EA043635B1 - DISubstituted pyrazole compounds as ketohexokinase inhibitors - Google Patents

Description

This invention relates to novel ketohexokinase (KHC) inhibitor compounds, pharmaceutical compositions containing these compounds, and the use of the compounds for the treatment of certain conditions such as type 2 diabetes mellitus (T2DM), heart failure, diabetic kidney disease and non-alcoholic steatohepatitis (NASH). ).

KGC, also called fructokinase, is a rate-limiting enzyme involved in the metabolism of fructose. It catalyzes the phosphorylation of fructose to fructose 1phosphate (F1P), causing a concomitant depletion of cellular ATP levels. Unlike glucose, fructose metabolism lacks feedback inhibition and triggers the accumulation of downstream intermediates involved, for example, in lipogenesis, gluconeogenesis and oxidative phosphorylation (Hannou, S.A. et al., J. Clin. Invest., 128 (2), 544-555, 2018). This has negative metabolic consequences that are associated with a number of serious metabolic disorders.

KGC exists in two alternatively spliced isoforms, consisting of KGC-C and KGC-A, distinguished by exon 3. KGC-C is expressed primarily in the liver, kidney, and intestine, whereas KGC-A is ubiquitous. Mice deficient in both isoforms are completely protected from fructose-induced metabolic syndrome. However, the adverse metabolic effects are exacerbated in mice lacking BGA-A alone (Ishimoto T. et al., Proc. Natl. Acad. Sci., USA, 109(11), 4320-4325, 2012).

Several epidemiological and experimental studies have reported that increased fructose intake, or more specifically increased fructose metabolism, may play an important role in the development of several disorders, including metabolic syndrome and, in particular, in the development of type 2 diabetes (Softic et al., J. Clin . Invest., 127(11), 4059-4074, 2017), heart failure (Mirtschink, P. et al., Eur. Heart J., 39, 2497-2505, 2018), diabetic kidney disease (Cirillo, P. et al., J. Am. Soc. Nephrol, 20, 545-553, 2009) and NAFLD/NASH (Vos, M. B. et al., Hepatology, 57, 2525-2531, 2013). Targeted inhibition of BGCs is expected to limit fructose metabolism and provide effective treatment options for a range of metabolic disorders.

US 2017/0183328 A1 describes substituted 3-azabicyclo[3.1.0]hexanes as KGC inhibitors. Recently published data show that the ketohexokinase inhibitor PF-06835919, administered for 6 weeks, reduces overall liver fat content as measured by proton density of the fat fraction on magnetic resonance imaging in subjects with nonalcoholic fatty liver disease (J. Hepatology , EASL International Liver Congress Abstracts, Supplement No. 1S, vol. 70, April 2019).

Compounds containing carboxyl functional groups carry a risk associated with the formation of acyl glucuronide metabolites (Vleet Van et al., Toxicology Letters, 272 (2017) 1-7). Acyl glucuronide metabolites are often unstable and can be chemically reactive, resulting in covalent binding to macromolecules and toxicity.

There is a need for alternative treatments for metabolic syndrome and its associated indications, including type 2 diabetes, heart failure, diabetic kidney disease and NASH. In particular, there is a need for compounds that are potent CHC inhibitors. There is a need for CHC inhibitor compounds that have beneficial properties, such as good oral bioavailability, to provide once-daily dosing. In addition, there is a need for KGC inhibitor compounds that do not have a carboxylic acid moiety and do not have the ability to form acyl glucuronides.

Accordingly, in one embodiment of the present invention there is provided a compound of formula I

N X^N

Formula I wherein X is N or C substituted with CN; R ¹ selected from H,

- 1 043635

R ² and R ³ are both H or one is H and the other is OH;

R ⁴ , R ⁵ , R ⁶ , R ⁷ and R ⁹ are independently H or CH ₃ ;

R ⁸ is H, CH ₃ , CH ₂ CH ₂ OH, C(=O)CH ₂ NH ₂ or C(=O)CH ₃ ; And

R ¹⁰ represents OH or NH ₂ ; or a pharmaceutically acceptable salt thereof.

In a specific embodiment of the invention, R ¹ is selected from H,

or a pharmaceutically acceptable salt thereof.

In a specific embodiment, the invention provides a compound of formula I, wherein X is N or C substituted with CN;

R ¹ selected from

and where R ² represents H and R ³ represents OH, or

R ² represents OH and R ³ represents H, or both R ² and R ³ represent H;

or a pharmaceutically acceptable salt thereof.

In a specific embodiment of the invention, the compound has formula Ia

- 2 043635 or is a pharmaceutically acceptable salt thereof.

In a specific embodiment of the invention, the compound has the formula Ic

or is a pharmaceutically acceptable salt thereof.

In a specific embodiment of the invention, the compound has the formula Ie

Formula 1e;

or is a pharmaceutically acceptable salt thereof.

In one embodiment of the invention, X is N.

In one embodiment of the invention, X is C substituted with CN.

In one embodiment of the invention, R ¹ is

In one embodiment of the invention, a compound of Formula I is

N

or a pharmaceutically acceptable salt thereof.

A specific embodiment of the invention provides a succinate salt

In a preferred embodiment of the invention, the succinate salt is a sesquisuccinate salt.

In one embodiment of the invention, a compound of Formula I is

- 3 043635 or a pharmaceutically acceptable salt thereof.

In one embodiment of the invention, a compound of Formula I is

or a pharmaceutically acceptable salt thereof.

In one embodiment of the invention, a compound of Formula I is

or a pharmaceutically acceptable salt thereof.

In one embodiment of the invention, a compound of Formula I is

or a pharmaceutically acceptable salt thereof.

In one embodiment of the invention, a compound of Formula I is

or a pharmaceutically acceptable salt thereof.

In one embodiment of the invention, a compound of Formula I is

or a pharmaceutically acceptable salt thereof.

- 4 043635

In a specific embodiment of the invention, the compound is selected from

2-[4-[2-[(25)-2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1 -yl]-1 piperazin-1-ylethanone;

[(27?)-1 - [4-[1-(4-piperidyl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]azetidin-2yl]methanol;

[(27?)-1 - [4-[1 -(azetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]azetidin-2yl]methanol;

(2S,3R)-1-[4-[1-(azetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]-2methylazetidin-3-ol;

2-[4-[5-cyano-6-[(2L')-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl]-A'(2-hydroxyethyl )acetamide;

2-[(27?)-2-(hydroxymethyl)azetidin-1-yl]-6-[1-(4-piperidyl)pyrazol-4-yl]-4(trifluoromethyl)pyridin-3-carbonitrile;

2-[(25.37?)-3-hydroxy-2-methylazetidin-1-yl]-6-[1-(4-piperidyl)pyrazol-4-yl]-4(trifluoromethyl)pyridin-3-carbonitrile ;

2-[(25)-2-methylazetidin-1-yl]-6-[1-(1-methyl-4-piperidyl)pyrazol-4-yl]-4(trifluoromethyl)pyridin-3-carbonitrile;

6-[1-[1-(2-hydroxyethyl)-4-piperidyl]pyrazol-4-yl]-2-[(28)-2-methylazetidin-1-yl]-4(trifluoromethyl)pyridin-3-carbonitrile ;

6-[1-[2-(dimethylamino)ethyl]pyrazol-4-yl]-2-[(25)-2-methylazetidin-1-yl]-4(trifluoromethyl)pyridin-3-carbonitrile;

6-[1-(2-hydroxyethyl)pyrazol-4-yl]-2-[(26)-2-methylazetidi n-1-yl]-4(trifluoromethyl)pyridin-3-carbonitrile;

2-[(26)-2-methylazetidin-1-yl]-6-[1-(4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridin-3 carbonitrile;

2-[(26)-2-methylazetidin-1-yl]-4-[1-(4-piperidyl)pyrazol-4-yl]-6(trifluoromethyl)pyrimidine;

(25.37?)-2-methyl-1 - [4- [ 1 - [(37?)-pyrrolidin-3 -yl]pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2yl]azetidin-3 -ola;

(25.37?)-2-methyl-1-[4-[1-[(35)-pyrrolidin-3-yl]pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2yl]azetidin-3- ola;

[(27?)-1 - [4-[1 - [(37?)-pyrrolidin-3-yl]pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2yl]azetidin-2-yl]methanol;

6-[1-(azetidin-3-yl)pyrazol-4-yl]-2-[(26)-2-methylazetidin-1-yl]-4-(trifluoromethyl)pyridin3-carbonitrile;

6-[1-[1-(2-Aminoacetyl)-4-piperidyl]pyrazol-4-yl]-2-[(26)-2-methylazetidin-1-yl]-4(trifluoromethyl)pyridin-3-carbonitrile ;

6-[1-(1-acetyl-4-piperidyl)pyrazol-4-yl]-2-[(25)-2-methylazetidin-1-yl]-4(trifluoromethyl)pyridin-3-carbonitrile;

2-[(26)-2-methylazetidin-1-yl]-6-[1-[2-(4-methylpiperazin-1-yl)-2-oxoethyl]pyrazol-4-yl]-4(trifluoromethyl)pyridine -3-carbonitrile;

2-[4-[5-cyano-6-[(25)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl] -N,N bis(2 -hydroxyethyl)acetamide;

- 5 043635

6-[1-[2-[(3R,4R)-3,4-dihydroxypyrrolidin-1-yl]-2-oxoethyl]pyrazol-4-yl]-2-[(2S)-2methylazetidin-1-yl ]-4-(trifluoromethyl)pyridine-3-carbonitrile;

6-[1-[2-[(3^,40)-3,4-dihydroxypyrrolidin-1-yl]-2-oxoethyl]pyrazol-4-yl]-2-[(20)-2-methylazetidin-1 -yl]-4-(trifluoromethyl)pyridin-3-carbonitrile;

6-[1-(2,3-dihydroxypropyl)pyrazol-4-yl]-2-[(25)-2-methylazetidin-1-yl]-4-(trifluoromethyl)pyridin-3-carbonitrile;

2-[(25)-2-methylazetidin-1-yl]-6-(1H-pyrazol-4-yl)-4-(trifluoromethyl)pyridin-3carbonitrile;

2-[(20)-2-methylazetidin-1-yl]-6-(1-tetrahydropyran-4-ylpyrazol-4-yl)-4(trifluoromethyl)pyridin-3-carbonitrile;

6-[1-(3-hydroxy-4-piperidyl)pyrazol-4-yl]-2-[(2L)-2-methyl lasetid and n-1-yl]-4-(trifluoromethyl)pyridin-3- carbonitrile;

M-(2-aminoethyl)-2-[4-[2-[(20)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1-yl]acetamide;

2-[4-[2-[(20)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl]-1-[(30)-

-methylpiperazin-1-yl]ethanone;

2-[4-[2-[(20)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl]-1-[(20)2-methylpiperazin- 1 -yl]ethanone;

-(3,3 - dimethylpiperazin-1 -yl)-2-[4-[2-[(2L)-2-methylazetid and n-1 -yl]-6(trifluoromethyl)pyrimidin-4-yl]pyrazol- 1-yl]ethanone;

- [(2X, 5R)-2,5-dimethylpiperazin-1-yl]-2-[4-[2-[(2L)-2-methylazeti d in-1 -yl]-6(trifluoromethyl)pyrimidin-4 -yl]pyrazol-1-yl]ethanone;

4-[1-(azetidin-3-yl)pyrazol-4-yl]-2-[(20)-2-methylazetidin-1-yl]-6(trifluoromethyl)pyrimidine;

2-[(25)-2-methylazetidin-1-yl]-4-[1-(1-methylazetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidine;

(2X,37?)-2-methyl-1 - [4-[1-(1-methylazetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2yl]azetidin-3-ol; or

[(27?)-1 - [4-[1-(1-methylazetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2yl]azetidin-2-yl]methanol;

or a pharmaceutically acceptable salt thereof.

In a specific embodiment, the compound is 2-[4-[2-[(2S)-2methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1 -yl]-1-piperazin- 1-ylethanone sesquisuccinate, which is also known as 2-[4-[2-[(2S)-2-methyl-1-azetidinyl]-6-(trifluoromethyl)-4pyrimidinyl]-1H-nirazol-1-yl]-1 -(1-piperazinyl)ethanone, butanedioate (1:1.5) or butanedioic acid 2-(4-{2-[(28)-2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl } -1 H-pyrazol-1-yl)-1 -(piperazin1-yl)ethan-1-one (1.5/1).

Formula I includes formulas Ia, Ib, Ic, Id, Ie, If, Ig and Ih, and reference to Formula I below, for example in methods of treatment and therapeutic applications, should also be construed as reference to all of these sub-formulas without exception.

In one embodiment, the present invention provides a method of treating type 2 diabetes in a patient in need of such treatment, which comprises administering to the patient an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof.

In one embodiment, the present invention provides a method of treating heart failure in a patient in need of such treatment, which comprises administering to the patient an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof.

In one embodiment, the present invention provides a method of treating diabetic kidney disease in a patient in need of such treatment, which comprises administering to the patient an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof.

In one embodiment, the present invention provides a method of treating NASH in a patient in need of such treatment, which comprises administering to the patient an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof.

In one embodiment, the present invention provides a method of treating a disease selected from the group consisting of metabolic syndrome, NAFLD, obesity, cardiovascular disease, coronary artery disease, chronic kidney disease, dyslipidemia, and complications of diabetes, such as diabetic retinopathy, in a patient in need of such treatment comprising administering to the patient an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof.

Moreover, in one embodiment of the present invention there is provided a compound of formula I or a pharmaceutically acceptable salt thereof for use in therapy. In one embodiment, the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof for use in the treatment of type 2 diabetes. In one embodiment, the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof for use in the treatment of heart failure. In one embodiment, the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof for use in the treatment of diabetic kidney disease. In one embodiment, the present invention provides a compound of Formula I or a pharmaceutically acceptable salt thereof for use in the treatment of NASH. In one embodiment, the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof for use in the treatment of metabolic syndrome, NAFLD, obesity, cardiovascular disease, coronary artery disease, chronic kidney disease, dyslipidemia, and complications of diabetes, such as diabetic retinopathy.

Moreover, one embodiment of the present invention provides the use of a compound of formula I or a pharmaceutically acceptable salt thereof for the preparation of a medicament for the treatment of type 2 diabetes. In one embodiment of the present invention, the use of a compound of Formula I or a pharmaceutically acceptable salt thereof is provided for the preparation of a medicament for the treatment of heart failure. In one embodiment, the present invention provides the use of a compound of formula I or a pharmaceutically acceptable salt thereof for the preparation of a medicament for the treatment of diabetic kidney disease. In one embodiment, the present invention provides the use of a compound of formula I or a pharmaceutically acceptable salt thereof for the preparation of a medicament for the treatment of NASH. One embodiment of the present invention provides the use of a compound of formula I or a pharmaceutically acceptable salt thereof for the preparation of a medicament for the treatment of metabolic syndrome, NAFLD, obesity, cardiovascular disease, coronary artery disease, chronic kidney disease, dyslipidemia, and complications of diabetes, such as diabetic retinopathy.

In one embodiment, the present invention provides a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt thereof with one or more pharmaceutically acceptable carriers, diluents or excipients. In one embodiment, the present invention provides a method for preparing a pharmaceutical composition, comprising admixing a compound of Formula I or a pharmaceutically acceptable salt thereof with one or more pharmaceutically acceptable carriers, diluents or excipients.

As used herein, the term treat or treat includes controlling, slowing, stopping, or reversing the progression or severity of an existing symptom or disorder.

As used herein, the term patient refers to a mammal. The patient is preferably a human.

