Glucagon-like peptide 1 receptor agonists

Abstract

In one embodiment, the present invention provides a compound of the formula: or a pharmaceutically acceptable salt thereof, and methods of using such compounds to treat type II diabetes.

Description

[0001] The present invention relates to a glucagon-like peptide-1 receptor agonist and the therapeutic use of the compound for treating type II diabetes.

[0002] Glucagon-like peptide-1 (GLP-1) is a member of the incretin family of peptide hormones secreted by enteroendocrine L-cells. GLP-1 induces insulin release from beta cells in a glucose-dependent manner. However, GLP-1 is rapidly metabolized so that only a small percentage of GLP-1 is available to induce insulin secretion. To compensate for this, GLP-1 receptor (GLP-1R) agonists have been developed to enhance insulin secretion as a treatment for type II diabetes.

[0003] Most GLP-1R agonists approved for the treatment of type 2 diabetes are injectable. Due to the disadvantages associated with injections, such as inconvenience, pain, and the potential for injection site irritation, patients generally prefer oral medications.

[0004] WO2018 / 109607 discloses certain benzimidazole derivatives described as GLP-1R agonists.

[0005] However, there is a need for alternative GLP-1R agonists. In particular, there is a need for GLP-1R agonists that can be administered orally. In particular, there is a need for GLP-1R agonists that have improved efficacy, favorable toxicological profiles, and / or pharmacokinetic profiles to support once-daily dosing.

[0006] Accordingly, the present invention provides a compound of the following formula or a pharmaceutically acceptable salt thereof:

[0007] ,

[0008] in

[0009] R 1 is H or F;

[0010] R 2 is H or F; and

[0011] R 3 It is H or CH3.

[0012] Formula I includes all individual enantiomers and mixtures thereof, as well as racemates, and pharmaceutically acceptable salts thereof.

[0013] In one embodiment, a compound of the formula is provided:

[0014]

[0015] or a pharmaceutically acceptable salt thereof.

[0016] In one embodiment, a compound of the formula: or a pharmaceutically acceptable salt thereof is provided:

[0017] ,

[0018] Where R 1 It is H or F.

[0019] In one embodiment, a compound of the formula: or a pharmaceutically acceptable salt thereof is provided:

[0020] ,

[0021] Where R 1 It is H or F.

[0022] In one embodiment, the compound is of the formula:

[0023]

[0024] Or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the compound is a compound of the formula:

[0025]

[0026] or a pharmaceutically acceptable salt thereof.

[0027] In one embodiment, the compound is of the formula:

[0028]

[0029] Or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the compound is a compound of the formula:

[0030]

[0031] Or a pharmaceutically acceptable salt thereof. In a particularly preferred embodiment, the tert-butylamine salt (also known as the erbumine salt) of the compound of the formula:

[0032] .

[0033] In one embodiment, a compound of the formula: or a pharmaceutically acceptable salt thereof is provided:

[0034] ,

[0035] Where R 2 It is H or F.

[0036] In one embodiment, a compound of the formula: or a pharmaceutically acceptable salt thereof is provided:

[0037]

[0038] Where R 2It is H or F.

[0039] In one embodiment, the compound is of the formula:

[0040]

[0041] Or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the compound is a compound of the formula:

[0042]

[0043] or a pharmaceutically acceptable salt thereof.

[0044] In one embodiment, the compound is of the formula:

[0045]

[0046] Or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the compound is a compound of the formula:

[0047]

[0048] or a pharmaceutically acceptable salt thereof.

[0049] Formula I encompasses formulae Ia, Ib, II, IIa, IIb, III, IIIa and IIIb and hereinafter, for example in the methods of treatment and therapeutic uses, a reference to formula I is also considered to be a reference to each and all of these subformulae.

[0050] In another embodiment, a pharmaceutically acceptable composition is provided, comprising a compound of Formula I or a pharmaceutically acceptable salt thereof and at least one pharmaceutically acceptable carrier, diluent or excipient. In a preferred embodiment, the pharmaceutically acceptable composition is formulated for oral administration.

[0051] In another embodiment, a method for treating type II diabetes in a mammal is provided, the method comprising administering to a mammal in need of treatment a pharmaceutically acceptable composition comprising an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof and at least one pharmaceutically acceptable carrier, diluent or excipient. In one embodiment, the pharmaceutically acceptable composition is formulated for oral administration. Preferably, the mammal is a human.

[0052] In another embodiment, a method of treating type II diabetes in a mammal is provided, the method comprising administering to a mammal in need of treatment an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the mammal is a human.

[0053] In another embodiment, a method of lowering blood glucose levels in a mammal is provided, the method comprising administering to a mammal in need of treatment an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the mammal is a human.

[0054] In another embodiment, a method of treating hyperglycemia in a mammal is provided, the method comprising administering to the mammal in need of treatment an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the mammal is a human.

[0055] In one embodiment, a compound of Formula I or a pharmaceutically acceptable salt thereof is provided for use in therapy.

[0056] In another embodiment, a compound of Formula I or a pharmaceutically acceptable salt thereof is provided for use in treating type II diabetes.

[0057] In another embodiment, a compound of Formula I or a pharmaceutically acceptable salt thereof is provided for use in lowering blood glucose levels.

[0058] In another embodiment, there is also provided a compound of Formula I or a pharmaceutically acceptable salt thereof for use in treating hyperglycemia.

[0059] In one embodiment, there is provided a use of a compound of Formula I or a pharmaceutically acceptable salt thereof for the preparation of a medicament for treating type II diabetes.

[0060] In one embodiment, there is provided a use of a compound of Formula I or a pharmaceutically acceptable salt thereof for the preparation of a medicament for lowering blood glucose levels.

[0061] In one embodiment, there is provided the use of a compound of Formula I or a pharmaceutically acceptable salt thereof for the preparation of a medicament for treating hyperglycemia.

[0062] In a preferred embodiment, the compound of formula I is administered orally. In a preferred embodiment, the compound of formula I is administered once daily. In another preferred embodiment, the therapeutic use is in humans.

[0063] The term "pharmaceutically acceptable salt" as used herein refers to salts of the compounds of the present invention that are considered acceptable for clinical and / or veterinary use. Examples of pharmaceutically acceptable salts and common methods for preparing them can be found in "Handbook of Pharmaceutical Salts: Properties, Selection and Use" P. Stahl et al., Second Revised Edition, Wiley-VCH, 2011 and SM Berge et al., "Pharmaceutical Salts", Journal of Pharmaceutical Sciences, 1977, 66(1), 1-19.

[0064] Examples of pharmaceutical compositions and methods for their preparation may be found in "Remington: The Science and Practice of Pharmacy", edited by Loyd, V. et al., 22 nd Ed., Mack Publishing Co., 2012. In one embodiment, the pharmaceutical composition can be formulated for oral administration. The pharmaceutical composition is preferably formulated as a tablet, capsule or solution. The tablet, capsule or solution may include a compound of Formula I in an effective amount for treating a patient in need of treatment.

