can be carried out by suspending the compound, optionally under cooling, in the presence of DBU, acetic acid, trifluoroacetic acid, diisopropylethylamine, and / or ammonium hydroxide.

[0117] In some embodiments, the method of the invention can include the steps of (a) to (d), wherein the reduction step (b) is carried out at a temperature of about - 12 °C to about - 3 °C, thereby minimizing the formation of DCU and DPU and indirectly CYU. CYU cannot be well removed from the rest of the synthesis. In some embodiments, the method of the invention can further include the workup of the deprotected compound from the deprotection step (c) under non-aqueous conditions (e.g., evaporation solvent exchange), thereby producing a non-tacky solid and / or avoiding hydrolysis instability and / or dimer formation. In some embodiments, the method of the invention can further include, wherein the deprotection step (c) further includes a grinding step to remove genotoxic impurities such as benzamide. In some embodiments, the method of the invention can further include the recrystallization of the final product, wherein the recrystallization is carried out at a pH of about 6.0 to about 7.4 and a temperature of about 50 °C to about 55 °C.

[0118] In some embodiments, the method of the present invention may include steps (a) to (d), wherein the hydrogenation step (a) is carried out using a palladium catalyst in a hydrogen atmosphere, and / or wherein the deprotection step (c) is carried out in the presence of an organic base. In particular, with respect to step (a), WO 2015 / 066162, as an improved method disclosed in U.S. Patent No. 8,268,800, successfully carried out high-yield hydrogenation by protecting the hydroxyl group and then performing the hydrogenation step with a large amount of formic acid in the presence of a considerable amount of Pd / C. However, from the perspective of industrial production, the improved steps of WO 2015 / 066162 still have other drawbacks, such as the large use of expensive Pd / C and the large use of formic acid, which is not suitable as a reagent for industrial production. The present invention enables the hydrogenation step to be carried out with high yield and high purity by performing the hydrogenation step (a) in a hydrogen atmosphere in the presence of a catalytic amount of a palladium catalyst. In addition, the deprotection reaction in WO 2015 / 066162 requires a long time with ammonia, which is troublesome for industrial-scale production and generates impurities that are difficult to remove. Compared with U.S. Patent No. 8,268,800, carrying out the hydrogenation step in the presence of a large amount of expensive rhodium catalyst under unprotected hydroxyl groups results in, for example, high manufacturing costs, low product purity, and difficulty in purification by chromatography. The problems of the above WO 2015 / 066162 and U.S. Patent No. 8,268,800 are overcome by the invention disclosed herein.

[0119] Use

[0120] The compound 1 produced by the present invention or a pharmaceutically acceptable salt thereof can be used to inhibit CDA activity. The compound 1 or a pharmaceutically acceptable salt thereof can be in the form of a pharmaceutical composition, for example, together with a pharmaceutically acceptable excipient. In some embodiments, the compound 1 or a pharmaceutically acceptable salt thereof can be used in combination with a CDA substrate drug, such as a CDA substrate drug useful for treating cancer, in a method for treating cancer in a subject in need thereof. Examples of CDA substrate drugs include, but are not limited to, decitabine, 5-azacytidine, gemcitabine, ara-C, tiazofurin, 5-fluoro-2'-deoxycytidine, and cytochlor. In some embodiments, the cancer can be selected from the group consisting of blood cancers and solid cancers. In certain embodiments, the blood cancer can be myelodysplastic syndrome or leukemia, such as acute myeloid leukemia or chronic myeloid leukemia. In certain embodiments, the solid cancer can be pancreatic cancer, ovarian cancer, peritoneal cancer, non-small cell lung cancer, metastatic breast cancer, bladder cancer, squamous cell carcinoma, transitional cell carcinoma, adenocarcinoma, gynecological cancer, fallopian tube cancer, liver cancer, hepatocellular carcinoma, lung cancer, cervical cancer, urogenital cancer, or gastrointestinal cancer. In some embodiments, the compound 1 or a pharmaceutically acceptable salt thereof can be administered substantially simultaneously with, before, or after the CDA substrate drug, optionally in a single unit dosage form or in multiple separate unit dosage forms. Embodiments according to the present invention are described in the following non-limiting examples.