As used herein, the term effective amount refers to an amount or dose of a compound of Formula I or a pharmaceutically acceptable salt thereof that, when administered single or multiple times to a patient, provides the desired effect in the patient being diagnosed or treated.

The effective amount can be determined by one skilled in the art using known techniques and observing results obtained under similar circumstances. When determining an effective amount for a patient, many factors are considered, including, but not limited to, the type of patient; its size, age and general health; the specific disease or disorder involved; degree of involvement or severity of disease or disorder; individual patient response; the specific compound administered; method of administration; characteristics of the bioavailability of the administered drug; selected dosing regimen; use of concomitant medications; and other relevant circumstances. The compounds of formula I are effective at a daily dosage that ranges from about 0.1 mg/kg to about 15 mg/kg body weight.

The compounds of formula I are formulated into pharmaceutical compositions that are administered by any route that ensures the bioavailability of the compound. Preferably, said compositions are for oral administration. Such pharmaceutical compositions and methods for their preparation are well known in the art (see, for example, Remington, J. P., Remington: The Science and Practice

- 7 043635 of Pharmacy, L.V. Allen, editor, 22nd ed., Pharmaceutical Press, 2012).

The compounds of Formula I and their pharmaceutically acceptable salts can be used in the methods of treatment and therapeutic purposes of this invention, with certain configurations being preferred. It should be understood that the following preferences apply to both treatments and therapeutic applications and to the compounds of this invention.

Compounds of this invention include

Formula 1e;

- 8 043635

HE

F ₃

N

R

Formula If;

R

Formula Ig; or

R

Formula Ih;

and pharmaceutically acceptable salts thereof.

Although the present invention covers all individual enantiomers and diastereomers, as well as mixtures of these compounds, including racemates, the compounds of formula Ia, formula Ic and formula Ie and their pharmaceutically acceptable salts are particularly preferred.

The individual enantiomers can be isolated or separated by one of ordinary skill in the art at any convenient time during the synthesis of compounds of formula I, by such methods as selective crystallization techniques, chiral chromatography (see, for example, J. Jacques et al., Enantiomers, Racemates, and Resolutions, John Wiley and Sons, Inc., 1981, and E. L. Eliel and S. H. Wilen, Stereochemistry of Organic Compounds, Wiley-Interscience, 1994), or supercritical liquid chromatography (SLC) (see, for example, T. A. Berger, Supercritical Fluid Chromatography Primer, Agilent Technologies, July 2015).

A pharmaceutically acceptable salt of a compound of formula I can be prepared, for example, by reacting the corresponding free base of a compound of formula I with an appropriate pharmaceutically acceptable acid in a suitable solvent under standard conditions well known in the art (see, for example, Bastin, R. J. et al., Org. Process. Res. Dev., 4, 427-435, 2000; and Berge, S. M. et al., J. Pharm. Sci, 66, 1-19, 1977). The preferred salt is the succinate salt. A particularly preferred salt is the sesquisuccinate salt. In the sesquisuccinate salt, the ratio of free base to succinate is 1:1.5. Succinate salt is also known as butanedioate salt.

The compounds of formula I or salts thereof can be prepared by a variety of procedures known to one of ordinary skill in the art, some of which are presented below in the diagrams, preparations and examples. The products of each step in the schemes below can be isolated by conventional methods well known in the art, including extraction, evaporation, precipitation, chromatography, filtration, trituration and crystallization. In the following diagrams, all substituents, unless otherwise noted, are as previously defined. The reagents and starting materials are generally available to those of ordinary skill in the art. Without limiting the scope of the invention, the following diagrams, preparations and examples are presented to further illustrate the invention. Additionally, one of ordinary skill in the art will understand that the compounds of Formula I can be prepared using a starting material or intermediate with the appropriate stereochemical configuration required, which can be prepared by one of ordinary skill in the art.

Certain abbreviations are defined as follows.

ACN refers to acetonitrile;

- 9 043635

BOC refers to tert-butoxycarbonyl;

DCM refers to methylene chloride or dichloromethane;

DIPEA refers to N,N-diisopropylethylamine;

DMF refers to N,N-dimethylformamide;

DMSO refers to dimethyl sulfoxide;

ELSD refers to evaporative light scattering detector;

ER/MS refers to electrospray mass spectrometry;

EtOAc refers to ethyl acetate;

EtOH refers to ethanol or ethyl alcohol;

h refers to an hour or hours;

HATU refers to 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium-3-oxide hexafluorophosphate;

HPLC refers to high-performance liquid chromatography;

IPA refers to isopropyl alcohol;

Me refers to methyl;

MeOH refers to methanol;

MTBE refers to methyl tert-butyl ether;

min refers to a minute or minutes;

m/z refers to the ratio of mass to charge;

Ph refers to phenyl;

FDC refers to a round bottom flask;

RT refers to room temperature;

MSD refers to selective cation exchange;

SEM refers to the standard error of the mean;

SLC refers to supercritical liquid chromatography;

TFA refers to trifluoroacetic acid;

THF refers to tetrahydrofuran.

Scheme 1

Scheme 1 depicts a general method for preparing compounds of formula I. R ^1a may be the same as the R ¹ in the final compound of formula I, or it may be a group that requires conversion to achieve the R ¹ of formula I. Common synthetic transformations of R ^1a . such as BOC deprotection, ester hydrolysis, and amide coupling reactions can be carried out before or after Step B. In Step A, a Suzuki cross-coupling reaction between heteroaryl dichloride 1 and pyrazole boronate ester 2 gives the substituted pyrazole compound 3. This reaction is carried out using a base , for example 2 M aqueous Na ₂ CO ₃ , in an organic solvent, for example 1,4-dioxane, in the presence of a palladium catalyst such as tetrakis(triphenylphosphine)palladium(0) or bis(triphenylphosphine)palladium(II) dichloride, at elevated temperature . In Step B, compound 3 is subjected to a nucleophilic aromatic substitution reaction with a substituted azetidine 4 to produce a compound of formula I. This reaction is carried out in the presence of an organic base such as DIPEA in an organic solvent such as 1,4-dioxane at elevated temperature.

- 10 043635

Scheme 2

Scheme 2 depicts the preparation of a subset of compounds of formula I in which the pyrazole is replaced by a nitrogen-containing heterocycle (formula I'). Pyrazole boronate ester 5 is substituted with a nitrogen-containing heterocycle, as shown in Scheme 2, in which Y and Z can each independently be -CH2- or -CH2-CH2-, and PG is a nitrogen protecting group such as BOC. In Step C, the Suzuki cross-coupling reaction between heteroaryl dichloride 1 and pyrazole boronate ester 5 gives the substituted pyrazole compound 6. As described in Step A of Scheme 1, this reaction is carried out using a base, such as 2 M aqueous Na ₂ CO ₃ , in an organic solvent, for example 1,4-dioxane, in the presence of a palladium catalyst such as tetrakis(triphenylphosphine)palladium(0) or bis(triphenylphosphine)palladium(II) dichloride, at elevated temperature. Two different paths can be chosen, as shown in diagram 2 - path A and path B.

In route A, the substituted pyrazole compound 6 is deprotected in step D to produce compound 7. For example, if PG is BOC, deprotection can be accomplished with TPA. In step E, the heterocyclic nitrogen of compound 7 can undergo substitution reactions, such as reductive amination, acylation, or amide coupling, to yield compound 8. Such substitutions are designated R in this diagram. In step F, compound 8 is subjected to a nucleophilic aromatic substitution reaction with a substituted azetidine 4 to yield compounds of formula I'. As described in step B of Scheme 1, this reaction is carried out in the presence of an organic base, such as DIPEA, in an organic solvent, such as 1,4-dioxane, at elevated temperature. This may be the final step or, alternatively, the R group may be subjected to further routine synthetic transformations such as deprotection.

In Path B, the substituted pyrazole compound 6 undergoes a nucleophilic aromatic substitution reaction in step G with a substituted azetidine 4 to produce compound 9. This can be achieved by a nucleophilic aromatic substitution reaction with a substituted azetidine 4 in the presence of an organic base such as DIPEA in an organic solvent such as 1,4dioxane, at elevated temperature. Alternatively, steps C and G can be performed in a single procedure in which the Suzuki cross-coupling reaction in step C is completed first and then azetidine 4 is added to the reaction mixture along with an organic base (e.g. DIPEA) followed by step G continues at elevated temperatures. The PG protecting group is removed in step H (e.g., by TPA if PG is BOC) to produce compound 10. In step I, the nitrogen group of compound 10 can undergo substitution reactions, such as reductive amination, acylation, or amide coupling, to produce compound formula I'. This may be the final step or, alternatively, the R group may be subjected to further routine synthetic transformations such as deprotection.

- 11 043635

Scheme 3

Scheme 3 depicts the preparation of a subset of compounds of formula I in which the pyrazole is replaced by an acetamide group (formula I). In Step J, the Suzuki cross-coupling reaction between heteroaryl dichloride 1 and pyrazole boronate ester 11 gives the substituted pyrazole compound 12. As described in Schemes 1 and 2, this reaction is carried out using a base, such as 2 M aqueous Na ₂ CO ₃ , in an organic solvent, for example 1,4-dioxane, in the presence of a palladium catalyst such as tetrakis(triphenylphosphine)palladium(0) or bis(triphenylphosphine)palladium(II) dichloride, at elevated temperature. In step K, heteroaryl chloride 12 undergoes nucleophilic aromatic substitution with azetidine 4 in the presence of an organic base such as DIPEA in an organic solvent such as 1,4-dioxane at elevated temperature to give compound 13. In step L, hydrolysis of the ester using a base is carried out. for example, sodium hydroxide, gives acid 14. In step M, acid 14 undergoes an amide coupling reaction with an amine of formula HNR'R to produce an amide compound of formula I. The amine HNR'R may be cyclic (eg, optionally substituted with piperazine). Step M may be the final step, or additional routine synthetic transformations such as deprotection may occur.

Receipts and examples

The following preparations and examples further illustrate various embodiments of the present invention and demonstrate typical methods for synthesizing a compound of the invention. The reagents and starting materials are readily available or can be easily synthesized by those of ordinary skill in the art. It should be understood that the preparations and examples are provided for illustration and not limitation and that various modifications may be made by those of ordinary skill in the art.

LC-ES/MS was performed on an AGILENT® HP1200 liquid chromatography system. Electrospray mass spectrometry measurements (obtained in positive and/or negative mode) were performed on a quadrupole mass spectrometer with a mass selective detector connected to an HPLC system, which may or may not be equipped with an ELSD.

LC-MS conditions (low pH).

Column: PHENOMENEX® GEMINI® NX C18 2.0x50 mm, 3.0 µm, 110 A;

gradient: 5-95% B for 1.5 min, then 95% B for 0.5 min;

column temperature: 50±10°C;

flow rate: 1.2 ml/min;

injection volume: 1 µl;

Solvent A: deionized water with 0.1% HCOOH;

solvent B: ACN with 0.1% formic acid;

wavelength 200-400 and 212-216 nm.

If the HPLC is equipped with an ELSD, the settings are: evaporator temperature 45°C, nebulizer temperature 40°C and gas flow 1.6 stb. l/min.

Alternative LC-MS conditions (high pH).

Column: Waters xBridge® C18 column 2.1x50 mm, 3.5 µm;

gradient: 5-95% B for 1.5 min, then 95% B for 0.50 min;

column temperature: 50±10°C;

flow rate: 1.2 ml/min;

injection volume: 1 µl;

- 12 043635 solvent A: 10 mM NH ₄ HCO ₃ , pH 9;

solvent B: ACN;

wavelength: 200-400 and 212-216 nm;

with ELSD: evaporator temperature 45°C, atomizer temperature 40°C and gas flow 1.60 st. l/min.

RPD profiles of crystalline solids were recorded on a Bruker D4 Endeavor X-ray powder diffractometer equipped with a CuKa source and a Vantec detector operating at 35 kV and 50 mA. The sample was scanned from 4 to 40 2θ° with a step size of 0.008 2θ° and a scan speed of 0.5 s/step using a divergence slit of 1.0 mm, a fixed anti-scattering slit of 6.6 mm, and a detector slit of 11.3 mm. The dry powder was packed into a quartz sample holder and a smooth surface was obtained using a glass slide. Diffraction profiles of the crystalline form were recorded at ambient temperature and relative humidity. Crystal peak positions were determined in MDI-Jade after fully shifting the diffraction patterns based on the internal standard NIST 675 with peaks at 8.853 and 26.774 2θ°. It is well known in the field of crystallography that for any given crystal form, the relative intensities of the diffraction peaks can vary due to preferred orientation caused by factors such as morphology and crystal structure. If the influence of preferred orientation is present, the peak intensities change, but the characteristic peak positions of the polymorph remain unchanged. See, for example, The United States Pharmacopeia #23, National Formulary #18, pages 1843-1844, 1995. Additionally, it is also well known in the field of crystallography that for any given crystal shape, the angular positions of the peaks may vary slightly. For example, peak positions may shift due to changes in the temperature at which the sample is analyzed, sample drift, or the presence or absence of an internal standard. In this case, it is assumed that the peak position variability of ±0.2 2θ° takes into account these possible deviations without interfering with the accurate determination of the specified crystal form. Confirmation of the crystal form can be based on any unique combination of characteristic peaks.

Receipt 1.

(2S)-1-Benzhydryl-2-methylazetidine-[(1R,4S)-7,7-dimethyl-2-oxonorbornan-1yl]methanesulfonic acid salt.

A 2000 mL three-neck FDC was connected to a dropping funnel, a nitrogen inlet, and a thermometer adapter. The vessel was purged with nitrogen and (3R)-butane-1,3-diol (25 g, 277 mmol), DIPEA (127 mL, 731 mmol) and ACN (556 mL) were added. The mixture was cooled to -30°C. Trifluoromethanesulfonic acid anhydride (101 mL, 601 mmol) was added dropwise over 3 hours so that the internal temperature was maintained at -35 to -30°C. After addition was complete, stirred for 10 minutes at -35 to -30°C. Trifluoromethanesulfonic acid anhydride (1.9 mL, 11 mmol) was added dropwise over 5 min to maintain the internal temperature at -35 to -30°C. After addition was complete, stirred for 10 minutes at -35 to -30°C. DIPEA (127 mL, 731 mmol) was added dropwise over 15 min to maintain the internal temperature at -35 to -30°C. After addition was complete, stirred for 10 minutes at -35 to -30°C. In a separate flask, aminodiphenylmethane (48.0 ml, 270 mmol) was dissolved in ACN (49 ml) under nitrogen atmosphere and the resulting solution was transferred to a dropping funnel. The amine solution was added dropwise to the cold triflate over 40 min to maintain the internal temperature between -20 and -35°C. After addition was complete, stirred for 30 minutes at -35 to -30°C. The reaction mixture was transferred to a water bath and allowed to warm slowly for 30 minutes. The bath was removed and the reaction mixture was allowed to warm to RT for 30 min. The vessel was transferred to a heating mantle and the reaction mixture was heated to 45°C for 30 min, then cooled to RT. The resulting mixture was poured into 1200 ml of water and extracted with toluene (400 mlx3). The extracts were combined and washed with water and saturated aqueous NaCl. The organic layers were dried over anhydrous Na ₂ SO ₄ , filtered and concentrated in vacuo. The residue was dried in vacuo overnight, then dissolved in DCM (400 ml). A silica gel layer was prepared on a sintered bed funnel and equilibrated with a 1:1 heptane/EtOAc mixture. The product solution was loaded onto a silica gel bed and washed with 1600 ml heptane/EtOAc 1:1. The filtrate was concentrated to obtain a red oily substance. The oily substance was dissolved in MeOH (250 ml) and the flask was placed in a water bath (~10°C). L(-)-camphorsulfonic acid (61.6 g, 265 mmol) was added in portions while maintaining the internal temperature below 20°C. The resulting mixture was stirred for 15 minutes and then concentrated in vacuo to obtain a brown foamy substance.