[0065] The term "effective amount" refers to the amount or dosage of a compound of Formula I or a pharmaceutically acceptable salt thereof that provides the desired effect in a patient being diagnosed or treated when administered to the patient in a single dose or multiple doses. The attending physician, as a person skilled in the art, can easily determine the effective amount by using conventional techniques and by observing the results obtained under similar circumstances. Factors considered in the determination of the effective amount or dosage of the compound include: whether the compound or its salt is administered; if used, the co-administration of other agents; the species of the mammal being treated; its size, age and general health; the involvement or severity of the condition; the response of the individual mammal; the mode of administration; the bioavailability characteristics of the administered formulation; the selected dosing regimen; and other relevant circumstances. The compounds of the present invention are effective at daily doses falling within the range of about 0.01 to about 15 mg / kg body weight.

[0066] As used herein, the term "treat," "treat," or "treating" refers to reducing, alleviating, or reversing an existing symptom, disorder, or condition, such as the progression or severity of hyperglycemia, which may include increasing insulin secretion.

[0067] The compound of formula I can be formulated as a pharmaceutical composition for administration by any route that can bioavailable the compound. Such compositions are preferably used for oral administration. Such pharmaceutical compositions and methods for preparing them are well known in the art (see, for example, Remington, JP, "Remington: The Science and Practice of Pharmacy", edited by LV Allen, 22nd edition, Pharmaceutical Press, 2012).

[0068] The compounds of formula I and their pharmaceutically acceptable salts are useful in the therapeutic applications of the present invention, with certain configurations being preferred.

[0069] The compounds of the present invention include:

[0070]

[0071]

[0072] or a pharmaceutically acceptable salt thereof.

[0073] Additional compounds of the invention include:

[0074]

[0075]

[0076] or a pharmaceutically acceptable salt thereof.

[0077] Additional compounds of the invention include:

[0078]

[0079]

[0080] or a pharmaceutically acceptable salt thereof.

[0081] While the present invention contemplates all individual enantiomers, mixtures and racemates thereof, compounds of Formula Ia, IIa and IIIa and pharmaceutically acceptable salts thereof are particularly preferred.

[0082] 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 fluid chromatography (SFC) (see, e.g., TA Berger; “ Supercritical Fluid Chromatography Primer ,” Agilent Technologies, July 2015) to separate or resolve the individual enantiomers.

[0083] Pharmaceutically acceptable salts of the compounds of the invention can be formed, for example, by reacting a compound of formula I with an appropriate pharmaceutically acceptable base in a suitable solvent under standard conditions well known in the art (see, for example, Bastin, RJ et al., Org. Process. Res. Dev ., 4, 427-435, 2000; and Berge, SM et al., J. Pharm. Sci. , 66, 1-19, 1977). The preferred salt is the tert-butylamine (or erbumine) salt.

[0084] Certain abbreviations used herein are based on Daub GH et al. "The Use of Acronyms in Organic Chemistry" Aldrichimica Acta , 1984, 17(1), 6-23. Certain abbreviations are defined as follows: "ACN" means acetonitrile; "ATP" means adenosine triphosphate; "BSA" means bovine serum albumin; "cAMP" means cyclic adenosine-3',5'-monophosphate; "DCM" means dichloromethane or methylene chloride; "DIPEA" means N,N-diisopropylethylamine; "DMF" means N,N-dimethylformamide; "DMSO" means dimethyl sulfoxide; "EC 50 ” refers to the concentration of the agent that produces a 50% response of the target activity compared to the predetermined positive control compound (absolute EC 50 ); "ES / MS" refers to electrospray mass spectrometry; "EtOAc" refers to ethyl acetate; "HATU" refers to 1-[bis(dimethylamino)methylene]-1-fluorophosphate H -1,2,3-triazolo[4,5- b ]pyridinium 3-oxide; “HEK” refers to human embryonic kidney; “HEPES” refers to 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; “h” refers to hour; “MeOH” refers to methanol or methyl alcohol; “min” refers to minute; “Pd(dppf)Cl2” refers to [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II); “RT” refers to room temperature; and “THF” refers to tetrahydrofuran.

[0085] The compounds of the present invention can be prepared by various procedures, some of which are described in the following preparations and examples. The specific synthetic steps of each of the routes described can be combined in different ways to prepare the compounds of the present invention or their salts. The products of the following steps can be recovered by conventional methods, including extraction, evaporation, precipitation, chromatography, filtration, trituration and crystallization. Reagents and raw materials are readily available to those of ordinary skill in the art. Independent isomers, enantiomers and diastereomers can be separated or split at any convenient point in the synthesis by methods such as selective crystallization techniques or chiral chromatography (see, for example, J. Jacques et al., " Enantiomers, Racemates, and Resolutions ", John Wiley andSons, Inc., 1981 and EL Eliel and SH Wilen, " Stereochemistry of Organic Compounds ”, Wiley-Interscience, 1994). The following preparations and examples are provided to further illustrate the present invention without limiting the scope of the present invention.

[0086] Solution 1

[0087]

[0088] Scheme 1 shows the synthesis of intermediate 6 for preparing compounds of Formula I. Benzoic acid 1 is first reduced with borane dimethyl sulfide complex in step 1 to produce alcohol 2. The alcohol is converted to a leaving group (LG, intermediate 3). For example, the alcohol in intermediate 2 can be converted to a mesylate group using methanesulfonyl chloride at -15°C in step 2, or it can be converted to a bromide using phosphorus tribromide at 0°C. Intermediate 3 is reacted with NaCN in step 3 to produce nitrile 4. Nitrile 4 is converted with KOH at elevated temperature in step 4 to produce acid 5, which is then esterified using oxalyl chloride, DMF and methanol in step 5 to produce intermediate 6.

[0089] Solution 2

[0090]

[0091] Scheme 2 depicts the preparation of key intermediate 12 for the preparation of compounds of Formula I via two routes. In the first route, aryl halide 6 undergoes a one-pot Miyura borylation / Suzuki coupling: aryl halide 6 is converted to boronate ester 7 in step 1a using bis(pinacolato)diboron, Pd(dppf)Cl2 and potassium acetate at elevated temperature, after which bromopyridine 8 and K2CO3 are added to the reaction (step 2a) to produce intermediate 12. In the second route, a two-step process is used: Suzuki coupling of aryl halide 6 with 6-hydroxypyridine-2-boronic acid pinacol ester 9 using Pd(dppf)Cl2 and K2CO3 at elevated temperature (step 1b) provides intermediate 10, which is subsequently alkylated with 4-(bromomethyl)-3-fluorobenzonitrile 6 using Ag2CO3 at elevated temperature (step 2b) to produce intermediate 12. Ester hydrolysis of intermediate 12 using LiOH in step 3 produces acid intermediate 13.

[0092] Solution 3

[0093]

[0094] Alternatively, key intermediates 12 and 13 can be prepared according to Scheme 3 by coupling bromopyridine 8 with boronate ester 7 or boronic acid 14 using Pd(dppf)Cl2 and potassium carbonate at elevated temperature.

[0095] Solution 4

[0096]

[0097] Scheme 4 shows the conversion of key intermediate 13 to compounds of formula I. Amide coupling using HATU and diphenylamine 15 in step 1 produces intermediate 16. Cyclization is achieved by heating intermediate 16 in acetic acid (step 2) to produce benzimidazole 17. Finally, in step 3, compounds of formula I are obtained by hydrolysis of 17 using LiOH.