[0121] Example

[0122] Example 1. Hydrogenation step (a)

[0123] In step (a), the hydrogenation of the compound of formula IV (1 equivalent) of the compound of production IIa is carried out in ethyl acetate (8.97 g / g) with hydrogen at a hydrogen pressure of 2 to 4 bar and acetic acid (80%, 2.62 g / g) in water (1.98 g / g) in the presence of Pd / C (5%, 0.05 g / g) at a temperature of 63 °C to 77 °C for at least 12 hours. After completion, the catalyst is removed by filtration, and the filter cake is washed with ethyl acetate (6.73 g / g). After cooling, the filtrate is washed with an aqueous potassium carbonate solution. The organic phase is washed with an aqueous sodium bicarbonate solution (7% w / w, 5.38 g / g) and with an aqueous NaCl solution (10% w / w, 3 g / g). The organic layer is distilled to a residual volume of about 8 volumes, and the residue is heated to 70 °C until dissolved and cooled to 45 °C to 55 °C. The reaction is seeded with the compound of formula IIa, and the reaction mixture is distilled to a residual volume of about 2 volumes, and methyl tert-butyl ether (4.44 g / g) is slowly added at a temperature of 40 °C to 50 °C. After cooling the mixture, the precipitated crystals are collected on a filter, washed with methyl tert-butyl ether (1.48 g / g) cooled to 0 °C to 10 °C, and the product is dried to obtain formula IIa. Yield: 86%. Chemical purity: 98.9%.

[0124] Example 2. Reduction of impurities during step (b)

[0125] Dissolve the compound of formula IIa (1 equivalent) in dichloromethane (14.6 g / g) and stir. Add ethanol (5.83 g / g), and cool the reaction mixture to -5 °C to 5 °C. Add cerium(III) chloride heptahydrate (0.08 g / g), and heat the reaction to 15 °C to 25 °C. Then add water (0.68 g / g), and stir the reaction for at least 20 min, then cool to 0 °C to -11 °C. Maintain the temperature at 0 °C to -11 °C, add sodium borohydride (0.11 g / g) portionwise, and stir the reaction until completion. Maintain the temperature at 0 °C to -11 °C, add acetone (0.73 g / g). Slowly add an aqueous solution of citric acid (3.7 to 4.4% w / w) until a pH of 6.5 to 7.5 is reached. Allow the phases to settle and separate the aqueous phase. Wash the organic phase with an aqueous solution of sodium bicarbonate, and then further wash with water. Separate the phases and wash the organic phase further with water. Vacuum distill the organic phase at ≤ 35 °C until the volume is about 2 volumes, then add methyl tert-butyl ether (7.38 g / g), and vacuum distill at ≤ 35 °C until the volume is 4.8 volumes. Add a second portion of methyl tert-butyl ether (3.7 g / g) and vacuum distill at ≤ 30 °C until the volume is 4.8 volumes. Add additional methyl tert-butyl ether (2.67 g / g), and cool the reaction mixture to 0 °C to 10 °C over ≥ 4 h. Add additional methyl tert-butyl ether (0.89 g / g), and stir the reaction mixture at this temperature, and then separate the solid product and wash it with methyl tert-butyl ether (1.46 g / g) pre-cooled to 0 °C to 10 °C, and dry the product to obtain formula IIIa. Yield: 76%. Chemical purity: 93%.

[0126] The procedures of Entries 1 - 5 were carried out in a similar manner as above, except that the reduction temperature was changed to 0 °C, -3 °C, -5 °C, -8 °C, and -11 °C, respectively. As shown in Table 2, the lower temperature minimized the formation of DCU, DPU, and CYU. The structures of the impurities are shown in Table 1.