- 13 043635

The foam was dried on a vacuum pump for 2 hours. The foam was dissolved in DCM (130 ml), then EtOAc (1100 ml) was slowly added to the stirred solution through a dropping funnel. The resulting mixture was transferred to a 4000 ml beaker and stirred in air overnight. The beaker was cooled in an ice bath for 10 minutes. The precipitate was collected in a sintered funnel by vacuum filtration, washing with a minimal amount of ice-cold EtOAc. The solid was dried on the sintered bed for 2 hours. The resulting white solid was dissolved in a minimal amount of DCM, transferred to a 2000 ml beaker and then diluted slowly with EtOAc until the clear solution became cloudy. The suspension was stirred for 4 hours in the open air. The solids were collected by vacuum filtration using a sintered bed funnel and dried on the sintered bed overnight to obtain the title compound (111.8 g, 238.06 mmol, 86% yield) as a white solid.

1H NMR (400 MHz, DMSOL): δ 10.54-10.47 (m, 1H), 7.61 (d, J=7.3 Hz, 5H), 7.47-7.37 (m, 7H ), 5.85 (d, J=10.3 Hz, 1H), 4.68-4.61 (m, 1H), 3.91-3.83 (m, 2H), 3.37 (s, 8H), 2.99 (d, J=14.6 Hz, 1H), 2.77-2.68 (m, 1H), 2.51-2.44 (m, 4H), 2.30-2 .16 (m, 2H), 1.91-1.81 (m, 2H), 1.42-1.28 (m, 3H), 1.08 (s, 3H), 1.01 (d, J =6.6 Hz, 3H), 0.77 (s, 4H); ee >98% (HPLC: Chiralcel® OJ (10 cm x 4.6 mm, 5 µm), 5 ml/min, 40°C isocratic 10% EtOH (0.2% iPrNH2)/CO2).

Receipt 2.

[(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonate(2S)-2-methylazetidin-1-ium salt.

Η

20 wt.% Pd(OH) ₂ on carbon (6.62 g) was added to a 2250 ml Parr flask. The flask was purged with nitrogen and MeOH (250 ml) was added. (2S)-1-Benzhydryl2-methylazetidine-[(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonic acid salt (111 g, 236 mmol) dissolved in MeOH was slowly added to the resulting suspension (250 ml). The vessel was sealed. Purge with nitrogen, then hydrogen, and increase the pressure to 60 psi. inch. The reaction vessel was shaken vigorously in a Parr mixer for 15 h at RT. The vessel was purged with nitrogen and the reaction mixture was then filtered through a pad of Celite®, rinsing with MeOH. The filtrate was concentrated to obtain a white solid and dried under vacuum. The solid was suspended in 780 ml of a 1:1 MTBE/EtOAc mixture and the mixture was heated to 65°C for 20 hours, then cooled to RT and stirred overnight. Solids were collected by filtration. The solids were suspended in 380 mL of MTBE and stirred at RT for 24 hours. The white solid was collected by filtration to give the title compound (41.5 g, 137 mmol, 58% yield).

1H NMR (400 MHz, DMSO-d6): δ 8.68-8.55 (m, 1H), 4.51-4.42 (m, 1H), 3.91-3.75 (m, 1H) , 3.36 (s, 3H), 2.91 (d, J=14.6 Hz, 1H), 2.69-2.61 (m, 1H), 2.52-2.46 (m, 2H ), 2.28-2.22 (m, 1H), 2.17-2.10 (m, 1H), 1.96 (t, J=4.5 Hz, 1H), 1.89-1, 79 (m, 1H), 1.43 (d, J=6.7 Hz, 2H), 1.36-1.26 (m, 1H), 1.05 (s, 2H), 0.75 (s , 2H).

Receipt 3.

(R)-2-azetidinemethanol hydrochloride.

OH

Qj CIH H

(R)-1-(tert-butoxycarbonyl)azetidine-2-carboxylic acid (30 g, 146 mmol), THF (300 mL), and 4-methylmorpholine (17.7 mL, 161 mmol). The mixture was cooled to -10°C and isobutyl chloroformate (21 ml, 161 mmol) was added dropwise. The mixture was stirred for 30 min, then heated to RT. The resulting solid was collected by filtration. The filtrate was cooled to 0°C and a solution of sodium borohydride (11.1 g, 292 mmol) in water (90 ml) was added dropwise (caution: gas evolution). After addition, the mixture was heated to RT and stirred for 30 minutes. The mixture was diluted with MTBE (300 ml) and water (100 ml). The mixture was washed with saturated aqueous NaHCO ₃ (200 ml) and then saturated aqueous NaCl (200 ml). The organic phase was dried over MgSO ₄ , filtered and concentrated to dryness to give an oil (27 g). HCl (4.0 M) in 1,4-dioxane (110 ml) was added carefully (caution: gas evolution) and the resulting mixture was stirred for 3 hours at RT. The solvent was evaporated in vacuo to obtain the title compound as an oil (16 g, 89%). This material was immediately used in production methods 9, 11, 16 and 36.

- 14 043635

Receipt 4.

(2S,3R)-2-methylazetidin-3-ol-[(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonic acid.

Stage 1.

A 500 ml three-neck CDC was equipped with a dropping funnel and a temperature sensor. But-2-en-1-ol (cis/trans) (23.7 mL, 267 mmol) and chloroform (200 mL) were added to the flask. The solution was cooled in an ice bath until the internal temperature reached 1.2°C. Bromine (13.7 mL, 267 mmol) was added dropwise through an addition funnel over 2 h at a rate of ~1 drop/6 s. After addition was complete, the reaction mixture was warmed to RT and stirred for 30 minutes. Stirring was turned off and the reaction mixture was allowed to stand for 3 days. They extinguished us. Na ₂ S ₂ O ₃ solution and stirred vigorously for 10 minutes. The mixture was allowed to sit for 3 days. The organic layer was removed and the aqueous fraction was extracted with DCM (x3). The organic layers were combined and dried over Na ₂ SO ₄ , filtered and concentrated in vacuo to give 2,3-dibromobutan-1-ol (62.3 g, 269 mmol).

Stage 2.

Procedure A.

A 1-liter, 3-neck CDC was equipped with a dropping funnel and a temperature sensor. 2,3-dibromobutan-1-ol (62.3 g, 269 mmol) and THF (180 ml) were added to the flask. The flask was placed in a water bath at RT. A solution of KOH (15.1 g, 269 mmol) in water (135 ml) was added dropwise through a dropping funnel over 10 min. Stir at RT for 2 hours. The organic phase is separated. The aqueous fraction was extracted with EtOAc 3x150 ml. The organic layers were combined, washed once with ~200 ml brine, dried over Na ₂ SO ₄ and filtered. The organics were carefully concentrated (100 mbar, 30°C to minimum volume, then 10 mbar, 30°C for 10 min) to obtain 37 g of a mixture of 2-(1 bromoethyl)oxirane (60 wt%), 2,3-dibromobutane -1-ol (36 wt%) and EtOAc (4 wt%), as determined by 1H NMR. EtOH (100 ml), aminodiphenylmethane (36 ml, 208.6 mmol) and NaHCO ₃ (26 g, 309 mmol) were added to the mixture. The reaction mixture was heated to 65°C overnight. Cooled until CT.

Procedure B.

2,3-dibromobutan-1-ol (72.6 g, 313 mmol) and THF (200 ml) were added to the first flask. The flask was placed in a water bath at RT. A solution of KOH (17.6 g, 314 mmol) in water (150 ml) was added. Stir at RT overnight. The organic phase was separated. The aqueous fraction was extracted with EtOAc (3x150 ml). The organic layers were combined, washed once with ~200 ml of saturated aqueous NaCl, dried over Na ₂ SO ₄ and filtered. The organic layer was concentrated in vacuo (100 mbar, 30°C to minimum volume, then 10 mbar, 30°C for 10 min) to obtain 41.1 g of a mixture of 2-(1 bromoethyl)oxirane (75 wt.%), 2. 3-dibromobutan-1-ol (22 wt%) and EtOAc (3 wt%), as determined by 1H NMR.

2,3-dibromobutan-1-ol (10 g, 43 mmol) and THF (30 ml) were added to the second flask. The flask was placed in a water bath at RT. A solution of KOH (2.42 g, 43.1 mmol) in water (20 ml) was added. Stir at RT overnight. The organic phase was separated. The aqueous fraction was extracted with EtOAc 3x50 ml. The organic layers were combined, washed once with ~200 ml of saturated aqueous NaCl, dried over Na ₂ SO ₄ and filtered. The organic layer was concentrated in vacuo (100 mbar, 30°C to minimum volume, then 10 mbar, 30°C for 10 min) to obtain 5.4 g of a mixture of 2-(1 bromoethyl)oxirane (66 wt.%), 2. 3-dibromobutan-1-ol (32 wt%) and EtOAc (2 wt%), as determined by 1H NMR.

2,3-dibromobutan-1-ol (10 g, 43 mmol) and THF (30 ml) were added to the third flask. The flask was placed in a water bath at RT. A solution of KOH (2.42 g, 43.1 mmol) in water (20 ml) was added. The mixture was heated to 50°C for 2 hours. Cooled to RT. The organic phase was separated. The aqueous fraction was extracted with EtOAc 3x50 ml. The organic layers were combined, washed once with ~200 ml brine, dried over Na ₂ SO ₄ and filtered. The organic layer was concentrated in vacuo (100 mbar, 30°C to minimum volume, then 10 mbar, 30°C for 10 min) to obtain 5.4 g of a mixture of 2-(1-bromoethyl)oxirane (70 wt%), 2,3-dibromobutan-1-ol (28 wt%) and EtOAc (2 wt%), as determined by 1H NMR.

Mixtures of the three reactions were combined together in the FDC to obtain 51.9 g of 2-(1-bromoethyl)oxirane (73 wt%). EtOH (100 ml), aminodiphenylmethane (44 ml, 255.0 mmol) and NaHCO ₃ (32 g, 380.926 mmol) were added. The resulting mixture was stirred at RT for 2 hours, then heated to 65°C and stirring continued overnight. Cooled until CT. The crude reaction mixtures from Procedures A and B were combined together for purification. Solids were removed by filtration, washing with EtOH. The filtrate was concentrated to dryness. The resulting oily substance was dissolved in DCM. The resulting solution was washed twice with NH ₄ Cl solution, dried over Na ₂ SO ₄ , filtered and concentrated to

- 15 043635 volume ~150 ml. The mixture was allowed to sit overnight. The solid was removed by filtration. The filtrate was purified by normal phase silica gel chromatography (70% MTBE:hexanes) to give crude 1-benzhydryl-2-methylazetidin-3-ol (66.8 g).

ER/MS (m/z): 254 (M+H).

Stage 3.

1-Benzhydryl-2-methylazetidin-3-ol (66.8 g) was dissolved in MeOH (608 ml). The resulting solution was purified using a Lux i-Cellulose 5 5x25 cm column using a solvent system of 85/15 CO ₂ /EtOH with 0.5% dimethylethylamine and a flow rate of 300 ml/min to obtain (2S,3R)-1-benzhydryl-2methylazetidine -3-ol (19.2 g).

Stage 4.

The CDC was loaded with (2S,3R)-1-benzhydryl-2-methylazetidin-3-ol (19.2 g, 75.8 mmol), L(-)-camphorsulfonic acid (19 g, 80.2 mmol), EtOH (100 ml) and Et ₂ O (50 ml). The mixture was heated until almost all solids were dissolved, then briefly sonicated. The mixture was heated to boiling, then cooled to rt and stored in the freezer overnight. The solids were collected by filtration, washed with a large volume of Et ₂ O and dried under reduced pressure to obtain (2S,3R)-1-benzhydryl-2-methylazetidin-3-ol, [(1R,4S)-7,7-dimethyl-2oxonorbornane -1-yl]methanesulfonic acid (31.9 g, 65.7 mmol).

Stage 5.

20% palladium hydroxide on carbon (50 wt.% water) (2 g) was loaded into the FDC. The catalyst was wetted with a small volume of EtOH. A partial solution of (2S,3R)-1-benzhydryl-2methylazetidin-3-ol was added to the suspension; [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonic acid (20 g, 41.2 mmol) in EtOH (400 ml). The suspension was bubbled with nitrogen for 5 min and then briefly bubbled with hydrogen. The reaction mixture was stirred in a hydrogen balloon until all starting material had disappeared as determined by LC-MS analysis. The reaction mixture was filtered through a pad of Celite®.

In a second flask, (2S,3R)-1-benzhydryl-2-methylazetidin3-ol was hydrogenated using the same procedure; [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonic acid (10 g, 20.6 mmol) with 20% palladium hydroxide on carbon (50 wt.% water) (1 d) in EtOH (200 ml).

In a third flask, (2S,3R)-1-benzhydryl-2-methylazetidin3-ol was hydrogenated using the same procedure; [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonic acid (2 g, 4.12 mmol) with 20% palladium hydroxide on carbon (50 wt.% water) (0 .2 g) in EtOH (40 ml).

The three filtrates were combined together and concentrated in vacuo. The material was suspended in nheptane and sonicated for 10 min, and then the solids were collected by filtration. The sequence of ultrasonication and filtration was repeated four more times. The solid was dried in vacuo overnight to give the title compound (20.11 g, 53% yield over 5 steps).

Ή NMR (399.85 MHz, MeOD): δ 4.35-4.25 (m, 2H), 4.07 (dd, J=6.9, 10.6 Hz, 1H), 3.78 (dd , J=7.0, 10.6 Hz, 1H), 3.30 (m, 1H), 2.79 (d, J=14.9 Hz, 1H), 2.70-2.63 (m, 1H), 2.40-2.33 (m, 1H), 2.08-2.03 (m, 2H), 1.92 (d, J=18.4 Hz, 1H), 1.69-1 .62 (m, 1H), 1.54 (d, J=6.6 Hz, 3H), 1.47-1.41 (m, 1H), 1.14 (s, 3H), 0.88 ( s, 3H).

Receiving 5.

Ethyl 2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrazol-1-yl)acetate.

4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-III-pyrazole (15 g, 77.32 mmol), ACN (150 ml) were loaded into a 500 ml FDC. , potassium carbonate (32.06 g, 232.0 mmol) and ethyl bromoacetate (9.09 ml, 81.2 mmol). The suspension was stirred at RT overnight. The reaction mixture was partitioned between EtOAc (300 ml) and water (250 ml). The layers were separated and the aqueous layer was extracted with EtOAc. The organic layers were combined and washed with saturated aqueous sodium chloride (500 ml), dried over sodium sulfate, filtered and concentrated in vacuo to give a light yellow oil. Purify by chromatography on silica gel using a gradient of 0 to 5% MeOH in DCM to give the title compound (16.8 g, 76% yield).

¹ H NMR (400 MHz, DMSO-d ₆ ): δ 7.94 (s, 1H), 7.60 (s, 1H), 5.08 (s, 2H), 4.14 (q, J=7 ,1 Hz, 2H), 1.26 (m, 12H), 1.02 (t, J=7.2 Hz, 3H).

- 16 043635

Receipt 6.

2-[4-[2-[(28)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl]acetic acid.

Ethyl 2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrazol-1yl)acetate (39.15 g, 132.8 mmol), 2,4-dichloro-6-(trifluoromethyl)pyrimidine (27.9 g, 126 mmol), 1,4-dioxane (800 ml), aqueous Na ₂ CO ₃ (2 M, 200 ml, 400 mmol) and bis(triphenylphosphine)palladium(II) dichloride (2.8 g, 4.0 mmol). The mixture was heated to 85°C. After 2 hours the mixture was cooled to RT. The reaction mixture was divided into two parts and processed according to the following methods.