[0098] Preparation and Examples

[0099] In AGILENT ® LC-ES / MS was performed on a HP1200 LC system. Electrospray mass spectrometry measurements (acquired in positive and / or negative mode) were performed on a mass selective detector quadrupole mass spectrometer connected to an HPLC with or without ELSD. LC-ES / MS conditions (low pH): Column: PHENOMENEX ® GEMINI ®NX C18 2.0 × 50 mm 3.0 μm, 110 Å; Gradient: 5-95% B in 1.5 min, then 95% B for 0.5 min, Column temp: 50°C + / -10°C; Flow rate: 1.2 mL / min; 1 μL injection volume; Solvent A: 0.1% HCOOH in deionized water; Solvent B: 0.1% formic acid in ACN; Wavelengths 200-400 nm and 212-216 nm. If the HPLC is equipped with an ELSD, set to 45°C evaporator temperature, 40°C nebulizer temperature, and 1.6 SLM gas flow rate. Alternative LC-MS conditions (high pH): Column: Waters xBridge ® C18 column 2.1×50 mm, 3.5 μm; Gradient: 5-95% B in 1.5 min, then 95% B for 0.50 min; Column temperature: 50℃ + / -10℃; Flow rate: 1.2 mL / min; 1 μL injection volume; Solvent A: 10 mM NH4HCO3 pH 9; Solvent B: ACN; Wavelengths: 200-400 nm and 212-216 nm; If with ELSD: 45℃ evaporator temperature, 40℃ nebulizer temperature and 1.60 SLM gas flow rate.

[0100] The X-ray powder diffraction (XRPD) pattern of the crystalline solid was obtained on a Bruker D4 Endeavor X-ray powder diffractometer equipped with a CuKα source and a Vantec detector, operating at 35 kV and 50 mA. The sample was scanned between 4 and 40 2θ° with a step size of 0.008 2θ° and a scan rate of 0.5 seconds / step and using a 1.0 mm divergence slit, a 6.6 fixed anti-scatter slit, and an 11.3 mm detector slit. The dry powder was mounted on a quartz sample holder and a glass slide was used to obtain a smooth surface. The crystal diffraction pattern was collected at ambient temperature and relative humidity. Based on the NIST 675 internal standard with peaks at 8.853 and 26.774 2θ°, the crystal peak positions were determined in MDI-Jade after full pattern shift. It is well known in the field of crystallography that for any given crystal form, the relative intensity of the diffraction peaks can be variable due to preferred orientation caused by factors such as crystal morphology and habit. If there is a preferred orientation effect, the peak intensity changes, but the characteristic peak position of the polymorph remains unchanged. See, for example, The United States Pharmacopeia #23, National Formulary #18, pp. 1843-1844, 1995. In addition, it is also well known in the field of crystallography that for any given crystal form, the angular peak position may change slightly. For example, the peak position may move due to temperature changes, sample displacement, or the presence or absence of an internal standard when analyzing the sample. In this case, a peak position variability of ± 0.2 2θ° is assumed to take into account these potential changes without hindering the clear identification of the crystal form shown. The crystal form can be confirmed based on any unique combination of characteristic peaks.

[0101] Preparation 1

[0102] (4-Bromo-2-fluoro-5-methylphenyl)methanol

[0103]

[0104] To a flask was added: 4-bromo-2-fluoro-5-methylbenzoic acid (100 g, 421 mmol), THF (200 mL) and borane (dimethyl sulfide complex, 2 mol / L solution in THF, 210 mL, 10 mmol). The mixture was stirred at room temperature overnight. The reaction mixture was quenched with HCl (1.0 N aqueous solution, 50 mL) and the mixture was filtered. The filtrate was concentrated in vacuo and the residue was partitioned between EtOAc (400 mL) and water (400 mL). The organics were washed with saturated aqueous NaCl solution (400 mL), dried over Na2SO4, filtered and concentrated to yield the title compound (93.5 g, 99%) as a solid. 1H-NMR (400MHz, CDCl3) δ 7.29 (d, J = 7.9 Hz, 1H), 7.26 (d, J = 9.1 Hz, 1H), 4.69 (s, 2H), 2.38 (s, 3H).

[0105] Preparation 2

[0106] (4-Bromo-2-fluoro-3-methyl-phenyl)methanol

[0107]

[0108] Prepare the title compound using 4-bromo-2-fluoro-3-methylbenzoic acid essentially as described in Preparation 1. Purify the product by silica gel chromatography using a 10 to 35% EtOAc / hexanes gradient. LC-ES / MS peak retention time: 1.01 min.

[0109] Preparation 3

[0110] 2-(4-Bromo-2-fluoro-5-methylphenyl)acetonitrile

[0111]

[0112] (4-Bromo-2-fluoro-5-methylphenyl)methanol (92 g, 420 mmol) was dissolved in DCM (500 mL) and triethylamine (120 mL, 861 mmol) was added. The mixture was cooled to -15 °C and a solution of methanesulfonyl chloride (40 mL, 517 mmol) in DCM (30 mL) was added dropwise to the reaction mixture. The mixture was stirred at room temperature for 30 minutes. The reaction mixture was partitioned between DCM (500 mL) and water (500 mL). The organics were washed with saturated aqueous NaCl solution (500 mL), dried over Na2SO4, filtered and concentrated. The residue was dissolved in DMF (400 mL) and the mixture was cooled with an ice bath. NaCN (21.0 g, 429 mmol) was added to the reaction mixture in one portion and stirred at room temperature overnight. The mixture was partitioned between EtOAc (400 mL) and water (500 mL). The organics were washed with saturated aqueous NaCl (500 mL), dried over Na2SO4, filtered and concentrated.The residue was purified by silica gel chromatography using a 10 to 30% EtOAc / hexanes gradient to give the title compound as an oil (47.0 g, 48%). 1 H-NMR (400 MHz, CDCl3) δ 7.34 (d, J= 8.7 Hz, 1H),7.32 (d, J = 8.1 Hz, 1H), 3.71 (s, 2H), 2.41 (s, 3H).

[0113] Preparation 4

[0114] 2-(4-Bromo-2-fluoro-3-methyl-phenyl)acetonitrile

[0115]

[0116] (4-Bromo-2-fluoro-3-methyl-phenyl)methanol (1.90 g, 8.67 mmol) and DCM (20 mL) were mixed together. The mixture was cooled to 0°C and phosphorus tribromide (1.0 mL, 11 mmol) was then added dropwise. The mixture was stirred at 0°C for 15 minutes and then the mixture was basified with saturated aqueous NaHCO3 (10 mL). The mixture was extracted with DCM (40 mL). The organics were washed with brine (30 mL), dried over (Na2SO4), filtered and concentrated to give a solid. The solid was dissolved in DMSO (10 mL) and NaCN (0.60 g, 13.0 mmol) was added and stirred for 1 h. The mixture was partitioned between EtOAc (50 mL) and water (50 mL). The organics were washed with brine (50 mL), dried over Na2SO4, filtered and concentrated to give the product as a solid (1.3 g, 64%). LC-ES / MS peak retention time: 1.17 min.