[0127] Table 2

[0128]

[0129] Example 3. Improvement during deprotection step (c)

[0130] Deprotection was carried out in a solution of ammonia (0.87 g / g) in methanol (4.95 g / g). The compound of formula IIIa (1 equivalent) and additional methanol (1.58 kg / kg) were added. The reaction mixture was adjusted to 20 °C to 30 °C and stirred at this temperature until completion. Work-up was carried out by distilling the reaction mixture under reduced pressure until the residual volume was about 3 volumes. Methanol (1.58 g / g) was added and distillation was continued under reduced pressure until the residual volume was about 3 volumes. Isopropanol (1.92 g / g) was added, then acetonitrile (3.93 g / g), and distillation was continued under reduced pressure until the residual volume was about 3 volumes. Acetonitrile (5.51 g / g) was added and distilled under reduced pressure until a residual volume of 5 volumes was reached. Additional acetonitrile (1.58 g / g) was added and distilled under reduced pressure until a residual volume of about 5 volumes was reached. After cooling the mixture, the solid product was separated and washed with acetonitrile (1.56 g / g) pre-cooled to 0 °C to 10 °C. The wet product was mixed with acetonitrile (2.37 g / g). After cooling the mixture, the wet product was then dried to give Compound 2. Yield: 87%. Chemical purity: 97%. Reference Example 4. Original deprotection step (c)

[0131] Deprotection was carried out by treatment with ammonia (7.0 M, in methanol, 25 equivalents). The mixture was stirred at 25 °C for 27 h and then concentrated under reduced pressure. The residue was dissolved in water (6.3 volumes) and washed twice with ethyl acetate (5.7 volumes each). The aqueous layer was concentrated under reduced pressure at a temperature below 35 °C to give Compound 2 (95% yield).

[0132] When proceeding with typical process conditions, Compound 2 generated from the original deprotection conditions generally provided Compound 1 with generally lower overall purity and a significantly higher number of impurities. See Table 3, Entries 1 and 2.

[0133] Example 5. Highly specific crystallization conditions at elevated temperature

[0134] The crystallization conditions of Compound 1 for improved purity were tested. It was found that highly specific crystallization conditions produced very pure material, improved the control of the particle size distribution into the optimal (monomodal) range, had a lower acetonitrile level, and avoided dimeric impurities. The specific conditions included using a higher temperature and adjusting to pH.

[0135] The preferred procedure for the recrystallization of Compound 1 is carried out by mixing Compound 2 (1 equivalent), 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU, 0.028 g / g), acetonitrile (5.02 g / g), and water (0.73 g / g) and stirring for 2 h at 15 °C to 25 °C. The mixture is cooled to 0 °C to 10 °C within ≥1 h and held at this temperature for 1 to 8 h. The solid product is collected by filtration and washed with acetonitrile:water (0.21 g / g:0.033 g / g) cooled to 0 °C to 10 °C. The wet product and acetonitrile (2.36 g / g) are stirred at 0 °C to 10 °C for 30 min to 8 h. The solid product is collected and washed with acetonitrile (0.78 g / g) cooled to 0 °C to 10 °C. The wet product is dried to obtain crude Compound 1. Yield: 80%. Chemical purity: 94%.

[0136] Mix crude Compound 1 (1 equivalent), acetone (6.26 g / g), and water (2 g / g) at a temperature of 20 °C to 30 °C. The pH of the reaction mixture is adjusted to 6.0 to 7.5 with an aqueous solution of formic acid (0.0076 g / g formic acid in 1.25 g / g water). If necessary, an aqueous solution of 1,8-diazabicyclo[5.4.0]undec-7-ene DBU (0.041 g / g DBU in 0.78 g / g water) can be used to adjust the pH to the target range. The reaction mixture is heated to 50 °C to 55 °C until complete dissolution is observed and filtered into a reactor set at 40 °C to 50 °C and washed with a mixture of water (0.1 g / g) and acetone (0.31 g / g). The solution is cooled to 33 °C to 43 °C and seeded with 0.005 g / g of Compound 1. The reaction is maintained at this temperature and then cooled to -10 °C to 0 °C. The suspension is held at -10 °C to 0 °C for 12 to 16 h and separated by filtration. The product is washed with acetone (0.78 g / g) and dried to obtain Compound 1. Yield: 61%. Chemical purity excluding epimers: 99.6%. The compound 6 of the following formula, which is an epimer of Compound 1, is produced at the following ratio. Compound 1:Compound 6 = 99.6:0.4. (PSD: D(0.9) = 252 μm, D(0.5) = 125 μm, D(0.1) = 21 μm).