Method A.

[(1R,48)-7,7-dimethyl-2-oxonorbornan-1yl]methanesulfonate (28)-2-methylazetidin-1-ium salt (18.69 g, 59.75 mmol) was added to the first portion of the reaction mixture and heated to 70°C for 2.5 hours. Cooled to RT. Aqueous NaOH (2 M, 166 mL, 332 mmol) was added and stirred at RT overnight. EtOAc (500 ml) was added and the mixture was stirred for 30 minutes. The mixture was acidified to pH 7 using aqueous HCl (5 M) and extracted with EtOAc (4x250 ml). The organic layers were combined, washed with saturated aqueous NaCl, dried over Na ₂ SO ₄ , filtered and concentrated in vacuo. The residue was dissolved in DCM (125 ml) and heptane (125 ml) was added dropwise. The mixture was stirred for 30 minutes. The solid was collected by filtration and washed with 1:1 DCM/heptane (50 mL). The solid was air dried.

Method B.

The second portion of the reaction mixture was concentrated in vacuo to remove 1,4-dioxane. The mixture was partitioned between EtOAc and water. The aqueous layer was separated and the organic layer was concentrated to dryness. To the flask containing the residue was added 1,4-dioxane (420 ml), aqueous Na ₂ CO ₃ (2 M, 75 ml, 150 mmol) and salt [(1R,4S)-7,7-dimethyl-2-oxonorbornane -1-yl]methanesulfonate (28)-2-methylazetidin-1-ium (18.69 g, 59.75 mmol). The mixture was heated to 70°C and stirred for 1.5 hours. The mixture was cooled to RT. Aqueous NaOH (2 M, 125 mL, 250 mmol) was added and stirred overnight. EtOAc (500 ml) was added and stirred for 30 minutes. The aqueous phase was separated and left to settle overnight. The aqueous mixture was acidified to pH 7 using aqueous HCl (5 M) and extracted with EtOAc (2x250 ml). The organic layers were combined, washed with saturated aqueous NaCl, dried over Na ₂ SO ₄ , filtered and concentrated in vacuo to give a solid.

Cleaning method.

The products from methods A and B were combined and dissolved in THF (485 ml). SiliaMetS® thiol resin (32 g) was added. The mixture was stirred for 1 hour and then filtered. The filtrate was concentrated in vacuo to obtain the title compound as a white powder (31.7 g, 72%).

ER/MS (m/z): 342 (M+H); 340 (M-H).

Receipt 7.

Tert-butyl-4-[2-[4-[2-[(28)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1yl] acetyl]piperazine-1 - carboxylate.

2-[4-[2-[(28)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1-yl]acetic acid (31.7 g) was loaded into the FDC per 1 liter , 91.0 mmol), tert-butylpiperazine-1-carboxylate (20.5 g, 109 mmol) and DCM (320 ml). Et ₃ N (25.6 ml, 182 mmol) was added to the resulting solution, followed by

- 17 043635 drops solution of 2,4,6-tripropyl-1,3,5,2,4,6-trioxatriphosphorinan-2,4,6-trioxide anhydride (50 wt.% in EtOAc, 68 ml, 114.2 mmol ). Stir at RT for 1 hour. The reaction mixture was washed with water (500 ml) and then with saturated aqueous NaCl. The organic layers were dried over Na ₂ SO ₄ , filtered and concentrated under high vacuum to give the title compound (52.5 g, purity estimated at 88 wt.%, based on theoretical quantitative yield of product).

ER/MS (m/z): 510 (M+H); 508 (M-H).

Receiving 8.

Tert-butyl-4-[4-[2-chloro-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl]piperidine-1-carboxylate.

2,4-Dichloro-6-(trifluoromethyl)pyrimidine (0.7 g, 3.05 mmol) and 1,4-dioxane (15 ml) were added to a microwave-safe tube. Tert-butyl-4-(4-(4,4,5,5-tetramethyl-1,3,2dioxaborolan-2-yl)-1P-pyrazol-1-yl)piperidin-1-carboxylate (1, 55 g, 4.0 mmol), tetrakis(triphenylphosphine) palladium (0) (0.195 g, 0.17 mmol) and aqueous Na ₂ CO ₃ (2 M, 5.5 ml, 11 mmol). The tube was sealed and heated in a microwave reactor at 85°C for 1 hour. In a second tube, the same reaction was carried out on the same scale. The reaction mixtures were combined in a separatory funnel. The reaction mixture was diluted with saturated aqueous sodium bicarbonate and extracted twice with EtOAc. The extracts were combined and dried over Na ₂ SO ₄ , filtered and concentrated. The residue was purified by silica gel chromatography using a gradient of 0 to 50% EtOAc/hexanes to give the title compound as a white solid (2.33 g, 88%).

ER/MS (m/z): 432, 434 (M+H); 430, 432 (M-H).

Receipt 9.

Tert-butyl-4-[4-[2-[(2R)-2-(hydroxymethyl)azetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1 -yl]piperidin-1 is a carboxylate.

Mix tert-butyl 4-[4-[2-chloro-6(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl]piperidin-1-carboxylate (1 g, 2.3 mmol) and 1,4-dioxane (16 ml). (R)-2-azetidinemethanol hydrochloride (0.52 g, 4.3 mmol) and DIPEA (1.6 mL, 9.3 mmol) were added. The tubes were sealed and heated in a microwave reactor at 130°C for 2.5 hours. The resulting reaction mixtures were combined, then the mixture was diluted with saturated aqueous NaHCO ₃ and extracted twice with EtOAc. The extracts were combined and dried over Na ₂ SO ₄ , filtered and concentrated. The residue was purified by silica gel chromatography using a gradient of 10 to 80% EtOAc/hexanes to give the title compound as a white foam (3.2 g, 95%).

ER/MS (m/z): 483 (M+H).

- 18 043635

Getting 10.

Tert-butyl 3-[4-[2-chloro-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl]azetidine-1-carboxylate.

2,4-Dichloro-6-(trifluoromethyl)pyrimidine (0.50 g, 2.30 mmol), 1,4-dioxane (22 ml) and water (2 ml) were added to a microwave reactor tube. Tert-butyl 3-(4-(4,4,5,5tetramethyl-1,3,2-dioxaborolan-2-yl)-III-pyrazol-1-yl)azetidin-1-carboxylate (0.630 g) was added to the solution , 1.71 mmol), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (II) (0.20 g, 0.26 mmol) and K ₂ CO ₃ (500 mg, 3.67 mmol). The tube was sealed and stirred at RT overnight. The crude reaction mixture was loaded into a silica cartridge, dried in a vacuum oven, and the residue was purified by flash chromatography (0 to 45% EtOAc/hexane gradient) to obtain the title compound as a light brown oil (0.375 g, 49%).

ER/MS (m/z): 348 (M+H-tBu); 402 (M-H).

Receipt 11.

Tert-butyl-3-[4-[2-[(2R)-2-(hydroxymethyl)azetidin-1-yl]-6-(trifluoromethyl)nirimidin-4yl]pyrazol-1 -yl]azetidin-1-carboxylate.

In a test tube, tert-butyl 3-[4-[2-chloro-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1yl]azetidine-1-carboxylate (190 mg, 0.47 mmol) and THF (18 ml). (R)-2-azetidinemethanol hydrochloride (150 mg, 1.23 mmol) and DIPEA (1.0 ml, 5.7 mmol) were added. The tube was sealed and heated in a microwave reactor at 100°C for 1 hour 40 minutes. After cooling to RT, the crude reaction mixture was loaded into a silica cartridge, dried in a vacuum oven, and the residue was purified by silica gel chromatography using a 0 to 80% (5% MeOH/EtOAc)/hexanes gradient to obtain the title compound as a colorless oily substance (153 mg, 72%).

ER/MS (m/z): 455 (M+H).

Receiving 12.

Tert-butyl-3-[4-[2-[(2S,3R)-3-hydroxy-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1 -yl]azetidin-1 -carboxylate.

The title compound was prepared essentially as described in Preparation 11 using (2S,3R)-2-methylazetidin-3-ol [(1R,4S)-7,7-dimethyl-2-oxo-norbornane-1- yl]methanesulfonic acid

- 19 043635 slot.

ER/MS (m/z): 455 (M+H).

Receipt 13.

Tert-butyl 3-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1yl]azetidin-1-carboxylate.

The title compound was prepared essentially as described in Preparation 11 using [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonate (2S)-2-methylazetidin-1-ium salt . The reaction mixture was purified by silica gel chromatography using 45% EtOAc/hexanes.

ER/MS (m/z): 439 (M+H).

Receipt 14.

2-[4-[5-cyαno-6-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl]acetic acid.

Ethyl 2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrazol-1-yl)acetate (2.95 g, 10.5 mmol) was loaded into the CDC ), 2,6-dichloro-4-(trifluoromethyl)pyridine-3-carbonitrile (2 g, 8.3 mmol), 1,4-dioxane (52.4 ml) and aqueous Na ₂ CO ₃ (2 M, 13 .3 ml, 26.6 mmol). The mixture was bubbled with nitrogen for 10 minutes. Bis(triphenylphosphine)palladium(II) dichloride (191 mg, 0.27 mmol) was added. The mixture was stirred at RT for 16.5 hours. The flask was equipped with a reflux condenser and [(1R,4S)-7,7-dimethyl2-oxonorbornan-1-yl]methanesulfonate (2S)-2-methylazetidin-1-ium salt (2, 73 g, 9.10 mmol). The mixture was heated to 70°C and stirred for 3 hours, then cooled to RT. Aqueous NaOH (2 M, 21 mL, 42 mmol) was added and the mixture was stirred at RT for 15 min. The organic solvent was evaporated in vacuum. The residue was diluted with water (50 ml) and the water was filtered from the solid. This process was repeated with water (50 ml) twice. The solid was collected by vacuum filtration and air dried overnight. 2-methyltetrahydrofuran (30.4 ml) and aqueous citric acid (6.5 wt%, 30.4 ml) were added. Stir for 5 minutes and then separate the layers. The organic layer was washed with aqueous citric acid (6.5 wt.%, 30.4 ml). The organic solution was dried over MgSO ₄ , filtered and concentrated to give the title compound (3.14 g, purity estimated at 97 wt.% based on theoretical quantitative yield of product).

ER/MS (m/z): 366 (M+H); 364 (M-H).

Receiving 15.

Tert-butyl 4-[4-[6-chloro-5-cyano-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl]piperidine-1carboxylate.

Method A.

2,6-Dichloro-4-(trifluoromethyl)pyridine-3 was added to the microwave reactor tube

- 20 043635 carbonitrile (497 mg, 2.063 mmol), tert-butyl-4-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2yl)pyrazol-1-yl]piperidin- 1-carboxylate (867.2 mg, 2.3 mmol), 1,1'-bis(diphenylphosphino)ferrocenepalladium(II) dichloride complex and DCM (91.2 mg, 0.109 mmol), aqueous potassium carbonate (3 M solution, 2.1 ml, 6.3 mmol) and 1,4-dioxane (10.5 ml). The mixture was bubbled with nitrogen for 5 min, sealed and heated to 80°C. After 1 hour, cool to RT. The mixture was filtered through a pad of Celite® and washed with EtOAc. The residue was concentrated and purified by silica gel chromatography using a gradient of 15 to 40% EtOAc in hexanes, then quickly dried in vacuo at 35°C to give the title compound (705 mg, 64%).

ER/MS (m/z): 454, 456 (M-H).

Method B.

2,6-Dichloro-4-(trifluoromethyl)pyridine-3-carbonitrile (1.07 g, 4.44 mmol) and 1,4-dioxane (25 ml) were added to the FDC. Add tert-butyl 4-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrazol-1yl]piperidine-1-carboxylate (2.08 g, 5. 51 mmol), tetrakis(triphenylphosphine) palladium (0) (200 mg, 0.17 mmol) and aqueous Na ₂ CO ₃ (2 M, 8.5 ml, 17 mmol). The reaction mixture was degassed with nitrogen and heated at 85°C for 1.5 hours. The organic phase was filtered from the reaction mixture and the crude reaction mixture was divided into two equal quantities for use in preparations 16 and 17.

ER/MS (m/z): 454 (M-H).

Getting 16.

Tert-butyl-4-[4-[5-cyano-6-[(2R)-2-(hydroxymethyl)azetidin-1-yl]-4-(trifluoromethyl)-2pyridyl]pyrazol-1 -yl]piperidin-1 -carboxylate.

To a microwave reactor tube was added _1/2 of the crude reaction product from Method B for Preparation 15, (R)-2-azetidinomethanol hydrochloride (0.5 g, 4 mmol), DIPEA (1.6 mL, 9.2 mmol) and additional amount of 1,4-dioxane (16 ml). The suspension was heated at 120°C for 2 hours. The reaction mixture was cooled and concentrated to the crude product. The residue was purified by silica gel chromatography using a gradient of 50 to 80% EtOAc/hexanes and purified again by silica gel chromatography using a gradient of 2 to 5% IPA/DCM to obtain the title compound as a tan solid ( 264 mg, 26%).

ER/MS (m/z): 507 (M+H).

Receipt 17.

Tert-butyl-4-[4-[5-cyano-6-[(2S,3R)-3-hydroxy-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2pyridyl]pyrazol-1 -yl] piperidine-1-carboxylate.

The title compound was prepared essentially as described in Preparation 16 using the second _1/2 of the crude reaction product from Preparation 15 and (2S,3R)-2-methylazetidin-3-ol [(1R,4S)-7,7 -dimethyl-2-oxonorbornan-1-yl]methanesulfonic acid.

ER/MS (m/z): 507 (M+H).

- 21 043635

Receiving 18.

2-chloro-6-[1-(4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridine-3-carbonitrile hydrochloride.

Tert-butyl 4-[4-[6-chloro-5-cyano-4-(trifluoromethyl)-2-pyridyl]pyrazol-1yl]piperidine-1-carboxylate (705 mg, 1.5 mmol) was added to the test tube. 4 M HCl in 1,4-dioxane (1.5 ml, 6.0 mmol) and DCM (1.5 ml). The mixture was stirred at RT. After 1 hour, the mixture was concentrated in vacuo and the residue was dried in vacuo overnight to give the title compound (608 mg), which was used as is without further purification in the preparation of 19 and 20.

ER/MS (m/z): 356, 358 (free base M+H).

Receiving 19.

2-chloro-6-[1 -(1 -methyl-4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridine-3-carbonitrile.

2-chloro-6-[1-(4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridine-3carbonitrile hydrochloride (303.3 mg, 0.74 mmol), 13.3 M formaldehyde was added to the test tube in water (0.2 ml, 3 mmol), sodium triacetoxyborohydride (378 mg, 1.73 mmol) and 1,2-dichloroethane (6.0 ml). The mixture was stirred at RT for 30 min. Saturated aqueous sodium bicarbonate was added and extracted twice with DCM. The extracts were combined and concentrated in vacuo to give the title compound (285 mg, 99% yield).

ER/MS (m/z): 370, 372 (M+H).

Getting 20.

Tert-butyl 4-[4-[5-cyano-6-[(28)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1yl]piperidine-1-carboxylate.

The title compound is prepared from 2,6-dichloro-4-(trifluoromethyl)pyridine-3 carbonitrile and tert-butyl-4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane- 2-yl)-1H-pyrazol-1-yl)piperidine 1-carboxylate using essentially the same procedure as in Example 10 below.