[0117] Preparation 5

[0118] 2-(4-Bromo-2-fluoro-5-methyl-phenyl)acetic acid methyl ester

[0119]

[0120] To a flask was added: 2-(4-bromo-2-fluoro-5-methylphenyl)acetonitrile (1.20 g, 5.10 mmol), ethanol (5 mL), water (3 mL), and potassium hydroxide (0.90 g, 16 mmol). The mixture was heated at 90 °C overnight. The mixture was cooled with an ice bath and acidified to pH 4-5 with 1.0 M HCl, then the mixture was partitioned between EtOAc (30 mL) and water (30 mL). The organics were washed with saturated aqueous NaCl solution (30 mL), dried over Na2SO4, filtered, and concentrated to yield 2-(4-bromo-2-fluoro-5-methyl-phenyl)acetic acid as a solid. This was dissolved in DCM (10 mL), then DMF (0.05 mL, 0.6 mmol) and oxalyl chloride (0.5 mL, 6 mmol) were added at room temperature. The mixture was stirred at room temperature for 30 minutes, then MeOH (2 mL, 49.4 mmol) was added dropwise. After 30 minutes, the solvent was removed in vacuo and the residue was partitioned between EtOAc (40 mL) and 5% NaHCO3 (30 mL). The organics were washed with saturated aqueous NaCl (40 mL), dried over Na2SO4, filtered and concentrated to give the title compound as an oil (1.1 g, 80%). ES / MS m / z ( 79 Br, 81 Br) 278,280 (M+NH4 + ).

[0121] Preparation 6

[0122] 2-(4-Bromo-2-fluoro-3-methyl-phenyl)acetic acid methyl ester

[0123]

[0124] Prepare the title compound using 2-(4-bromo-2-fluoro-3-methyl-phenyl)acetonitrile essentially as described in Preparation 5. LC-ES / MS peak retention time: 1.22 min.

[0125] Preparation 7

[0126] Methyl 2-(4-bromo-2,6-difluorophenyl)acetate

[0127]

[0128] 4-Bromo-2,6-difluorophenylacetic acid (3.30 g, 12.5 mmol), DCM (20 mL), DMF (0.05 mL, 0.6 mmol) and oxalyl chloride (1.3 mL, 15 mmol) were mixed. The mixture was stirred at room temperature for 30 minutes, and then MeOH (1.5 mL, 37 mmol, 100 mass %) was added dropwise. The mixture was concentrated and partitioned between EtOAc (30 mL) and saturated aqueous NaHCO3 (15 mL). The organics were washed with saturated aqueous NaCl (30 mL), dried over Na2SO4, filtered and concentrated to give the title compound (3.41 g, quantitative yield) as an oil, which was used to prepare 10 without further purification. ES / MS m / z ( 79 Br, 81 Br) 265,267 (M+H).

[0129] Preparation 8

[0130] 4-[(6-Bromo-2-pyridyl)oxymethyl]-3-fluoro-benzonitrile

[0131]

[0132] 2-Bromo-6-fluoropyridine (2.50 g, 13.8 mmol) and 3-fluoro-4-(hydroxymethyl)benzonitrile (2.15 g, 13.8 mmol) were dissolved in 1,4-dioxane (25 mL) and a solution of potassium tert-butoxide (20 wt % in THF, 10.0 mL, 16.6 mmol) was added dropwise at room temperature over 12 hours. The reaction mixture was heated at 40°C for 30 minutes. The mixture was poured into aqueous K2CO3 solution (1 M) and extracted twice with EtOAc. The organics were washed with water and saturated aqueous NaCl solution, dried over Na2SO4, filtered and concentrated. The residue was dried in a vacuum oven at 50°C to give the title compound (4.23 g, 95%) as a light yellow solid. ES / MS m / z ( 79 Br, 81 Br) 307,309 (M+H).

[0133] Preparation 9

[0134] 2-[2-Fluoro-4-(6-hydroxy-2-pyridinyl)-5-methyl-phenyl]acetic acid methyl ester

[0135]

[0136] To a flask was added 6-hydroxypyridine-2-boronic acid pinacol ester (1.6 g, 6.9 mmol), methyl 2-(4-bromo-2-fluoro-5-methyl-phenyl)acetate (2.2 g, 8.4 mmol), THF (15 mL), water (1 mL), and potassium carbonate (2.0 g, 14 mmol). The mixture was purged with nitrogen for 10 minutes, then Pd(dppf)Cl2 (0.26 g, 0.35 mmol) was added and heated at 75 °C for 2 hours. The mixture was partitioned between EtOAc (30 mL) and water (30 mL). The organics were washed with saturated aqueous NaCl solution (30 mL), dried over Na2SO4, filtered and concentrated to give the title compound as a solid (1.4 g, 74%). ES / MS m / z 276 (M+H), 274 (MH).

[0137] Preparation 10

[0138] Methyl 2-[2,6-difluoro-4-(6-hydroxy-2-pyridinyl)phenyl]acetate

[0139]

[0140] The title compound was prepared essentially as described in Preparation 9 using methyl 2-(4-bromo-2,6-difluorophenyl)acetate and the reaction was heated at 75°C overnight. ES / MS m / z 280 (M+H).

[0141] Preparation 11

[0142] 2-[2-Fluoro-4-(6-hydroxy-2-pyridinyl)-3-methyl-phenyl]acetic acid methyl ester

[0143]

[0144] The title compound was prepared essentially as described in Preparation 9 using methyl 2-(4-bromo-2-fluoro-3-methyl-phenyl)acetate and the reaction was heated at 75 °C overnight (18 h). ES / MS m / z 276 (M+H), 274 (MH).

[0145] Preparation 12

[0146] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-3-methyl-phenyl]acetic acid methyl ester

[0147]

[0148] 2-(4-Bromo-3-methylphenyl)acetic acid (10.7 g, 45.8 mmol) was dissolved in DCM (50 mL). The mixture was cooled in an ice / water bath, and oxalyl chloride (4.8 mL, 55 mmol) and DMF (0.1 mL) were added. The ice / water bath was removed and stirred at room temperature for 2 hours. MeOH (6.0 mL) was added dropwise over 2 minutes and stirred at room temperature for 1 hour. The reaction mixture was concentrated in vacuo and the residue was dissolved in EtOAc. The organics were washed with saturated aqueous NaHCO3 and saturated aqueous NaCl. The organics were dried over Na2SO4, then filtered and concentrated. To the residue were added bis(pinacolato)diboron (12.8 g, 50.4 mmol) and potassium acetate (13.6 g, 137 mmol). Nitrogen was bubbled through the reaction mixture for 15 minutes, and then Pd(dppf)Cl2 (complex with DCM, 1.13 g, 1.37 mmol) was added. The reaction was heated at 85 °C in an oil bath under nitrogen for 15 hours, then the reaction flask was removed from the oil bath. Potassium carbonate (9.49 g, 68.7 mmol) was dissolved in water (60 mL), nitrogen was bubbled through the solution for 10 minutes, and then the solution was added to the reaction mixture, followed by 4-[(6-bromo-2-pyridyl)oxymethyl]-3-fluoro-benzonitrile (14.1 g, 45.8 mmol). Nitrogen was bubbled through the entire reaction mixture for 5 minutes and heated at 85 °C under nitrogen for 6 hours. The reaction was cooled to near room temperature and concentrated in vacuo to remove most of the 1,4-dioxane. The mixture was diluted with EtOAc (200 mL) and washed with water and saturated aqueous NaCl solution. The organics were dried over Na2SO4, then filtered and concentrated. The crude product was purified by silica gel chromatography using a 5 to 50% EtOAc / hexane gradient to produce the title compound (13.3 g, 70%) as a light yellow solid. ES / MS m / z 391 (M+H).