[0137] Example 6. Comparison with the original purification process

[0138] The crude compound 1 (1 equivalent) was suspended in a mixture of acetone (2.5 ml / g) and water (2.5 ml / g) and stirred at 25 ± 2 °C for 2 hours. The mixture was cooled to 5 ± 2 °C and stirred for 2 hours, and then filtered. The filter cake was rinsed with acetone (2 x 0.55 ml / g), and then dried in vacuo at 55 °C. Compound 1 was provided in 67% yield. Chemical purity excluding epimers: 99.4%. The compound 6 of the following formula, which is an epimer of compound 1, was produced in the following ratio. Compound 1:Compound 6 = 98.7:1.3.

[0139] Compared with the updated purification process, the overall purity of compound 1 and the ratio to compound 6 were lower in the original process. The impurity distribution is shown in Table 3. The structures of the monitored impurities are shown in Table 4.

[0140] Table 3

[0141]

[0142] ND = not detected.

[0143] Table 4

[0144]

[0145] Example 7. Alternative methods for steps (a) and (c)

[0146] Step (a):

[0147]

[0148] A mixture of the compound (2-1) (20.0 g, 42.4 mmol), ethyl acetate (200 mL), acetic acid (80 mL), and water (50 mL) was dissolved with stirring at 50 to 60 °C under a nitrogen atmosphere, and then 0.2 g of Pd / C (10% by weight, 5 wt% on a dry basis, 50% water-wet, type NEs-5DR) was added thereto and the atmosphere in the reaction vessel was replaced with nitrogen. Subsequently, the atmosphere was replaced with hydrogen, and the reaction mixture was stirred at 50 to 60 °C under an increased hydrogen pressure (0.5 MPa) for 18 hours. The reaction mixture was filtered and the residue on the filter was washed with 20 mL of 80% acetic acid. The ethyl acetate in the filtrate was removed in vacuo (100 Torr, 50 °C). The residual solution was heated to 80 °C to dissolve it, and 400 mL of water preheated to 80 °C was added thereto. After cooling the mixture, the precipitated crystals were collected on a filter, washed with water and ethanol, and dried to obtain 19.24 g of the compound (3-1) (yield: 95.6%, chemical purity: 98.4%).

[0149] Steps (b) and (c)

[0150]

[0151] Synthesis of compound (4-1): Compound (3-1) (19.0 g, 40.0 mmol) was dissolved in dichloromethane (228 mL) under stirring. Ethanol (152 mL) and cerium(III) chloride heptahydrate (1.49 g, 4.0 mmol) were added to the solution, and the mixture was cooled to a temperature below 6 °C. A solution of sodium borohydride (3.77 g, 100 mmol) in water (19 mL) was added dropwise to the cooled mixture while maintaining its temperature below 6 °C. After the addition, the reaction mixture was reacted at a temperature below 6 °C for 2 hours. While maintaining the temperature of the reaction mixture below 6 °C, the reaction mixture under stirring was quenched with 9.5 mL of acetone, and then 114 mL of 0.5 M aqueous hydrochloric acid was added thereto while maintaining its temperature below 6 °C to adjust the quenched mixture to pH 7. After the mixture was warmed to 30 to 40 °C, 114 mL of saturated aqueous sodium bicarbonate was added to the mixture. The mixture was separated with a separatory funnel, the organic layer was washed with 114 mL of water, and the solvent was removed in vacuo (50 Torr, 40 °C). The resulting compound (4-1) was used in the next step without purification.