ER/MS (m/z): 491 (M+H).

Receiving 21.

Tert-butyl-4-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1yl]piperidin-1-carboxylate.

- 22 043635

2,4-Dichloro-6-(trifluoromethyl)pyrimidine (0.258 g, 1.1 mmol) was dissolved in 1,4-dioxane (6 ml) and tert-butyl-(4-(4-(4,4,5 ,5-tetramethyl)-1,3,2-dioxaborolan-2-yl)-1H-nirazol-1-yl)piperidine-1carboxylate (0.591 g, 1.52 mmol), tetrakis(triphenylphosphine)palladium (0) (68 mg, 0.059 mmol) and 2 M aqueous sodium carbonate (2.2 ml, 4.4 mmol). The mixture was heated at 85°C in a microwave reactor for 1 hour. Cooled to RT and [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1yl]methanesulfonate (2S)-2-methylazetidine-1- salt was added. ia (0.414 g, 1.36 mmol) and DIPEA (0.6 ml, 3.4 mmol). The mixture was heated at 110°C for 2 hours. The mixture was diluted with saturated aqueous sodium bicarbonate and extracted twice with EtOAc. The extracts were combined, dried over sodium sulfate, then filtered and evaporated. The residue was purified by silica gel chromatography using a gradient of 0 to 50% EtOAc/hexanes to give the title compound (215 mg, 41%) as a white solid.

ER/MS (m/z): 467 (M+H).

Receiving 22.

Tert-butyl-3-[4-[6-chloro-5-cyano-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl]azetidine-1-carboxylate.

3-cyano-2,6-dichloro-4-(trifluoromethyl)pyridine (0.360 g, 1.49 mmol) was added to the test tube.

1,4-dioxane (7.5 ml), tert-butyl-3-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-III-pyrazol-1 yl)azetidine-1-carboxylate (0.727 g, 1.98 mmol), tetrakis(triphenylphosphine) palladium (0) (95 mg, 0.08 mmol) and 2 M aqueous sodium carbonate (2.8 ml, 5.6 mmol , 2 mol/l). The vessel was sealed and heated in a microwave reactor at 85°C for 1 hour. Diluted with saturated sodium bicarbonate and extracted twice with EtOAc. The extracts were combined, dried over sodium sulfate, filtered and evaporated. The residue was purified by silica gel chromatography using a gradient of 0 to 90% EtOAc/hexanes to give the title compound (0.609 g, 95%).

ER/MS (m/z): 426, 428 (M-H).

Receiving 23.

2-chloro-6-(1-tetrahydropyran-4-ylpyrazol-4-yl)-4-(trifluoromethyl)pyridine-3-carbonitrile.

The title compound is prepared from 1-tetrahydropyran-4-yl-4-(4,4,5,5-tetramethyl-1,3,2dioxaborolan-2-yl)pyrazole using essentially the same procedure as in the preparation 22.

ER/MS (m/z): 357, 359 (M+H).

Receiving 24.

Tert-butyl-3-[4-[5-cyano-6-[(28)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1yl]azetidin-1-carboxylate.

Tert-butyl-3-[4-[6-chloro-5-cyano-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl]azetidine-1-carboxylate (603 mg, 1.410 mmol) was dissolved in THF ( 18 ml). [(1R,4S)-7,7-dimethyl-2-oxonorbornan1-yl]methanesulfonate (2S)-2-methylazetidin-1-ium salt (519 mg, 1.711 mmol) and DIPEA (0.75 ml, 4. 3 mmol),

- 23 043635 the mixture was then heated at 130°C for 2.5 hours. The solvent was evaporated and the residue was purified by silica gel chromatography using a gradient of 0 to 50% EtOAc/hexanes to give the title compound (498 mg, 76% ) in the form of a colorless oily substance.

ER/MS (m/z): 463 (M+H), 461 (M-H).

Receiving 25.

Tert-butyl-N-[2-[4-[4-[5-cyano-6-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-

-yl]-1-piperidyl]-2-oxoethyl]carbamate.

\ / ¹¹¹

N 7 ^F F U' ^N

I C-n

2-[(2S)-2-methylazetidin-1-yl]-6-[1-(4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridine-3carbonitrile (0.129 g, 0.330 mmol) was dissolved in DXM (1 ml). 2-(tert-butoxycarbonylamino)acetic acid (69 mg, 0.39 mmol), HATU (0.175 g, 0.460 mmol) and DIPEA (0.18 ml, 1.0 mmol) were added. The mixture was stirred at RT for 3 hours, then diluted with saturated aqueous sodium bicarbonate and extracted twice with DCM. The organic layers were combined, dried over sodium sulfate, then filtered and evaporated. The residue was purified by reverse phase chromatography on C18 silica (solvent A=10 mM aqueous ammonium bicarbonate, solvent B=ACN; gradient 10 to 91% B) to give the title compound (129 mg, 71%) .

ER/MS (m/z): 548 (M+H).

Receiving 26.

1-(2,2-dimethyl-[1,3]dioxolan-4-ylmethyl)-4-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-1Hpyrazole.

4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (0.500 g, 2.50 mmol) was suspended in DMF (10 ml), then 4- (chloromethyl)-2,2-dimethyl-1,3-dioxolane (0.73 ml, 5.0 mmol) and cesium carbonate (1.64 g, 5.03 mmol). The mixture was heated at 75°C overnight. The reaction mixture was diluted with saturated aqueous sodium bicarbonate and extracted with EtOAc. The organic layers were combined and washed four times with saturated aqueous NaCl, then dried over sodium sulfate, filtered and evaporated to give the title compound (780 mg), which was used without further purification.

ER/MS (m/z): 309 (M+H).

Receiving 27.

2-chloro-6-[1-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]pyrazol-4-yl]-4-(trifluoromethyl)pyridine-3carbonitrile.

3-Cyano-2,6-dichloro-4-(trifluoromethyl)pyridine (0.352 g, 1.46 mmol) was dissolved in 1,4-dioxane (7.3 ml). 1-(2,2-dimethyl-[1,3]dioxolan-4-ylmethyl)-4-(4,4,5,5-tetramethyl[1,3,2]dioxaborolan2-yl)-1H-nirazole ( 0.602 g, 1.89 mmol), tetrakis(triphenylphosphine) palladium (0) (0.715 g, 0.619 mmol) and aqueous 2 M sodium carbonate (2.8 ml, 5.6 mmol). The mixture was purged with nitrogen for 15 minutes, then heated to 85°C for 1.5 hours. The reaction mixture was diluted with saturated aqueous sodium bicarbonate and extracted twice with EtOAc. The organic layers were combined and dried over sodium sulfate, then filtered and evaporated. The residue was purified by silica gel chromatography using a gradient of 0 to 40% EtOAc/hexanes to give the title compound (207 mg, 36%) as a colorless oil.

- 24 043635

ER/MS (m/z): 387, 389 (M+H).

Receiving 28.

Tert-butyl 4-[6-chloro-5-cyano-4-(trifluoromethyl)-2-pyridyl]pyrazole-1-carboxylate.

3-cyano-2,6-dichloro-4-(trifluoromethyl)pyridine (195 mg, 0.785 mmol), tert-butyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane- 2-yl)pyrazole-1-carboxylate (250 mg, 0.85 mmol), 2 M aqueous sodium carbonate (2.5 ml, 5.0 mmol) and 1,4-dioxane (4 ml). The reaction mixture was degassed at RT by bubbling nitrogen through the reaction mixture while stirring for 5 min. [1,1'bis(diphenylphosphino)ferrocene]dichloropalladium(II) (185 mg, 0.240 mmol) was added and the reaction mixture was heated at 100°C for 2 hours. Water was added and extracted with EtOAc. The extracts were combined and concentrated in vacuo. The residue was purified by silica gel chromatography using a gradient of 25 to 50% EtOAc/hexanes to give the title compound (40 mg, 14%) as an orange solid.

ER/MS (m/z): 271, 273 (M-H-BOC).

Receipt 29.

Tert-butyl-N -[2-[[2-[4-[2-[(2 S)-2-methylazetidin-1 -yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol1-yl]acetyl ]amino]ethyl]carbamate.

Added to the test tube

2-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4 yl]pyrazol-1-yl]acetic acid (200 mg, 0.586 mmol), DMF ( 2 ml), DIPEA (0.31 ml, 1.76 mmol), HATU (0.267 g, 0.703 mmol) and N-BOC-ethylenediamine (0.102 ml, 0.644 mmol). Stirred for 8 hours at RT, then purified by preparative HPLC (parameters: solvents - 10 mM aqueous ammonium bicarbonate, pH 10/5% MeOH (solvent A) and ACN (solvent B); pre-column - Waters BEH HILIC 100x30 mm, 5 µm , 110 A with 15x30 mm BEH HILIC guard column; column: Phenomenex® Kinetex® EVO C18, 100x30 mm, 5 µm, 100 A with 10 mm EVO guard column using built-in heater at 50°C; gradient from 33 to 67% V ) to obtain the title compound (179 mg, 63%).

ER/MS (m/z): 484 (M+H).

The compounds given in table. 1 is prepared using essentially the same procedure as in Preparation 29 and the corresponding commercially available amine.

- 25 043635

Table 1

Receipt Number Name Structure ER/MS (m/z) (M+H) thirty tert-butyl-(28)-2methyl-4-[2-[4-[2-[(25)2-methylazetidin-1 yl]-6- (trifluoromethyl)pyrim idin-4-yl]pyrazol-1 yl]acetyl]piperazine 1-carboxylate *0* Y> ^F o ^N —\ y—N ^-0 ⁰ 524 31 tert-6util-(38)-3methyl-4-[2-[4-[2-[(25)2-methylazeti din-1 yl]-6- (trifluoromethyl)pyrim idin-4-yl]pyrazol-1 yl]acetyl]piperazine 1-carboxylate О , У °γ ^Η Ο ° 524 32 mpem-6yTnn-(2R,5S)2,5-dimethyl-4-[2-[4-[2[(25)-2- methylazetidin-1 -yl]- 6- (trifluoromethyl)pyrim idin-4-yl]pyrazol-1 yl]acetyl]piperazin- 1-carboxylate |Ύν F.YYL/\ ^f 7 Y\n ^F N—/ VN ^~-0 ⁰ 538 33 tertiary-butyl-2,2dimethyl-4-[2-[4-[2- [(25)-2- methylazeti din -1 -yl] 6- (trifluoromethyl)pyrim idin-4-yl]pyrazol-1 yl]acetyl]piperazine 1-carboxylate N N<^N ^f YY\n ^{F n} ~5 -An ' Y-0 ⁰ 538

- 26 043635

Receiving 34.

Tert-butyl-(3R)-3-[4-[2-[(2S,3R)-3-hydroxy-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1-yl ]pyrrolidine-1-carboxylate.

HE

In a reaction vessel, combine 2,4-dichloro-6-(trifluoromethyl)pyrimidine (600 mg, 2.71 mmol) with (R)-tert-butyl-3-(4-(4,4,5,5-tetramethyl- 1,3,2-dioxaborolan-2-yl)-1 H-pyrazol-1-yl)pyrrolidin-1 carboxylate (1 g, 2.67 mmol), 2 M aqueous sodium carbonate (3 ml, 6 mmol) and bis (triphenylphosphine)palladium(II) dichloride (50 mg, 0.068 mmol) in 1,4-dioxane (6 ml). The mixture was degassed with nitrogen and heated at 80°C for 2 hours. Diluted with EtOAc (75 ml), washed with water and saturated aqueous NaCl. The organic layer was dried over sodium sulfate, filtered and concentrated. The residue was combined in a test tube with (2S,3R)-2-methylazetidin-3-ol [(1R,4S)-7,7-dimethyl-2oxonorbornan-1-yl]methanesulfonic acid (420 mg, 1.32 mmol) in 1 ,4-dioxane (4 ml) and DIPEA (0.7 ml, 4 mmol). The vessel was sealed and heated at 120°C in a microwave reactor for 1 hour. The reaction mixture was applied directly to silica gel and purified by silica gel chromatography using a gradient of 0 to 80% EtOAc/hexanes to give the title compound (570 mg, 92 %) in the form of a whitish foamy substance.

ER/MS (m/z): 469 [M+H].

Getting 35.

Tert-butyl-(3S)-3-[4-[2-[(2S,3R)-3-hydroxy-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1-yl ]pyrrolidine-1-carboxylate.

^{OH <V>}

Q^O 0^

The title compound was prepared essentially as described in Preparation 34 using (S)-tert-butyl-3-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2- yl)-1H-πpyrazol-1-yl)πpyrrolidine-1carboxylate.

ER/MS (m/z): 469 [M+H].

Getting 36.

Tert-butyl-3-[(1R)-4-[2-[(2R)-2-(hydroxymethyl)azetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1-yl]pyrrolidin- 1-carboxylate.

- 27 043635

The title compound was prepared essentially as described in Preparation 34 using (R)-2-azetidinemethanol hydrochloride.

ER/MS (m/z): 469 [M+H].

Receiving 37.

Tert-butyl-4-[4-[5-cyano-6-[(2S)-2-methylazetidin-1-ul]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1yl]-3-hydroxypiperidin-1 -carboxylate.

4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (560 mg, 2.9 mmol), tert-butyl-7-oxa- 4-azabicyclo[4.1.0]heptane-4-carboxylate (580 mg, 2.9 mmol), DMF (10 ml) and cesium carbonate (1.7 g, 5.2 mmol). The mixture was heated at 80°C for 6 hours. The reaction mixture was diluted with water and extracted with EtOAc. The extracts were combined and dried over sodium sulfate, filtered and concentrated to give crude tert-butyl-3-hydroxy-4-[4-(4,4,5,5-tetramethyl-1,3,2dioxaborolan-2-yl)pyrazol- 1-yl]piperidine-1-carboxylate (650 mg). This material was placed in the FDC and 2,6-dichloro-4-(trifluoromethyl)pyridine-3-carbonitrile (300 mg, 1.2 mmol), 1,4-dioxane (5 ml) and 2 M aqueous sodium carbonate (1 .3 ml, 2.6 mmol). The reaction mixture was degassed at RT by bubbling nitrogen through the reaction mixture while stirring for 5 min. Tetrakis(triphenylphosphine)palladium(0) (80 mg, 0.06 mmol) was added and degassed for an additional 3 min. The reaction mixture was heated at 80°C for 4 h, then cooled to RT. DIPEA (0.6 mL, 3 mmol) and [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonate (2S)-2-methylazetidinium 1-ium salt (410 mg, 1.0 mmol) were added. 4 mmol). Stir for 30 minutes at RT and then heat at 80°C for 1 hour. The mixture is diluted with water and extracted with EtOAc. The extracts were combined and dried over sodium sulfate, filtered and concentrated. The residue was purified by reverse phase chromatography (C18, gradient 10 to 100% ACN/10 mM aqueous ammonium carbonate+5% methanol) to give the title compound (95 mg, 15%).

ER/MS (m/z): 507 (M+H).

Example 1.

2-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl]-1-piperazin1-ylethanone.

tert-butyl-4-[2-[4-[2-[(2S)-2-methylazetidin-1-yl]-6(trifluoromethyl)pyrimidin-4-yl]pyrazol-1-yl was loaded into a 2 L FDC ]acetyl]piperazine-1-carboxylate (47.0 g, 89.5 mmol) and DCM (470 ml) was added. TFA (60 mL, 778 mmol) was added dropwise to the mixture and stirred at RT for 19 hours. Additional TFA (8 mL, 110 mmol) was added and stirring continued for 18 hours. The reaction mixture was slowly added to the flask containing cooled aqueous NH ₄ OH (35 wt.%, 150 ml, 1300 mmol). The layers were separated. The organic layer was washed with saturated aqueous NaCl, dried over Na ₂ SO ₄ , filtered and concentrated. The residue was dissolved in EtOAc (400 ml) and concentrated in vacuo. The residue was dissolved in IPA (400 ml) and concentrated in vacuo to give the title compound as an off-white foam (32 g, 84%).