[0149] Preparation 13

[0150] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]acetic acid methyl ester

[0151]

[0152] To a flask was added methyl 2-[2-fluoro-4-(6-hydroxy-2-pyridinyl)-5-methyl-phenyl]acetate (1.40 g, 5.09 mmol), 1,4-dioxane (35 mL), silver carbonate (1.7 g, 6.2 mmol) and 4-(bromomethyl)-3-fluorobenzonitrile (1.4 g, 6.2 mmol). The mixture was heated at 60 °C overnight. The solid was filtered off and the filtrate was concentrated. The residue was purified by silica gel chromatography using 12 to 55% EtOAc / hexanes to produce the title compound (1.60 g, 77%) as a solid. ES / MS m / z 409 (M+H), 407 (MH).

[0153] Preparation 14

[0154] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-phenyl]acetic acid methyl ester

[0155]

[0156] A flask was charged with 4-[(6-bromo-2-pyridinyl)oxymethyl]-3-fluoro-benzonitrile (2.02 g, 6.58 mmol), methyl 2-(2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)acetate (2.99 g, 9.88 mmol), K2CO3 (2.30 g, 16.5 mmol), 1,4-dioxane (30 mL) and water (10 mL). Nitrogen was bubbled through the mixture for 10 minutes. Pd(dppf)Cl2 DCM complex (492 mg, 0.658 mmol) was added to the mixture and heated to 80 °C under nitrogen for 5 hours. The reaction mixture was cooled, diluted with EtOAc (75 mL) and filtered through Celite ® The filtrate was washed with water and saturated aqueous NaCl solution, dried over Na2SO4, filtered and concentrated. The resulting residue was purified by silica gel chromatography using a 5 to 90% EtOAc / hexanes gradient to afford the title compound (2.68 g, 94%). ES / MS m / z 395 (M+H).

[0157] Preparation 15

[0158] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2,6-difluoro-phenyl]acetic acid methyl ester

[0159]

[0160] The title compound was prepared essentially as described in Preparation 13 using methyl 2-[2,6-difluoro-4-(6-hydroxy-2-pyridinyl)phenyl]acetate and the reaction was heated at 80°C overnight. ES / MS m / z 413 (M+H).

[0161] Preparation 16

[0162] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-3-methyl-phenyl]acetic acid methyl ester

[0163]

[0164] The title compound was prepared essentially as described in Preparation 13 using methyl 2-[2-fluoro-4-(6-hydroxy-2-pyridinyl)-3-methyl-phenyl]acetate and the reaction was heated at 80°C for 3 hours. ES / MS m / z 409 (M+H).

[0165] Preparation 17

[0166] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-3-methyl-phenyl]acetic acid

[0167]

[0168] To a flask was added methyl 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-3-methyl-phenyl]acetate (1.20 g, 3.07 mmol), ACN (20 mL), water (10 mL) and lithium hydroxide (0.35 g, 15 mmol). The mixture was heated at 45 °C for 3 hours. The mixture was cooled with an ice bath and acidified to pH = 4-5 with 1.0 M HCl. The mixture was partitioned between EtOAc (30 mL) and water (30 mL). The organics were washed with brine (30 mL), dried over Na2SO4, filtered and concentrated to give the title compound as a solid (1.1 g, 95%). ES / MS m / z 377 (M+H).

[0169] Preparation 18

[0170] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]acetic acid

[0171]

[0172] To a vial was added methyl 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]acetate (1.6 g, 3.9 mmol), ACN (20 mL), water (6 mL) and lithium hydroxide (0.45 g, 19 mmol). The mixture was heated at 45 °C for 2 hours, cooled with an ice bath and acidified to pH = 4-5 with 1.0 M HCl. The mixture was partitioned between EtOAc (50 mL) and water (50 mL). The organics were washed with saturated aqueous NaCl solution (50 mL), dried over Na2SO4, filtered and concentrated to give the title compound as a solid (1.55 g, 100%). ES / MS m / z 395 (M+H).

[0173] Preparation 19

[0174] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-phenyl]acetic acid

[0175]

[0176] Methyl 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-phenyl]acetate (2.68 g, 6.25 mmol) was dissolved in THF (50 mL), and then lithium hydroxide (797 mg, 32.9 mmol) and water (20 mL) were added. After stirring at room temperature for 5 hours, the pH of the reaction mixture was adjusted to 5 with aqueous HCl (1 M). The volatile solvents were removed in vacuo to produce an aqueous slurry. The solid was filtered and dried to obtain the title compound (2.24 g, 88%). ES / MS m / z 381 (M+H).

[0177] Preparation 20

[0178] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2,6-difluoro-phenyl]acetic acid

[0179]

[0180] Prepare the title compound essentially as described in Preparation 18 using 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2,6-difluoro-phenyl]acetic acid methyl ester. ES / MS m / z 399 (M+H).

[0181] Preparation 21

[0182] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-3-methyl-phenyl]acetic acid

[0183]

[0184] The title compound was prepared using 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-3-methyl-phenyl]acetic acid methyl ester essentially as described in Preparation 17. ES / MS m / z 395 (M+H).

[0185] Preparation 22

[0186] 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]phenyl]acetic acid

[0187]

[0188] 4-[(6-Bromo-2-pyridyl)oxymethyl]-3-fluoro-benzonitrile (0.70 g, 2.3 mmol) and 2-(4-dihydroxyborylphenyl)acetic acid (0.64 g, 3.4 mmol), THF (15 mL), water (5 mL) and potassium carbonate (0.63 g, 4.6 mmol) were mixed together. The mixture was purged with nitrogen for 10 minutes, then Pd(dppf)Cl2 (0.085 g, 0.11 mmol) was added and the mixture was heated at 75 °C for 8 hours. The mixture was acidified to pH 4-5 with aqueous HCl (1 M). The mixture was partitioned between EtOAc (50 mL) and water (50 mL). The organics were washed with brine (50 mL), dried over (Na2SO4), then filtered and concentrated. The residue was purified by silica gel chromatography using a 25 to 65% EtOAc / hexanes gradient to give the title compound as a solid (800 mg, 97% yield). ES / MS m / z 363.0 (M+H).

[0189] Preparation 23

[0190] Methyl 4-amino-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate

[0191]

[0192] To a solution of methyl 3-fluoro-4-nitrobenzoate (2.0 g, 10 mmol) in THF (10 mL) and DMF (10 mL) at room temperature was added triethylamine (3.1 mL, 22 mmol). To the pale yellow solution was added [(2S)-oxetan-2-yl]methylamine (Austin Chemical Company, 1.0 g, 11 mmol) and the rust-colored solution was stirred overnight. The reaction was diluted with EtOAc (100 mL) and water (50 mL). The organic layer was separated and the aqueous layer was back extracted with EtOAc (2 × 50 mL). The organics were combined and washed with saturated aqueous NaCl. The organics were dried over Na2SO4, filtered, concentrated and the residue was dried under high vacuum. This resulted in crude methyl 4-nitro-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate (2.8 g, 10 mmol) as a yellow solid (ES / MS m / z 267 (M+H)).