[0152] Synthesis of compound 5: Methanol (190 mL) and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) (0.30 mL, 2.0 mmol) were added to the compound (4-1) obtained in step (b), and the reaction mixture was warmed to 40 °C and stirred at 40 °C for 3 hours. After the reaction, methanol was removed in vacuo (50 Torr, 40 °C), and 95 mL of acetonitrile was added to the residual solution. After cooling, the precipitated crystals were collected on a filter, washed with cooled acetonitrile, and dried to obtain 8.96 g of compound 5 (yield: 83.4%, chemical purity: 97.1%).

[0153] Step (d)

[0154]

[0155] Synthesis of crude compound (1):A mixture of compound (5) (0.9 g, 3.35 mmol), acetonitrile (5.8 mL), water (0.65 mL) and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) (0.025 mL, 0.17 mmol) was suspended, and the suspension was continuously stirred at 20 to 30 °C for 48 hours. After cooling, the precipitated crystals were collected on a filter, washed with cooled 90% acetonitrile and acetonitrile, and dried to obtain 0.66 g of compound 1 (yield: 73.0%, chemical purity: 98.3%). Compound 5: Compound 6 (epimer of compound 5) = 98.9:1.1.

[0156] Purification of crude compound 1: Compound 1 (0.3 g, 1.11 mmol) was suspended in a mixture of acetonitrile (0.96 mL) and water (0.24 mL), and the suspension was stirred at 40 to 50 °C for two hours. Then, the suspension was stirred at a temperature below 5 °C for two hours. The resulting precipitate was collected on a filter, washed with cooled 90% acetonitrile and acetonitrile, and dried to obtain 0.66 g of compound 1 (yield: 73.5%, chemical purity: 99.6%). The following compound 6, which is an epimer of compound 1, was produced at the following ratio. Compound 1: Compound 6 = 99.7:0.3.

[0157]

[0158] The total amount of impurities contained in the purified compound 1 was 0.46% (by area percentage). Separately, compound 1 was prepared and purified by the method disclosed in Patent Document 2 (WO 2015 / 066162) from the same material, but the total amount of impurities therein (2.46%) was higher than the total amount of impurities in the above method, which means that the method of the present invention can provide a method for preparing compound 1 in a higher yield.

[0160] Embodiments of the present invention include, but are not limited to, the following.

[0161] 1. A method for producing compound 1 or a salt thereof:

[0162]

[0163] Comprising the following steps:

[0164] (a) Hydrogenating a compound of formula IV:

[0165]

[0166] wherein R is a hydroxyl protecting group,

[0167] to produce a compound of formula IIa:

[0168]

[0169] (b) Reduce the compound of formula IIa to produce a compound of formula IIIa:

[0170]

[0171] (c) Deprotect the compound of formula IIIa to produce compound 2:

[0172] and

[0173] (d) Precipitate or crystallize compound 2 in the presence of a catalyst to produce compound 1:

[0174]

[0175] or a salt thereof;

[0176] wherein the method comprises one or more of the following, in any combination:

[0177] (i) wherein the hydrogenation step (a) is carried out using a palladium catalyst in a hydrogen atmosphere;

[0178] (ii) wherein the reduction step (b) is carried out at a temperature of about -12 °C to about -5 °C;

[0179] (iii) wherein the deprotection step (c) is carried out in the presence of an organic base; and / or

[0180] (iv) wherein the deprotection step (c) is carried out under non-aqueous conditions.

[0181] 2. The method according to embodiment 1, wherein the catalyst is about 1 mol% to about 20 mol% of DBU.

[0182] 3. The method according to embodiment 2, wherein the catalyst is about 5 mol% to about 10 mol% of DBU.

[0183] 4. The method according to embodiment 3, wherein the catalyst is about 5 mol% of DBU.

[0184] 5. The method according to embodiment 1, wherein the catalyst is acetic acid, trifluoroacetic acid, diisopropylethylamine or ammonium hydroxide.

[0185] 6. The method according to embodiments 1-5, wherein the deprotection step (c) further comprises a grinding step.

[0186] 7. The method according to any one of Embodiments 1-6 further comprises recrystallizing or slurrying Compound 1.

[0187] 8. The method according to Embodiment 7, wherein the recrystallization is carried out at a pH of about 6.0 to about 7.4 and a temperature of about 50 °C to about 55 °C.