ER/MS (m/z): 410 (M+H).

- 28 043635

Example 1a.

2-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)nurimidin-4-yl]pyrazol-1-yl]-1-piperazin1-ylethanone sesquisuccinate.

2-[4-[2-[(28)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol1-yl]-1-piperazin-1-yl-ethanone was added to the reactor (959 g, 2.34 mol) followed by IPA (11.5 L). Succinic acid (550 g, 4.69 mol) was added and the mixture was heated to 70-80°C to obtain a solution. The mixture was stirred at 70 to 80°C for 2 hours, then cooled to 25°C for 6 hours. The mixture was filtered and washed with IPA (1 L). The resulting solids were dried at 40-50°C for 6 hours to obtain the title compound (1070 g, 78%) as a white solid.

^1H NMR (400 MHz, DMSO-d6): δ 8.41 (s, 1H), 8.18 (s, 1H), 7.33 (s, 1H), 5.24 (s, 2H), 4 .53-4.43 (m, 1H), 4.06-3.93 (m, 2H), 3.58-3.47 (m, 4H), 2.95-2.82 (m, 4H) , 2.47-2.40 (m, 1H), 2.37 (s, 6H - succinate methylene groups, 1.5 eq.), 2.02-1.92 (m, 1H), 1.50 ( d, J=4 Hz, 3H).

High resolution ER-MS (m/z): theoretical 410.1911 (free base M+H), observed 410.1916.

The RID peaks from example 1a are given in table. 2.

table 2

Powder X-ray diffraction peaks of Example 1a

Peak Angle (°2-theta) Relative intensity (% of the most intense peak) 1 12.1 27.5% 2 14.1 22.7% 3 16.3 33.3% 4 17.7 30.5% 5 18.9 100.0% 6 20.0 22.1% 7 20.5 53.8% 8 21.3 44.9% 9 22.0 36.6% 10 24.3 23.2%

Example 2.

[(2R)-1 -[4-[1 -(4-piperidyl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]azetidin-2-yl]methanol.

Tert-butyl 4-[4-[2-[(2R)-2-(hydroxymethyl)azetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol1-yl]piperidin-1-carboxylate ( 2.55 g, 5.28 mmol) was dissolved in DCM (50 ml) and TFA (10 ml, 132 mmol) was added. The mixture was stirred at RT for 30 min, then applied directly to the MCO resin. The resin was washed with MeOH and then with ammonia in MeOH solution (7 M). Fractions containing the desired product were combined and concentrated in vacuo. The residue was purified by silica gel chromatography using a 0 to 9% (7 M ammonia/MeOH)/DCM gradient to give the title compound as a white powder (2.02 g, 94%).

ER/MS (m/z): 383 (M+H); 381 (M-H).

- 29 043635

Example 3.

[(2R)-1-[4-[1-(azetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]azetidin-2yl]methanol.

Tert-butyl-3-[4-[2-[(2R)-2-(hydroxymethyl)azetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol1-yl]azetidin-1-carboxylate ( 153 mg, 0.336 mmol) was dissolved in DCM (15 ml) and TFA (5 ml, 66 mmol) was added. The mixture was stirred at RT for 1 hour, then concentrated under reduced pressure at RT. The residue was evaporated together with DCM and dried in vacuo. The residue was purified by preparative HPLC (parameters: solvent A=10 mM aqueous ammonium bicarbonate, pH 10/5% MeOH, solvent B=ACN; column - Phenomenex® Kinetex® EVO C18, 100x30 mm, 5 µm, 100 A with a guard column EVO 15x30 mm) to obtain the title compound (67 mg, 40%) as a white powder.

ER/MS (m/z): 355 (M+H); 353 (M-H).

Example 4.

(2S,3R)-1-[4-[1-(azetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]-2-methylazetidin3-ol.

Tert-butyl-3-[4-[2-[(2S,3R)-3-hydroxy-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1-yl]azetidin-1 β-carboxylate (153 mg, 0.336 mmol) was dissolved in DCM (15 ml) and TFA (5 ml, 66 mmol) was added. The mixture was stirred at RT for 1 hour, then concentrated under reduced pressure at RT. The residue was evaporated together with DCM and dried in vacuo. The residue was purified by preparative HPLC (parameters: solvent A = 10 mM aqueous NH4HCO3 with 5% MeOH, pH 10, solvent B = ACN; column - Phenomenex® Kinetex® EVO C18, 100x30 mm, 5 μm, 100 A with a guard column EVO 15x30 mm ) to obtain the title compound (79 mg, 57%) as a white powder.

ER/MS (m/z): 355 (M+H); 353 (M-H).

Example 5.

2-[4-[5-cyano-6-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl]-N-(2hydroxyethyl)acetamide.

2-[4-[5-cyano-6-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1-yl]acetic acid (351 mg, 0. 96 mmol) was dissolved in DMF (2 ml) and HATU (482 mg, 1.24 mmol), ethanolamine (0.1 ml, 2 mmol) and DIPEA (0.5 ml, 3 mmol) were added. The mixture was stirred at RT for 2 d. The reaction mixture was immediately purified by reverse phase chromatography (C18 bound silica) using a 5 to 95% MeCN/aqueous (NH ₄ ) ₂ CO ₃ (10 mM) gradient to give indicated in the title

- 30 043635 compounds (110 mg, 28%).

ER/MS (m/z): 409 (M+H); 407 (M-H).

Example 6.

2-[(2R)-2-(hydroxymethyl)αzetidin-1-yl]-6-[1-(4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridin3-carbonitrile.

Tert-butyl-4-[4-[5-cyano-6-[(2R)-2-(hydroxymethyl)azetidin-1-yl]-4-(trifluoromethyl)-2pyridyl]pyrazol-1-yl]piperidin-1 β-carboxylate (264 mg, 0.52 mmol) was dissolved in DCM (2 ml). TFA (2 ml) was added to the reaction mixture in one portion and stirred for 1 hour. The reaction mixture was concentrated, diluted with DCM, and the reaction mixture was concentrated. The reaction material was diluted with MeOH (1 ml) and saturated aqueous NaHCO ₃ was added to obtain a pH of ~8. The reaction mixture was purified by reverse phase chromatography (40 g, C18, gradient 20-100% ACN/10 mM aqueous (NH ₄ ) ₂ CO ₃ with 5% MeOH) to give the title compound (135.5 mg, 64 %).

ER/MS (m/z): 407 (M+H).

Example 7.

2-[(2S,3R)-3-hydroxy-2-methylαzetidin-1-yl]-6-[1-(4-piperidyl)pyrazol-4-yl]-4(trifluoromethyl)pyridin-3-carbonitrile.

He

Tert-butyl-4-[4-[5-cyano-6-[(2S,3R)-3-hydroxy-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2pyridyl]pyrazol-1-yl] piperidine-1-carboxylate (220 mg, 0.43 mmol) was dissolved in DCM (2 ml). TFA (2 ml) was added to the reaction mixture in one portion and stirred for 1 hour. The reaction mixture was concentrated, diluted with DCM, and the reaction mixture was concentrated. The reaction material was diluted with MeOH (1 ml) and saturated aqueous NaHCO ₃ was added to obtain a pH of ~2. The reaction mixture was purified by reverse phase chromatography (40 g, C18, gradient 20-100% ACN/10 mM aqueous (NH ₄ ) ₂ CO ₃ with 5% MeOH) to give the title compound (127 mg, 72%) .

ER/MS (m/z): 407 (M+H).

Example 8.

2-[(2S)-2-methylazetidin-1-yl]-6-[1-(1-methyl-4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridine3-carbonitrile.

2-chloro-6-[1-(1-methyl-4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridine3-carbonitrile (261 mg, 0.68 mmol), salt [(1R ,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonate (2S)-2-methylazetidin-1-ium (338 mg, 1.1 mmol), DIPEA (0.4 ml, 2 mmol) and THF (3.7 ml). The tube was sealed and heated at 130°C for 2 hours. The reaction mixture was partitioned between DCM and 1 N aqueous HCl. The aqueous phase was neutralized with 1 H NaOH, then extracted three times with DCM. Organic

- 31 043635 Chinese extracts were mixed. Dried over sodium sulfate, filtered and concentrated in vacuo. The material was dried at 50°C in a vacuum oven for 1 hour. The residue was purified by chromatography on silica gel using a gradient of 5 to 10% MeOH/DCM, fractions containing the title compound were combined, concentrated and dried in a vacuum oven at 50 °C overnight. The residue was purified by preparative HPLC (parameters: solvent A = 10 mM aqueous ammonium bicarbonate with 5% MeOH, solvent B = ACN; column - Xbridge™ 30x75 mm 5 μm, 45 ml/min; gradient from 5 to 100% B) to obtain the indicated in the title compound (73 mg, 26%).

ER/MS (m/z): 405 (M+H).

Example 9.

6-[1-[1-(2-hydroxyethyl)-4-piperidyl]pyrazol-4-yl]-2-[(2S)-2-methylazetidin-1-yl]-4(trifluoromethyl)pyridine-3-carbonitrile .

HE

2-chloro-6-[1-(4-piperidyl)pyrazol-4-yl]-4(trifluoromethyl)pyridin-3-carbonitrile hydrochloride (304.3 mg, 0.63 mmol) was added to a 20 ml test tube, 2 -[tert-butyl(dimethyl)silyl]oxyacetaldehyde (219 mg, 1.19 mmol), DIPEA (0.52 ml, 3.1 mmol), sodium triacetoxyborohydride (277.2 mg, 1.3 mmol) and DCM (2, 0 ml). The reaction mixture was stirred at RT for 2 hours. 2-[tert-Butyl(dimethyl)silyl]oxyacetaldehyde (219 mg, 1.2 mmol) was added and stirred overnight at RT. Saturated aqueous sodium bicarbonate solution was added and extracted twice with DCM. The organic phases were combined, washed with saturated aqueous NaCl, dried over sodium sulfate, filtered and concentrated in vacuo. The residue was purified by silica gel chromatography using a gradient of 40 to 70% EtOAc in hexanes to give 6-[1-[1-[2-[tert-butyl(dimethyl)silyl]oxyethyl]-4-piperidyl]pyrazol-4-yl] -2-chloro-4-(trifluoromethyl)pyridine-3carbonitrile (220 mg, 58%).

6-[1-[1-[2-[tert-Butyl(dimethyl)silyl]oxyethyl]-4-piperidyl]pyrazol-4-yl]2-chloro-4-(trifluoromethyl)pyridine-3-carbonitrile was added to the test tube (175 mg, 0.289 mmol), [(1R,4S)-7,7-dimethyl-2oxonorbornan-1-yl]methanesulfonate (2S)-2-methylazetidin-1-ium salt (136 mg, 0.45 mmol), DIPEA (0.15 ml, 0.85 mmol) and THF (1.5 ml, 18 mmol). The vessel was sealed and heated at 130°C for 60 minutes. The reaction mixture was partitioned between EtOAc and 1 N HCl, the organic layer was removed and the aqueous layer was extracted three times with EtOAc. The organic layers were combined, washed with saturated aqueous NaCl, dried over sodium sulfate, filtered and concentrated in vacuo. The residue was dissolved in THF (1.3 ml) and the mixture was cooled to 0°C, then 1 M tetrabutylammonium fluoride in THF (0.37 ml, 0.37 mmol) was added dropwise. The reaction mixture was allowed to warm to RT and stirred overnight. The reaction mixture was concentrated in vacuo. The residue was purified by silica gel chromatography using 100% hexanes, then using a 1 to 10% (0.7 N ammonia in MeOH)/DCM gradient to give the title compound (93 mg, 73%).

ER/MS (m/z): 435 (M+H).

Example 10.

6-[1-[2-(dimethylamino)ethyl]pyrazol-4-yl]-2-[(28)-2-methylazetidin-1-yl]-4(trifluoromethyl)pyridin-3-carbonitrile.

2,6-Dichloro-4-(trifluoromethyl)pyridine-3-carbonitrile (250 mg, 1.04 mmol), N,N-dimethyl-(2-(4-(4,4,5,5 -tetramethyl)-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl)ethanamine (271 mg, 1.07 mmol), 1,4-dioxane (10 ml), aqueous potassium carbonate ( 3 M, 1.04 ml 3.12 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (II)) (40 mg, 0.052 mmol). The vessel was sealed and heated at 80°C for 2 hours, then the mixture was cooled to RT. Filtered through 3 g cartridge

- 32 043635

Celite® using EtOAc for elution. The filtrate was concentrated by evaporation. [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonate (28)-2-methylazetidin-1-ium salt (378 mg, 1.24 mmol), DIPEA (0 .9 ml, 5.2 mmol) and DMSO (10 ml). The mixture was heated at 120°C for 6 hours. The reaction mixture was diluted with water and loaded into an 8trata-XL® cartridge (8 g; pre-washed with MeOH, dried and then washed with water). The cartridge was washed with water and then with a 1:1 mixture of MeOH/water and the eluents were removed. The product was eluted with MeOH, then DCM and finally with DCM/MeOH 1:1, combining the eluted fractions. The combined fractions containing the title compound were concentrated. The residue was purified by preparative HPLC (parameters: solvent A = 10 mM aqueous ammonium bicarbonate with 5% MeOH pH 10, solvent B = ACN; precolumn - Waters BEH HILIC 100x30 mm, 5 μm, 110 Ac with a BEH HILIC 15x30 mm guard column; Phenomenex® column Kinetex® EVO C18, 100x30 mm, 5 µm, 100 A with 15x30 mm EVO guard column using built-in heater at 50°C; gradient 33 to 100% B) to obtain the title compound (53 mg, 13%).

ER/MS (m/z): 379 (M+H).

Example 11.

6-[1-(2-hydroxyethyl)pyrazol-4-yl]-2-[(28)-2-methylazetidin-1-yl]-4-(trifluoromethyl)pyridin-3carbonitrile.

The title compound was prepared essentially using the method described in Example 10 with 2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol- 1-yl)ethanol.

ER/MS (m/z): 352 (M+H).

Example 12.

2-[(28)-2-methylazetidin-1-yl]-6-[1-(4-piperidyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridin-3carbonitrile.

Tert-butyl 4-[4-[5-cyano-6-[(28)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1yl]piperidine-1-carboxylate ( 1.99 g, 4.06 mmol) was dissolved in DCM (50 ml) and TFA (10 ml, 132.3 mmol) was added slowly. Stir at RT for 30 min, then the reaction mixture is loaded into four 10 g OSD cartridges. The cartridges were washed with MeOH and then 7 H ammonia in MeOH. The main washing waters were concentrated in vacuo. The residue was purified by reverse phase chromatography on C18 coupled to silica (solvent A: 10 mM ammonium bicarbonate with 5% MeOH; solvent B: ACN; gradient: 10-71% solvent B) to give the title compound (709 mg, 45%) in the form of a white solid.

ER/MS (m/z): 391 (M+H).

Example 13.

2-[(28)-2-methylazetidin-1-yl]-4-[1-(4-piperidyl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidine.

Tert-butyl 4-[4-[2-[(28)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1yl]piperidine-1-carboxylate (0.213 g, 0.457 mmol) was dissolved in DCM (5 ml) and TFA (1 ml) was added. The mixture was stirred for 25 min, then the reaction mixture was loaded directly into a 10 g OSD cartridge. The cartridge was washed with MeOH and then eluted with 7 H ammonia in MeOH. Basic

- 33 043635 the washings were concentrated to obtain the title compound (129 mg, 77%).