[0193] Next, methyl 4-nitro-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate (2.8 g, 10 mmol) was dissolved in THF (50 mL) and palladium on carbon (pre-wetted with 5% water, 0.5 g) was added. The reaction mixture was purged with hydrogen in vacuo and then stirred at room temperature under a hydrogen balloon for 2 hours, during which the yellow color disappeared. ® The mixture was filtered and concentrated to give the title compound (2.4 g, 99%). ES / MS m / z 237 (M+H).

[0194] Preparation 24

[0195] 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-3-methyl-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoic acid methyl ester

[0196]

[0197] To a vial was added 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-3-methyl-phenyl]acetic acid (1.10 g, 2.92 mmol), DMF (10 mL), HATU (1.4 g, 3.6 mmol), methyl 4-amino-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate (0.76 g, 3.2 mmol), and DIPEA (1.5 mL, 8.6 mmol). The mixture was stirred at room temperature for 30 minutes and then partitioned between EtOAc (30 mL) and water (30 mL). The organics were washed with saturated aqueous NaCl solution (30 mL) and then dried over Na2SO4, filtered and concentrated. The residue was purified by silica gel chromatography using a 10 to 35% EtOAc / DCM gradient to give the title compound as a solid (1.2 g, 69%). ES / MS m / z 595 (M+1), 593 (M-1).

[0198] Preparation 25

[0199] 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoic acid methyl ester

[0200]

[0201] To a flask was added: 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]acetic acid (1.20 g, 3.04 mmol), DMF (15 mL), HATU (1.2 g, 3.1 mmol), methyl 4-amino-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate (0.80 g, 3.4 mmol) and DIPEA (1.5 mL, 8.6 mmol). The mixture was stirred at room temperature for 30 minutes and then partitioned between EtOAc (30 mL) and water (30 mL). The organics were washed with saturated aqueous NaCl solution (30 mL) and then dried over Na2SO4, filtered and concentrated. The residue was purified by silica gel chromatography using a 10 to 35% EtOAc / DCM gradient to give the title compound as a solid (1.20 g, 64%). ES / MS m / z 613 (M+1), 611 (MH).

[0202] Preparation 26

[0203] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid methyl ester

[0204]

[0205] To a round bottom flask was added 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-phenyl]acetic acid (205 mg, 0.540 mmol), methyl 4-amino-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate (116 mg, 0.490 mmol), HATU (224 mg, 0.589 mmol), DIPEA (0.26 mL, 1.5 mmol) and DMF (5 mL). After stirring at room temperature for 3.5 hours, the reaction mixture was diluted with EtOAc (30 mL) and washed with water and saturated aqueous NaCl solution. The organics were dried over Na2SO4, filtered and concentrated. The residue was purified by silica gel chromatography using a 0 to 10% MeOH / DCM gradient to obtain the intermediate amide (326 mg). ES / MS m / z 599 (M+H).

[0206] The intermediate amide was heated with acetic acid (5 mL) at 50 °C for 15 hours. The reaction mixture was concentrated in vacuo and the remaining residue was dissolved in EtOAc (25 mL). The organics were washed with saturated aqueous NaHCO3 and saturated aqueous NaCl. The organics were dried over Na2SO4, filtered and concentrated. The resulting product was purified by silica gel chromatography using a 20 to 100% EtOAc / hexane gradient to obtain the title compound (152 mg, 52%). ES / MS m / z 581 (M+H).

[0207] Preparation 27

[0208] 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2,6-difluoro-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoic acid methyl ester

[0209]

[0210] Prepare the title compound essentially as described in Preparation 24 using 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2,6-difluoro-phenyl]acetic acid. The product that precipitated during the aqueous workup was collected by filtration and used without further purification. ES / MS m / z 617 (M+H), 615 (MH).

[0211] Preparation 28

[0212] 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-3-methyl-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoic acid methyl ester

[0213]

[0214] Prepare the title compound essentially as described in Preparation 24 using 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-3-methyl-phenyl]acetic acid. ES / MS m / z 613 (M+H), 611 (MH).

[0215] Preparation 29

[0216] 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoic acid methyl ester

[0217]

[0218] Prepare the title compound essentially as described in Preparation 25 using 2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]phenyl]acetic acid. ES / MS m / z 581.0 (M+H), 579.0 (MH).

[0219] Example 1

[0220] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-3-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid

[0221]

[0222] To a vial was added methyl 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-3-methyl-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate (1.2 g, 2.0 mmol) and acetic acid (6 mL). The mixture was heated at 80 °C for 2 h and then the solvent was removed in vacuo. The residue was partitioned between EtOAc (30 mL) and aqueous NaHCO3 (5%, 20 mL). The organics were washed with saturated aqueous NaCl (30 mL), dried over Na2SO4, filtered and concentrated. The residue was dissolved in ACN (5 mL) and water (3 mL) and then LiOH (0.22 g, 9.2 mmol) was added to the mixture and stirred at 50 °C for 2 h. The solvent was removed in vacuo. The residue was purified by reverse phase flash chromatography using a 20 to 35% ACN / 5% NH4HCO3 aqueous solution gradient to give the title compound as a solid (900 mg, 79%). ES / MS m / z 563 (M+H), 561 (MH).

[0223] Example 2

[0224] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid

[0225]

[0226] To a vial was added methyl 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate (1.20 g, 1.96 mmol) and acetic acid (15 mL), then the mixture was heated at 80 °C for 2 hours. The solvent was removed in vacuo. The residue was partitioned between EtOAc (30 mL) and aqueous NaHCO3 (5%, 20 mL). The organics were washed with saturated aqueous NaCl (30 mL), dried over Na2SO4, filtered and concentrated. The residue was dissolved in ACN (10 mL) and water (4 mL), then LiOH (0.24 g, 10 mmol) was added to the mixture and stirred at 50 °C for 2 hours. The mixture was acidified to pH = 4-5 with saturated aqueous citric acid. The solvent was removed in vacuo. The residue was purified by reverse phase flash chromatography using a 20 to 35% ACN / 5% NH4HCO3 aqueous solution gradient to give the title compound as a solid (745 mg, 66%). ES / MS m / z 581 (M+H), 579 (MH).

[0227] Example 2a

[0228] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid tert-butylammonium

[0229] .

[0230] Method 1 – Seedless preparation

[0231] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid (555 mg, 0.96 mmol) was suspended in acetone (6 mL) at 50 °C while stirring at 800 rpm to produce a slurry of white solid. Tert-butylamine (115 µL, 1.09 mmol, 1.14 eq) was added and a brief clarification of the mixture was observed followed by precipitation of a white solid. The slurry was stirred at 50 °C for 1 hour, then heating was stopped and the sample was stirred as it reached room temperature. The solid was filtered off by vacuum filtration and dried in place under a stream of nitrogen for 15 minutes, then dried in vacuo at 50 °C for 1 hour to produce the title compound (612 mg, 98%).