[0188] 9. The method according to any one of Embodiments 1-8, wherein step (d) is carried out in the presence of a solution comprising acetonitrile.

[0189] 10. The method according to any one of Embodiments 1-8, wherein step (d) is carried out in the presence of a solution comprising acetone or tetrahydrofuran.

[0190] 11. The method according to any one of Embodiments 1-10, wherein step (b) is carried out in the presence of CeCl 3 present.

[0191] 12. The method according to any one of Embodiments 1-11, wherein the R is a benzoyl group.

[0192] 13. The method according to any one of Embodiments 1-12, wherein the palladium catalyst used in step (i) is palladium on carbon (Pd / C).

[0193] 14. The method according to Embodiment 13, wherein the amount of the Pd / C used in step (i) is a catalytic amount not exceeding 0.1 part by weight per 1 part by weight of the compound of Formula IV.

[0194] 15. The method according to Embodiment 13, wherein the amount of the Pd / C used in step (i) is 0.025 to 0.05 part by weight per 1 part by weight of the compound of Formula IV.

[0195] 16. The method according to any one of Embodiments 1-15, wherein step (i) is carried out in a mixed solvent comprising ethyl acetate, acetic acid and water.

[0196] 17. The method according to any one of Embodiments 1-16, wherein step (i) is carried out at ambient pressure or increased pressure.

[0197] 18. The method according to any one of Embodiments 1-17, wherein step (i) is carried out at an increased pressure of 0.1 to 0.5 MPa.

[0198] 19. The method according to any one of Embodiments 1-18, wherein the organic base used in step (iii) is one or more bases selected from the group consisting of DBU, triethylamine, DMAP and DABCO.

[0199] 20. The method according to any one of Embodiments 1-19, wherein the amount of the organic base used in step (iii) is 0.01 to 2.2 moles per mole of the compound of Formula IIIa.

[0200] 21. The method according to any one of Embodiments 1-20, wherein the organic base used in step (iii) is DBU.

[0201] 22. A composition of Compound 1 or a salt thereof:

[0202]

[0203] comprising less than 10, 9, 8, 7, 6 or 5 measurable impurities.

[0204] 23. The composition according to Claim 24, wherein there are no impurities at a level greater than 0.5 wt%, 0.25 wt%, 0.2 wt%, 0.15 wt%, 0.1 wt%, 0.05 wt% or 0.01 wt%.

[0205] 24. A composition of Compound 1 or a salt thereof:

[0206]

[0207] wherein there are no impurities at a level greater than 0.5 wt%, 0.25 wt%, 0.2 wt%, 0.15 wt%, 0.1 wt%, 0.05 wt% or 0.01 wt%.

[0208] 25. The composition according to Claim 26, comprising less than 10, 9, 8, 7, 6 or 5 measurable impurities.

[0209] The foregoing is a description of the invention and is not to be construed as a limitation thereof. The invention is defined by the following claims, including equivalents of the claims.

[0210] All publications, patent applications, patents and other references cited herein are incorporated by reference in their entirety for the teachings relevant to the sentences and / or paragraphs in which the references are presented.

Claims

1. A composition comprising Compound 1 or a salt thereof: wherein the composition comprises fewer than 7 measurable impurities, wherein there is no impurity with a level greater than 1.0 wt% of the composition, wherein Compound 1 has a purity of at least 99% by weight of the composition, and wherein the composition comprises an impurity at a level of at least 0.25 wt% of the composition.

2. The composition according to claim 1, wherein the composition comprises fewer than 6 measurable impurities.

3. The composition according to claim 1 or 2, wherein the composition comprises fewer than 5 impurities.

4. The composition according to any one of claims 1 - 3, wherein there is no impurity with a level greater than 0.5 wt%.

5. The composition according to any one of claims 1 - 4, wherein there is no impurity with a level greater than 0.25 wt%.

6. The composition according to any one of claims 1 - 5, wherein Compound 1 has a purity of at least 99.5% by weight of the composition.

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