ER/MS (m/z): 367 (M+H).

Examples in table. 3 was prepared using essentially the method described in Example 13 and the corresponding protected amine.

Table 3

Example number Name Structure ER/MS (m/z): (M+H) 14 (2X,3R)-2-methyl-1 - [4-[1-[(3R)pyrrolidin-3-yl]pyrazol-4-yl] 6-(trifluoromethyl)pyrimidin-2yl]azetidin-3-ol OH ^< V ^> N^N ^F T A ^ΝΗ 369 15 (25,3)-2-methyl-1-[4-[1-[(35)- pyrrolidin-3 -yl]pyrazol-4-yl] 6-(trifluoromethyl)pyrimidin-2yl]azetidin-3-ol OH Λ..... Ν Ν^Ν fTTA^x fY ta ^F Ν ^C-NH 369 16 [(27?)-1 - [4- [ 1 - [(ZL)-pyrrolidin -3 yl]pyrazol-4-yl]-6- (trifluoromethyl)pyrimidine-2- yl]azetidin-2-yl]methanol _Λ he ...../ N N<A _n f^aaL^x fY ^? ^N.H. 369

Example 17.

6-[1-(αzetidin-3-yl)pyrazol-4-yl]-2-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)pyridin-3carbonitrile.

The title compound was prepared essentially using the same procedure as Example 12, starting with Ίret-butyl-3-[4-[5-cyano-6-[(2S)-2-methylαzetidin-1-yl] -4-(Ίriforomethyl)-2pyridyl]pyrazol-1-yl]azetidine-1-carboxylate. Purified by preparative HPLC (parameters: solvent A - 10 mM aqueous ammonium bicarbonate with 5% MeOH, solvent B - ACN; gradient from 35 to 59% B; column - Phenomenex® Kinetex® EVO C18, 100x30 mm, 5 μm) to obtain the indicated in the connection header.

ER/MS (m/z): 363 (M+H).

- 34 043635

Example 18.

6-[1-[1 -(2-Aminoacetyl)-4-piperidyl]pyrazol-4-yl]-2-[(2S)-2-methylazetudin-1 -yl]-4(trifluoromethyl)pyridin-3-carbonitrile .

The title compound was prepared using essentially the same procedure as Example 12, starting with tert-butyl-N-[2-[4-[4-[5-cyano-6-[(2S)-2- methylαzetudin-1-yl]-4-(trifluoromethyl)-2pyridyl]pyrazol-1-yl]-1-piperidyl]-2-oxoethyl]carbamate. Purification was done by preparative HPLC (parameters: solvent A - 10 mM ammonium bicarbonate with 5% MeOH; solvent B - MeOH; gradient from 20 to 50% B, 55 ml/min; column - Phenomenex® Kinetex® EVO C18 30 mmx250 mm, 5 μm ).

ER/MS (m/z): 448 (M+H).

Example 19.

6-[1-(1-acetyl-4-piperidyl)pyrazol-4-yl]-2-[(2S)-2-methylazetidin-1 -yl]-4(trifluoromethyl)pyridin-3-carbonitrile.

2-[(2S)-2-methylazetidin-1-yl]-6-[1-(4-nuperidyl)nyrazol-4-yl]-4-(trifluoromethyl)nirudin-3carbonitrile (Example 12) (0.111 g, 0.284 mmol) was dissolved in DCM (1 ml) and pyridine (0.026 ml, 0.32 mmol) was added. The mixture was cooled to 0°C and acetyl chloride (0.024 ml, 0.34 mmol) was added. The mixture was heated to RT and stirred for 30 min. The mixture was cooled to 0°C and pyridine (0.026 ml, 0.32 mmol) and acetyl chloride (0.024 ml, 0.34 mmol) were added. Heated to RT and stirred for another 30 minutes. The reaction mixture was diluted with saturated aqueous sodium bicarbonate and extracted twice with DCM. The organic layers were combined, dried over sodium sulfate, filtered and evaporated. The residue was purified by preparative HPLC (parameters: solvent A - 10 mM ammonium bicarbonate with 5% MeOH; solvent B - MeOH; gradient 25-55%, 55 ml/min; column - Phenomenex® Kinetex® EVO C18 30x250 mm, 5 μm) for obtaining the title compound (67 mg, 55%).

ER/MS (m/z): 433 (M+H).

Examples 20-23.

Examples in table. 4 was prepared using essentially the procedure of Example 5, a suitable carboxylic acid and a commercially available amine.

- 35 043635

Table 4

Example number Name Structure ER/MS (m/z): (M+H) 20 2-[(25)-2-methylazetidin-1-yl]-6-[1 [2-(4-methylpiperazin-1-yl)-2oxoethyl]pyrazol-4-yl]-4(trifluoromethyl)pyridin- 3 carbonitrile X / ^{1 11} N 1\К 1 fVVY^ О С- N\ 448 21 2-[4-[5-cyano-6-[(25)-2methylazetidin-1 -yl]-4(trifluoromethyl)-2- pyridyl]pyrazol-1-yl]-M,M-bis(2hydroxyethyl)acetamide ΞΕ O ] O z- Z LL LL 453 22 6-[1-[2-[(ZYA,4YA)-3,4- dihydroxypyrrolidin-1 -yl] -2oxoethyl] pyrazol-4-yl] -2- [(2S)-2methylazetidin-1 -yl] -4- (trifluoromethyl)pyridin-3 carbonitrile Ϋ..... ^F Wh HO 451 23 6-[1-[2-[(35.45)-3.4- dihydroxypyrrolidin-1 -yl] -2oxoethyl] pyrazol-4-yl] -2- [(25)-2 - methylazetidin-1 -yl] -4- (trifluoromethyl)pyridin-3 carbonitrile mV/th$ 6 I 451

Example 24.

6-[1-(2,3-dihydroxypropyl)pyrazol-4-yl]-2-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)pyridin3-carbonitrile.

2-chloro-6-[1-[(2,2-dimethyl-1,3-dioxolan-4-yl)methyl]pyrazol-4-yl]-4-(trifluoromethyl)pyridine-3carbonitrile (0.204 g 0.527 mmol) dissolved in THF (6 ml). Aqueous hydrochloric acid (2 M, 3 mL, 6 mmol) was added to the mixture and stirred at RT for 1.5 hours. The mixture was diluted with saturated aqueous sodium bicarbonate and extracted twice with EtOAc. The organic layers were combined and dried over sodium sulfate, then filtered and evaporated to give crude 2-chloro-6-[1-(2,3dihydroxypropyl)pyrazol-4-yl]-4-(trifluoromethyl)pyridine-3-carbonitrile (183 mg ).

- 36 043635

The crude material was completely dissolved in DMF (2 ml), then [(1R,4S)-7,7dimethyl-2-oxonorbornan-1-yl]methanesulfonate (2S)-2-methylazetidin-1-ium salt (222 mg, 0 .73 mmol) and DIPEA (0.37 ml, 2.1 mmol). The mixture was heated in a microwave reactor at 130°C for 2.5 hours. The reaction mixture was purified by preparative HPLC (parameters: solvent A - 10 mM ammonium bicarbonate with 5% MeOH; solvent B - ACN; gradient from 20 to 50%, 60 ml /min; column - Phenomenex® Kinetex® EVO C18 30x250 mm, 5 µm) to obtain the title compound (102 mg, 51%).

ER/MS (m/z): 382 (M+H).

Examples 24a and 24b.

Example 24a.

6-[1-(2,3-dihydroxypropyl)nazol-4-yl]-2-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)pyridin3-carbonitrile - isomer 1.

Example 24b.

6-[1-(2,3-dihydroxypropyl)nirazol-4-ul]-2-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)pyridine3-carbonitrile - isomer 2.

Isomers of 6-[1 -(2,3-dihydroxypropyl)pyrazol-4-yl]-2-[(2S)-2-methylazetidin-1 -yl]-4(trifluoromethyl)pyridin-3-carbonitrile (86 mg) were separated using chiral SLC (parameters: column - Phenomenex® Lux® Cellulose-2, 21x250 mm; column temperature - 40°C; solvent 15% EtOH/CO ₂ , 80 ml/min) to obtain the title compounds (isomer 1, first eluting isomer: 35 mg, ER/MS (m/z): 382 (M+H); isomer 2, second eluting isomer: 39 mg, ER/MS (m/z): 382 (M+H) ). Analytical chiral SLC (parameters: column - Phenomenex® Lux® Cellulose2, 4.6x150 mm; solvent - 15% EtOH/CO ₂ , 5 ml/min): isomer 1 - retention time 3.10 min, 95.6% ee; isomer 2 - retention time 3.52 min - 94.4% ee.

Example 25.

2-[(2S)-2-methylazetidin-1-yl]-6-(1H-pyrazol-4-yl)-4-(trifluoromethyl)nuridin-3-carbonitrile.

Combine tert-butyl 4-[6-chloro-5-cyano-4-(trifluoromethyl)-2-pyridyl]pyrazole-1carboxylate (75 mg, 0.20 mmol), 1,4-dioxane (1.5 ml) and EtOH (1 ml). DIPEA (130 mg, 1.0 mmol) and [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonate (2S)-2-methylazetidin-1ium salt (117 mg, 0.0 mmol) were added. 39 mmol). The reaction mixture was heated in a microwave reactor at 200°C for 2 hours. The reaction mixture was concentrated in vacuo. The residue was purified by reverse phase chromatography on C18 silica using a gradient of 10 to 100% ACN/water with 0.1% formic acid to give the title compound (35 mg, 41%) as a pale solid -yellow color.

ER/MS (m/z): 308 (M+H), 306 (M-H).

Example 26.

2-[(2S)-2-methylazetidin-1-yl]-6-(1-tetrahydropyran-4-ylpyrazol-4-yl)-4-(trifluoromethyl)pyridin3-carbonitrile.

Combine 2-chloro-6-(1-tetrahydropyran-4-ylpyrazol-4-yl)-4-(trifluoromethyl)pyridine3-carbonitrile (105 mg, 0.29 mmol), 1,4-dioxane (1 ml) in a test tube , EtOH (0.5 ml), DIPEA (0.2 ml, 1 mmol) and [(1R,4S)-7,7-dimethyl-2-oxonorbornan-1-yl]methanesulfonate salt (28)-2-methylazetidine -1-ia (100 mg, 0.33 mmol). The reaction mixture was heated in a microwave reactor at 150°C for 2 hours. The reaction mixture was concentrated in vacuo. The residue was purified by silica gel chromatography using a gradient of 40 to 50% EtOAc/hexanes to give the title compound (88 mg, 76%) as a light yellow solid.

ER/MS (m/z): 392 (M+H).

- 37 043635

Example 27.

6-[1 -(-3-hydroxy-4-piperudyl)pyrazol-4-yl]-2-[(2S)-2-methylazetudin-1 -yl]-4-(trifluoromethyl)pyridine3-carbonitrile.

Tert-butyl-4-[4-[5-cyano-6-[(2S)-2-methylazetidin-1-yl]-4-(trifluoromethyl)-2-pyridyl]pyrazol-1yl]-3-hydroxypiperidin-1 β-carboxylate (85 mg, 0.17 mmol) was dissolved in DCM (1 ml), then TFA (0.5 ml) was added and stirred for 5 min. The reaction mixture was concentrated in vacuo. The residue was purified by reverse phase chromatography (C18, gradient 20 to 100% ACN/10 mM aqueous ammonium carbonate+5% methanol) to obtain the title compound (27 mg, 40%) as a white solid.

ER/MS (m/z): 407 (M+H).

Example 28.

N-(2-aminoethyl)-2-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1yl]acetamide.

Tert-butyl-N-[2-[[2-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol1-yl]acetyl] amino]ethyl]carbamate (340 mg) was dissolved in pure TFA (5 ml) and stirred at ambient temperature. After 2 min, quench with aqueous NaOH to an alkaline state. Extracted with DCM and EtOAc. The extracts were combined, dried over sodium sulfate, filtered and concentrated. The residue was purified by preparative HPLC (parameters: solvents - 10 mM aqueous ammonium bicarbonate, pH 10/5% MeOH (solvent A) and ACN (solvent B); pre-column - Waters BEH HILIC 100x30 mm, 5 µm, 110 A with a BEH HILIC guard column 15x30 mm; column - Phenomenex® Kinetex® EVO C18, 100x30 mm, 5 µm, 100 A with EVO 15x30 mm guard column using flow heater at 50°C; gradient from 14 to 48% B) to obtain the title compound ( 45 mg, 17%).

ER/MS (m/z): 384 (M+H).

The compounds given in table. 5 was prepared using essentially the same procedure as in Example 28 and the corresponding protected amine.

- 38 043635

Table 5

Example number Name Structure ER/MS (m/z): (M+H) 29 2-[4-[2-[(25)-2-methylazetidin-1 yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1 -yl]-1 - [(35)-3 methylpiperazin-1 -yl ]ethanone N<^N ^F 424 thirty 2-[4-[2-[(25)-2-methylazetidin-1 yl]-6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1 -yl]-1 - [(25)-2methylpiperazin-1 -yl] ethanon ^< V ^> N^N .....D '—N H 424 31 1-(3,3-dimethylpiperazin-1-yl)-2[4-[2-[(25)-2-methylazetidin-1 -yl] 6-(trifluoromethyl)pyrimidin-4yl]pyrazol-1 -yl]ethanone LL LL 438 32 1-[(2S,5R)-2,5-dimethylpiperazin-1 yl]-2-[4-[2-[(25)-2-methylazeti din- 1 -yl]-6-(trifluoromethyl)pyrimi din4-yl]pyrazol-1 -yl]ethanone ^< V ^> N^N fY T n ^F o c λ-N ''' H 438

Example 33.

4-[1-(azetidin-3-yl)pyrazol-4-yl]-2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidine.

Tert-butyl 3-[4-[2-[(2S)-2-methylazetidin-1-yl]-6-(trifluoromethyl)pyrimidin-4-yl]pyrazol-1yl]azetidin-1-carboxylate (530 mg, 1.21 mmol) was dissolved in DCM (50 ml). TFA (6 mL) was added and the mixture was stirred at RT for 1 hour. The mixture was concentrated in vacuo to give the crude title compound. Half of this material was purified using preparative HPLC (parameters: solvents - 10 mM aqueous ammonium bicarbonate, pH 10/5% MeOH (solvent A)

- 39 043635 and ACN (solvent B); pre-column - Waters BEH HILIC 100x30 mm, 5 µm, 110 A with protective column BEH HILIC 15x30 mm; column - Phenomenex® Kinetex® EVO C18, 100x30 mm, 5 µm, 100 A with EVO guard column 15x30 mm using a flow heater at 50°C; gradient from 23 to 58% B) to obtain the title compound (162 mg, 80% yield from half starting material) as a white solid.

ER/MS (m/z): 339 (M+H).

Example 34.

2-[(2S)-2-methylazetidin-1-yl]-4-[1-(1-methylazetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidine.

Half of the crude 4-[1-(azetidin-3-yl)pyrazol-4-yl]-2-[(2S)-2-methylazetidin-1-yl]-6(trifluoromethyl)pyrimidine (0.6 mmol) obtained in example 33, dissolved in MeOH (6 ml). Acetic acid (0.1 ml) and 3.45 M aqueous formaldehyde (1 ml) were added and the mixture was stirred at RT for 30 min. Sodium triacetoxyborohydride (300 mg, 1.4 mmol) was added and the mixture was stirred for 2 hours. The mixture was concentrated in vacuo and the residue was purified by preparative HPLC (solvents: 10 mM aqueous ammonium bicarbonate pH 10/5% MeOH (solvent A) and ACN (solvent B); pre-column - Waters BEH HILIC 100x30 mm, 5 µm, 110 A with guard column BEH HILIC 15x30 mm; column - Phenomenex® Kinetex® EVO C18, 100x30 mm, 5 µm, 100 A with guard column EVO 15x30 mm using built-in heater at 50°C; gradient 33-67% B) to obtain the title compound (128 mg, yield 61% of half starting material in Example 33) as a white solid.