[0232] Method 2 – Preparation using seed crystals

[0233] Mix 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid (50 g, 86.1 mmol), acetone (658 mL), and water (42 mL) together and heat the mixture to 50°C. Filter the mixture through GF / F paper and rinse with 94:6 v:v acetone:water (25 mL). Heat the resulting solution at 50°C. Prepare a solution of tert-butylamine (10 mL, 94.7 mmol, 1.1 eq) and 94:6 v:v acetone:water (25 mL). A portion of the tert-butylamine solution (7 mL) was added, followed by seeds of tert-butylammonium 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylate (50 mg). The remainder of the tert-butylamine solution was added via a syringe pump at a rate of 0.47 mL / min over approximately 1 hour. The resulting suspension was heated at 50°C for 2 hours, and then the mixture was cooled to ambient temperature overnight. The slurry was filtered and rinsed with acetone (2 × 100 mL). The wet cake was dried in vacuo at 50°C to constant weight to yield the title compound (51.8 g, 92%) as a light yellow solid.

[0234] A sample of the prepared title compound was characterized by an XRD pattern using CuKα radiation as having diffraction peaks (2-θ values) as described in Table 1 below, specifically having a peak at 6.9 and one or more peaks selected from 16.3 and 22.5; the tolerance for the diffraction angle was 0.2 degrees.

[0235] Table 1. X-ray powder diffraction peaks of tert-butylammonium 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-5-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylate

[0236]

[0237] Example 3

[0238] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-phenyl]methyl]-3-[(2S)-oxetan-2-ylmethyl]benzimidazole-5-carboxylic acid

[0239]

[0240] Methyl 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylate (152 mg, 0.256 mmol) was dissolved in THF (6 mL) and lithium hydroxide (31 mg, 1.26 mmol) and water (2 mL) were added. The mixture was stirred at room temperature for 16 hours and then the pH was adjusted to 6 with aqueous HCl (1 N). The THF was removed in vacuo and the remaining solid was collected by filtration. Purify by reverse phase flash chromatography using a gradient of 10 to 40% ACN / NH4HCO3 aqueous solution (10 mM, pH 10) to obtain the title compound (60 mg, 41%). ES / MS m / z 567 (M+H).

[0241] Example 4

[0242] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2,6-difluoro-phenyl]methyl]-3-[(2S)-oxetan-2-ylmethyl]benzimidazole-5-carboxylic acid

[0243]

[0244] The title compound was prepared essentially as described in Example 1 using 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2,6-difluoro-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoic acid methyl ester. The product was purified by reverse phase flash chromatography using a gradient of 30 to 50% ACN / NH4HCO3 in water (10 mM, pH 10). ES / MS m / z 585 (M+H), 583 (MH).

[0245] Example 5

[0246] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-3-methyl-phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid

[0247]

[0248] The title compound was prepared essentially as described in Example 2 using methyl 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]-2-fluoro-3-methyl-phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate. The product was purified by reverse phase flash chromatography using a 5 to 40% ACN / 5% NH4HCO3 aqueous gradient. ES / MS m / z 581 (M+H), 579 (MH).

[0249] Example 6

[0250] 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridyl]phenyl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylic acid

[0251]

[0252] The title compound was prepared essentially as described in Example 1 using methyl 4-[[2-[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridinyl]phenyl]acetyl]amino]-3-[[(2S)-oxetan-2-ylmethyl]amino]benzoate. The crude product was purified by reverse phase flash chromatography using a 20 to 35% ACN / 5% NH4HCO3 aqueous gradient. ES / MS m / z 549.0 (M+H), 547.1 (MH).

[0253] Biological Detection

[0254] Human GLP-1 receptor cAMP detection in HEK293 cells

[0255] GLP-1 receptor functional activity was determined using cAMP formation in HEK293 clonal cell lines expressing human GLP-1R (NCBI accession number NP_002053) at expression densities of 581 ± 94 (n = 6) and 104 ± 12 (n = 5) fmol / mg protein (using [ 125 I] GLP-1(7-36)NH2 homologous competition binding assay). In a 20 µl assay volume (final DMSO concentration of 0.5%), 1X GlutaMAX TMhGLP-1R receptor expressing cells were treated with compounds (20 point concentration response curve in DMSO, 2.75x direct dilution in Labcyte Echo, 384 well plates Corning Cat# 3570) in DMEM (Gibco Cat# 31053) containing 1% (Gibco Cat# 35050), 0.1% bovine casein (Sigma C4765-10ML), 250 µM IBMX (3-isobutyl-1-methylxanthine, Acros Cat# 228420010) and 20 mM HEPES (Gibco Cat# 15630). After incubation for 30 minutes at 37°C, the resulting increase in intracellular cAMP was quantified using the CisBio cAMP Dynamic 2 HTRF Assay Kit (62AM4PEJ). Briefly, the cells were lysed by adding cAMP-d2 conjugate (10 µL) in cell lysis buffer followed by the addition of antibody anti-cAMP-Eu 3+ -cryptate (also in cell lysis buffer) (10 µL) to measure intracellular cAMP levels. The resulting competitive assay was incubated at room temperature for at least 60 minutes and then analyzed using the PerkinElmer Envision ® The instrument detects at 320 nm excitation and 665 nm and 620 nm emission. Envision units (emission at 665 nm / 620 nm*10,000) are inversely proportional to the amount of cAMP present and are converted to nM cAMP per well using a cAMP standard curve. The amount of cAMP generated in each well (nM) was converted to a percentage of the maximal response observed with human GLP-1(7-36)NH2. Relative EC values ​​were derived by nonlinear regression analysis using the percentage of maximal response vs. the concentration of compound added. 50 Value and percent top (E max ), fitted to a four-parameter logistic equation. EC values ​​for compounds of Examples 1-6 when tested in the above cAMP assay using HEK293 cells expressing 581 and 104 fmol / mg GLP-1R 50 and E max The data are shown in Tables 2 and 3, respectively. These data indicate that the compounds of Examples 1-6 are agonists of the human GLP-1 receptor.

[0256] Table 2. Intracellular cAMP response of HEK293 cell line with GLP-1R expression density of 581 fmol / mg

[0257]

[0258] Table 3. Intracellular cAMP response of HEK293 cell line with GLP-1R expression density of 104 fmol / mg

[0259]

[0260] In vivo intraperitoneal glucose tolerance test in human GLP-1R knock-in mice

[0261] Use from mice Glp-1r The efficacy of the example compounds in reducing blood glucose concentration in vivo was determined in mice expressing human GLP-1R (NCBI accession number NP_002053) (Jun, LS et al., PLoS One. 2014 9:e93746). The mice fasted overnight were orally administered with a solution of 10% Kolliphor. ® (HS15) / polyethylene glycol 400 (PEG400) in a 1:1 ratio. One hour after dosing, animals were given glucose by intraperitoneal injection (2 g / kg), and blood glucose levels were measured intermittently over the next two hours using a glucometer. A range of doses of the test compound was delivered, and the calculated area under the curve was determined for each dose group and fitted to a four-parameter logistic model as the ED with 95% confidence intervals. 50 Calculation of in vivo efficacy. When tested in the above-described in vivo intraperitoneal glucose tolerance test, the compounds of Examples 1-3 exhibited efficacy in lowering blood glucose concentrations in mice expressing human GLP-1R, ED 50 The (and 95% confidence interval) values ​​are shown in Table 4, indicating that these compounds are potent and orally available GLP-1R agonists in mice.

[0262] Table 4. Hypoglycemic efficacy in mice expressing human GLP-1R

[0263] Example <![CDATA[Hypoglycemic ED 50 (mg / kg)]]> 95% Confidence Interval 1 0.09 0.0301-0.2592 2 0.07 0.0246-0.1808 3 0.06 0.013-0.246

[0264] Non-human primate (NHP) pharmacokinetics:

[0265] The test compounds were administered intravenously (IV) at 0.5 mg / kg (using a dose volume of 1 mL / kg) to fasted male cynomolgus monkeys. Serial blood samples were collected at 0.08, 0.25, 0.5, 1, 2, 4, 8, 12 and 24 hours after the IV bolus administration. After treatment with EDTA coagulant, plasma was obtained by centrifugation and stored at -70°C until analysis by LC-MS / MS. The test article concentration in plasma was determined. Plasma clearance and steady-state distribution volume were calculated using non-compartmental analysis. Table 5 shows the pharmacokinetic data for the compounds of Examples 1-3 in this assay. These data were used in part to report human mechanistic PK projections, which suggest that the human pharmacokinetic profile supports once-daily dosing.

[0266] Table 5. Pharmacokinetic data in cynomolgus monkeys

[0267] Example Plasma clearance (mL / min / kg) Distribution volume (L / kg) vehicle* 1 13 1.2 A 2 11 1.1 A 3 6 1.1 B

[0268] *Vehicle A - 5% DMSO and 95% (20% CAPTISOL® (w / v) in water); Vehicle B - 20% captisol (w / v) in water + 1 molar equivalent of NaOH.

[0269] Phosphodiesterase 10 (PDE10) enzyme activity assay

[0270] To generate the phosphodiesterase 10A1 (PDE10A1) protein, the full-length PDE10A1 clone corresponding to GenBank ID AAD32595.1 was cloned into pFastBac1 (Invitrogen). The PDE10A1 protein with a C-terminal FLAG-tag was expressed by baculovirus infection of insect cells, purified using anti-FLAG M2-agarose (Sigma) and size exclusion chromatography on a Superdex 200 column (GE Healthcare), and stored in small aliquots (20 mM Tris-HCl, Ph 7.5, 150 mM NaCl, 10% glycerol) at −80°C.

[0271] PDE10A1 enzyme activity was measured using a yttrium silicate-based scintillation proximity assay (SPA) that detects radioactive monophosphate nucleotides but not cyclic monophosphates. The assay buffer consisted of 50 mM Tris-HCl pH 7.5, 8 mM MgCl2, 3.4 mM EDTA, and 0.1% BSA (Sigma). The assay was performed in a 384-well plate (3706, Corning) in a total volume of 50 μl: 24 μl PDE10A1 enzyme, 1 μl test compound, and 25 μl cyclic nucleotides. Test compounds were diluted in neat DMSO using a 10-point concentration response curve with a 3-fold dilution factor and 1 μl was acoustically dispensed into the assay plate using an Echo555 (LabCyte). 24 μl PDE10A1 protein was incubated with 1 μl compound for 30 min and then quenched by the addition of [8- 3 The reaction was initiated with the substrate [H]-cGMP (6.5 Ci / mmol, Perkin Elmer). The final concentrations of the components in the assay buffer were 70 pM PDE10A1, 80 nM ( 3 H -cGMP) and 2% DMSO. The maximum compound concentration in the reaction mixture was 10 μM. The reaction was incubated at room temperature for 60 min, then quenched and 400 mg of SPA beads (RPNQ0150, Perkin Elmer) were added per well. Bead-bound radioactivity (product) was quantified after 12 h using a Microbeta counter (Perkin Elmer). Data were normalized to % inhibition and IC was calculated using the 4-parameter logistic equation as described 50 Values ​​(Campbell, RM; Dymshitz, J.; Eastwood, BJ; et al. "Data Standardization for Results Management.": Sittampalam, GS; Grossman, A.; Brimacombe, K.; et al., eds. Assay Guidance Manual . Bethesda (MD): Eli Lilly & Company and the National Center for Advancing Translational Sciences; 2004.). Table 6 shows the activity of the compounds of Examples 1-4 in this assay. These data indicate that the compounds of Examples 1 to 4 have weak binding affinity for PDE10A, which suggests a reduced risk of toxicity.

[0272] Table 6. In vitro potency of PDE10A1 inhibition

[0273] Example <![CDATA[IC 50 (µM),n = 1]]> 1 >10 2 7.43 3 >10 4 5.41

[0274] human hERG K + Channel affinity radioligand binding assay

[0275] Compounds were evaluated for their effects on human hERG Kappa B in transfected HEK-293 cells in a radioligand binding assay as described herein. + The cell membrane homogenate (approximately 40 μg of protein) was incubated with 3 nM [ 3 H] Dofetilide was incubated for 60 minutes. The assay was performed in a 96-well plate format with a volume of 200 μL, containing up to 1% DMSO from the initial solubilization of the test compound. After incubation, the sample was quickly filtered under vacuum through a 0.3% PEI pre-soaked glass fiber filter (GF / B, Packard) and rinsed several times with ice-cold 50 mMTris-HCl, 10 mM KCl, and 1 mM MgCl2 using a 96-sample cell collector (Unifilter, Packard). The filter was dried and then radioactivity was counted using scintillation cocktail (Microscint 0, Packard) in a scintillation counter (Topcount, Packard). Table 7 shows the activity of Examples 1-3 in this assay, expressed as a percentage of inhibition of specific binding of the control radioligand. These data show that the compounds of Examples 1 to 3 have weak hERG inhibitory activity, which indicates a reduced risk of toxicity.

[0276] Table 7. Human hERG K + Channel affinity Radioligand inhibition percentage

[0277] Example Percent inhibition (%) at 100 µM compound concentration, n = 1 1 0 2 54 3 37

Claims

1. A compound or a pharmaceutically acceptable salt thereof, wherein the compound is selected from:

2. A compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein the compound is selected from:

3. The compound according to claim 2 or a pharmaceutically acceptable salt thereof, wherein the compound is:

4. The compound according to claim 3, which is the tert-butylamine salt of the compound of the formula:

5. A compound which is:

6. A pharmaceutical composition comprising a compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof and at least one pharmaceutically acceptable carrier, diluent or excipient.

7. A compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, for use in therapy.

8. A compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, for use in the treatment of type II diabetes.

9. A compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, for use in lowering blood glucose levels.

10. A compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, for use in the treatment of hyperglycemia.

11. The compound according to any one of claims 7 to 10, or a pharmaceutically acceptable salt thereof, wherein the compound is administered orally.

12. Use of a compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating type II diabetes.

13. Use of a compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for lowering blood sugar levels.

14. Use of a compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating hyperglycemia.

15. Use according to any one of claims 12 to 14, wherein the compound is administered orally.

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