ER/MS (m/z): 353 (M+H).

Example 35.

(2S,3R)-2-methyl-1-[4-[1-(1-methylazetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2yl]azetidin-3-ol.

The title compound was prepared using essentially the same procedure as Example 34, starting with (2S,3R)-1-[4-[1-(azetidin-3-yl)pyrazol-4-yl]-6 -(trifluoromethyl)pyrimidin-2-yl]-2methylazetidin-3-ol (example 4).

ER/MS (m/z): 369 (M+H), 367 (M-H).

Example 36.

[(2R)-1-[4-[1-(1-methylazetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]azetidin-2yl]methanol.

The title compound was prepared using essentially the same procedure as in

- 40 043635 least 34, starting with [(2R)-1-[4-[1-(azetidin-3-yl)pyrazol-4-yl]-6-(trifluoromethyl)pyrimidin-2-yl]azetidin2-yl] methanol (example 3).

ER/MS (m/z): 369 (M+H), 367 (M-H).

Methods of analysis.

KGC enzyme activity assay for human KGC-S and human KGC-A.

The true potency of inhibition of KGC C or A activity can be measured using an enzymatic assay that measures F1P production. Compounds were prepared in DMSO and tested using a 10-point concentration curve, yielding 3-fold serial dilutions of compounds in a 96-well plate ranging from 20 μM to 1.02 nM. The enzyme was prepared in assay buffer (50 mM 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (HEPES), 10 mM potassium chloride, 100 mM magnesium chloride, 2 mM tris(2-carboxyethyl)phosphine (TCEP), 0 .01% n-octyl glucoside) and incubated with the compounds at RT for 15 min. The reaction was carried out in 100 μl volumes containing substrate concentrations of fructose (250 μM for the KGC-S assay and 1.25 mM for the KGC-A assay) and ATP (150 μM for both isoforms), which were additionally incubated at RT for 20 min . The reaction was then stopped by adding a stopping buffer consisting of 0.2% formic acid and 1 μg/ml internal standard ^13C6 _- fructose-6-phosphate ( ^13C6 -F6P). The plates were stored at -20°C until RapidFire MS analysis.

RapidFire MS assay for F1P quantification.

An Agilent 300 RapidFire automated extraction system (Agilent, Santa Clara, CA) with three HPLC quaternary pumps was coupled to an Agilent 6495 triple quadrupole mass spectrometer (Agilent Technologies, Santa Clara, CA) equipped with an electrospray ionization (ESI) source. The RapidFire Mass Spec system is equipped with a reusable cartridge (G9205A) RapidFire C18 (Type C) for solid phase extraction (SPE).

Solvent A used for sample loading and washing is 6 mM octylamine (Acros Organics 129495000) adjusted to pH 5.0 with acetic acid. Solvent B used to elute the sample is 20% water in ACN containing 0.1% formic acid. Samples were analyzed sequentially by withdrawing 10 μL directly from the multiwell plates onto a collection loop under vacuum. 10 μL of sample was loaded into the C18 cartridge and washed using solvent A at a flow rate of 1.25 mL/min for 5000 ms. The retained analyte was eluted into the mass spectrometer using solvent B at a flow rate of 1.25 mL/min for 5000 ms. The system was re-equilibrated using solvent A at a flow rate of 1.25 ml/min for 2000 ms.

The triple quadrupole mass spectrometer was equipped with an ESI source, and the analytes were monitored using selected reaction monitoring (SRM) in negative [M-H]- mode. F1P was monitored at m/z 259.02/96.9 and ¹³ C6-fructose-6-phosphate was monitored at m/z 264.99/97. Area ratio values for F1P were calculated using ¹³ C ₆ -fructose-6-phosphate as an internal standard.

Compounds of Examples 1 to 36 were tested essentially as described above.

Table 6

Example number hKGK-S 1C ₅ o (nM) hCGK-A 1C ₅ o (nM) 1 20.7 ± 3.88 (n=4) 24.2 ± 2.9 (n=4) 2 134 ±43.1 (n=7) 98.2 ± 25.0 (n=7) 3 41.9 ±6.77 (n=3) 45.4 ±29.5 (n=4) 4 14.4 ± 1.12 (n=3) 11.4 ± 4.34 (n=4) 5 57.0 ± 32.6 (n=5) 37.7 ± 19.9 (n=5) 6 25.7 ± 5.14 (n=2) 15.4 ±5.05 (n=3) 7 18.1 ±4.85 (n=2) 6.02 ± 2.27 (n=3) 8 56.1 ± 62.5 (n=3) 25.2 ± 15.7 (n=3) 9 6.26 (n=1) 3.58 (n=1)

- 41 043635

10 418 ±503 (n=3) 246 ± 248 (n=3) And 8.03 (n=1) 5.23 (n=1) 12 3.87 ± 0.391 (n=4) 3.65 ± 0.817 (n=4) 13 15.5 ±20.4 (n=3) 7.69 ± 7.97 (n=3) 14 15.9 ± 4.37 (n=2) 19.1 ±2.98 (n=3) 15 27.2 ± 8.60 (n=2) 26.7 ±3.66 (n=3) 16 28.0 ±17.3 (n=2) 30.0 ± 5.09 (n=3) 17 16.1 ±7.96 (n=4) 13.2 ± 12.7 (n=4) 18 59.2 ± 67.8 (n=3) 35.6 ±32.4 (n=3) 19 54.3 (n=1) 29.2 (n=1) 20 150 ±243 (n=3) 81.8 ± 92.6 (n=3) 21 55.2 ±28.1 (n=3) 39.2 ± 13.6 (n=3) 22 61.3 ±57.5 (n=3) 33.7 ± 17.8 (n=3) 23 41.6 ±47.1 (n=3) 24.9 ± 15.3 (n=3) 24 7.91 (n=1) 7.29 (n=1) 24a 6.06 (n=1) 9.99 (n=1) 246 4.89 (n=1) 6.75 (n=1) 25 13.6 ±7.2 (n=4) 16.9 ±9.7 (n=4) 26 28.4 (n=1) 22.2 (n=1) 27 9.21 (n=1) 4.63 (n ⁼ 1) 28 68.0 (n=1) 51.0 (n=1) 29 38.6 (n=1) 103 (n=1) thirty 46.3 (n=1) 57.8 (n=1) 31 34.7 (n=1) 70.5 (n=1) 32 13.3 (n=1) 41.5 (n=1) 33 4.90 ±1.31 (n=2) 3.63 ± 1.13 (n=3) 34 3.71 ±0.178 (n=2) 5.41 ±6.13 (n=3) 35 2.63 (n=1) 1.59 (n=1) 36 10.0 (n=1) 3.98 (n=1)

Data are expressed as mean ± SEM (n).

The results presented in table. 6 above demonstrate that compounds of Examples 1 to 36 inhibit the enzymatic activity of both KGC-S and KGC-A.

Analysis of cellular activity of KGC.

Efficacy can be measured using a cell-based assay for inhibition of fructose conversion to F1P by cellular KGC. HepG2 cells were seeded into 96-well cell culture plates in growth medium (Dulbecco's modified Eagle's medium (DME) high glucose, 10% heat inactivated fetal bovine serum (HI FBS), 1x penicillin/streptomycin) and allowed to attach overnight in an incubator at 37°C. The growth medium was washed and replaced with an analytical medium consisting of Gibco OptiMEM 1 reduced serum medium, 0.1% casein, 8.33 mM D-fructose- ¹³ C6 and compound at concentrations ranging from 100 to 0.0051 μM (10- point concentration curve). The plates were incubated at 37°C for 3 h, after which the assay medium was pumped out from the cell wells. A stopping solution consisting of 80% MeOH, 2 mM ammonium acetate, and 50 ng/ml fructose-6phosphate- ^13C6 was then added to the cells. The plates were stored at -20°C until RapidFire MS analysis (described above).

Compounds of Examples 1 to 36 were tested essentially as described above.

- 42 043635

Table 7

Example number HepG2 IC50 1 41.1 ±8.13 (n=3) 2 98.4 ± 8.69 (n=3) 3 40.2 ± 10.9 (n=4) 4 16.8 ±4.97 (n=4) 5 83.6 ± 8.16 (n=3) 6 22.9 ±5.15 (n=3) 7 8.06 ± 1.41 (n=3) 8 20.4 ± 8.09 (n=3) 9 15.0 (n=1) 10 118 ±24.2 (n=3) AND 48.4 (n=1) 12 5.36 ± 0.0912 (n=3) 13 26.2 ± 4.30 (n=3) 14 34.9 ± 8.26 (n=3) 15 52.7 ±3.08 (n=3) 16 73.2 ± 14.9 (n=3) 17 1.66 ±0.435 (n=3) 18 16.3 ± 1.84 (n=3) 19 95.9 (n=1) 20 35.7 ±3.47 (n=3) 21 102 ±5.47 (n=3) 22 109 ±9.69 (n=3) 23 76.2 ± 7.07 (n=3) 24 27.0 (n=1) 24a 18.2 (n=1) 246 27.2 (n=1) 25 33.5 ± 9.07 (n=3) 26 81.6 (n=1) 27 6.93 (n=1) 28 144(n=1) 29 145(n=1) thirty 143(n=1) 31 119(n=1) 32 62.9 (n=1) 33 6.13 ±0.528 (n=3) 34 2.93 ±0.156 (n=3) 35 6.41 (n=1) 36 13.4 (n=1)

Data are expressed as mean ± SEM (n).

The results presented in table. 7 above demonstrate that compounds of Examples 1 to 36 inhibit fructose metabolism to F1P in HepG2 cells.

Liquid chromatographic method with tandem mass spectrometry (LC-MS/MS) for pro

Claims (19)

conducting pharmacokinetic analyses. Samples were extracted by protein precipitation by adding 180 μL of MeOH:ACN (1:1, v/v) containing internal standard to 50 μL of plasma. Samples were then diluted with MeOH:water (1:1, v/v) to obtain concentrations within the range of the standard curve. Diluted samples were analyzed by LC-MS/MS using a Sciex API 4000 triple quadrupole mass spectrometer (Applied Biosystems/MDS; Foster City, CA) equipped with a TurboIonSpray interface and operating in positive ion mode. The analyzed substances were separated chromatographically using an ECHELON C18 4 µm 20X2.1 mm column. LC conditions were: water/1 M ammonium bicarbonate (2000:10, v/v) (mobile phase A) and MeOH/1 M ammonium bicarbonate, (2000:10, v/v) (mobile phase B). Pharmacokinetics in Sprague Dawley rats. The in vivo pharmacokinetic properties of Example 1 and Example 2 were demonstrated using Sprague Dawley rats (fasted; n=3/route). The compound was administered as a single oral (PO; 2 or 3 mg/kg; volume 10 ml/kg) or intravenous (IV; 1 mg/kg; volume 1 ml/kg) dose in vehicle. Blood was collected from each animal at several time points from 0 to 48 hours after dosing. Plasma concentrations for Example 1 and Example 2 were determined by LC-MS/MS as described above. For Example 1, the mean half-life was 12.9 hours and the bioavailability was 83% as determined by PO administration, while by IV administration the mean half-life was 12.8 hours and the mean clearance was 5.86 ml/min. /kg. For Example 2, the mean half-life was 5.12 hours and the bioavailability was 95% as determined by PO administration, while by IV administration the mean half-life was 4.29 hours and the mean clearance was 56.4 ml/min. /kg. These data indicate that Examples 1 and 2 have different clearance levels, but both have high oral bioavailability and prolonged elimination, as evidenced by an adequate mean half-life. Pharmacokinetics in dogs. The in vivo pharmacokinetic properties of Example 1 were demonstrated in Beagle dogs (after feeding, n=3). The compound was administered as a single oral (PO; 2 or 3 mg/kg; volume 2 ml/kg) or intravenous (IV; 1 mg/kg; volume 1 ml/kg) dose in vehicle. Blood was collected from each animal at several time points from 0 to 72 hours after dosing. Plasma concentrations for Example 1 and Example 2 were determined by LC-MS/MS as described above. For Example 1, the mean half-life was 36.6 hours and the bioavailability was 87% as determined by PO administration, while by IV administration the mean half-life was 28 hours and the mean clearance was 3.41 ml/min/kg . For Example 2, the mean half-life was 9.79 hours and bioavailability was ~100% as determined by PO administration, while by IV administration the mean half-life was 10.3 hours and the mean clearance was 19.6 mL. min/kg. These data indicate that Examples 1 and 2 have different levels of clearance, but both have high oral bioavailability and prolonged elimination, as evidenced by an adequate mean half-life. CLAIM 1. Compound formula where X represents N or C substituted with CN; R ¹ selected from H, - 44 043635 R ² and R ³ are both H or one is H and the other is OH; R ⁴ , R ⁵ , R ⁶ , R ⁷ and R ⁹ are independently H or CH ₃ ; R ⁸ is H, CH ₃ , CH ₂ CH ₂ OH, C(=O)CH ₂ NH ₂ or C(=O)CH ₃ ; And R ¹⁰ represents OH or NH ₂ ; or a pharmaceutically acceptable salt thereof. 2. The connection according to claim 1, characterized in that R ¹ is selected from or a pharmaceutically acceptable salt thereof. 3. The connection according to claim 1 or 2, characterized in that the connection is or a pharmaceutically acceptable salt thereof. 4. A compound according to any one of claims 1 to 3, characterized in that X represents N, or a pharmaceutically acceptable salt thereof. 5. A compound according to any one of claims 1 to 3, characterized in that X represents C substituted with CN, or a pharmaceutically acceptable salt thereof. 6. The connection according to any one of claims 1-4, characterized in that the connection is or a pharmaceutically acceptable salt thereof. 7. The compound or a pharmaceutically acceptable salt thereof according to claim 6, characterized in that the pharmaceutically acceptable salt is a succinate salt. 8. A method of treating type 2 diabetes mellitus in a patient, comprising administering to a patient in need of such treatment an effective amount of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof. 9. A method of treating heart failure in a patient, comprising administering to a patient in need of such treatment an effective amount of a compound according to any one of claims. 1-7 or a pharmaceutically acceptable salt thereof. 10. A method of treating diabetic kidney disease in a patient, comprising administering to the patient in need of such treatment an effective amount of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof. 11. A method of treating non-alcoholic steatohepatitis in a patient, comprising administering to a patient in need of such treatment an effective amount of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof. - 45 043635 12. A method of treating chronic kidney disease in a patient, comprising administering to the patient in need of such treatment an effective amount of a compound according to any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof. 13. Use of a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 for the treatment of type 2 diabetes mellitus. 14. Use of a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 for the treatment of heart failure. 15. Use of a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 for the treatment of diabetic kidney disease. 16. Use of a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 for the treatment of non-alcoholic steatohepatitis. 17. Use of a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 for the treatment of chronic kidney disease. 18. A pharmaceutical composition containing a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 with one or more pharmaceutically acceptable carriers, diluents or excipients. 19. A method for preparing a pharmaceutical composition, comprising mixing a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 with one or more pharmaceutically acceptable carriers, diluents or excipients. Eurasian Patent Organization, EAPO Russia, 109012, Moscow, Maly Cherkassky lane, 2