Novel cyanopyridine KHK inhibitor compounds

Abstract

The present invention encompasses compounds of formula (I), wherein the substituents R1 to R4 have the meanings given in the claims and herein, their use as inhibitors of KHK inhibitors, pharmaceutical compositions comprising compounds of this type, and their use as medicaments, in particular as agents for the treatment and / or prevention of NAFLD, NASH and type 2 diabetes. JPEG2025539698000087.jpg59150

Description

[Technical Field]

[0001] The present invention relates to novel cyanopyridine compounds of formula (I) and derivatives which inhibit ketohexokinase (KHK), wherein the substituents R1 to R4 have the meanings given in the claims and herein, their use as inhibitors of KHK, pharmaceutical compositions comprising compounds of this type, and their use as medicaments, in particular as agents for the treatment and / or prevention of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH) and type 2 diabetes mellitus (T2DM). [ka] [Background technology]

[0002] Ketohexokinase (KHK) catalyzes the phosphorylation of fructose to fructose-1-phosphate (F-1-P). The enzymatic activity of human KHK-C in the liver leads to increased synthesis of fatty acids and triglycerides, along with increased fructose consumption. Fructose metabolism by KHK is thought to contribute to numerous diseases, such as NAFLD, NASH, and T2DM.

[0003] Haurd et al. (J. Med. Chem. 2017, 60, 7835-7849) describe KHK inhibitors with a cyanopyridine nucleus, such as 6-[(3S,4S)-3,4-dihydroxy-1-pyrrolidinyl]-2-[(3R)-3-hydroxy-3-methyl-1-pyrrolidinyl]-4-(trifluoromethyl)-3-pyridinecarbonitrile (CAS Registry Number 2711012-28-9).

[0004] It is an object of the present invention to provide additional cyanopyridine compounds that are more potent KHK inhibitors.

[0005] In the present invention, "KHK inhibitor" means a compound that inhibits the enzymatic activity of human ketohexokinase-C, as described below.

[0006] Detailed Description of the Invention It has been surprisingly found that the compounds of formula (I), in which the substituents R1 to R4 have the following meanings, act as inhibitors of KHK and can inhibit the production of fructose-1-phosphate in cells. Thus, the compounds of the present invention can be used, for example, for the treatment of NAFLD, NASH, or T2DM.

[0007] Thus, the present invention relates to a compound of formula (I) or a salt thereof: [ka] (In the formula, R1 is [ka] and R2 is [ka] selected from the group consisting of where: [ka] teeth, [ka] NH2 in etc. [ka] NHMe in etc. [ka] NMe2 in etc. [ka] Hydroxymethyl in, for example, [ka] Hydroxy (OH) in and is substituted with one or two substituents selected from R3 is [ka] selected from the group consisting of Here, R5 is [ka] Me or Et, optionally substituted with up to three F, such as R4 is [ka] is selected from the group consisting of

[0008] In particular, R1 is preferably [ka] is.

[0009] In particular, R2 is preferably [ka] and 3-dimethylamino-pyrrolidin-1-yl such as

[0010] In particular, R3 is preferably [ka] is.

[0011] In particular, R4 is preferably [ka] is.

[0012] The present invention is directed to compounds of formula (I) or salts thereof, which are useful in the prevention and / or treatment of diseases and / or conditions in which the inhibition of ketohexokinase is of therapeutic benefit, including, but not limited to, the treatment of NAFLD, NASH, and T2DM. Accordingly, in another aspect of the invention, compounds of formula (I) or pharmaceutically acceptable salts thereof are used as medicaments, and the invention also relates to compounds of formula (I) or pharmaceutically acceptable salts thereof for use in methods of treatment of the human or animal body.

[0013] Of particular interest is the use of the compound of formula (I) or one of its salts in the treatment of NAFLD, NASH and T2DM, in particular in the treatment of NASH. Accordingly, another aspect of the present invention is the use of the compound of formula (I) or a pharmaceutically acceptable salt thereof for preparing a pharmaceutical composition comprising at least one compound of formula (I) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier for the treatment of NAFLD, NASH and T2DM. Particularly preferred is their use in the preparation of a pharmaceutical composition for the treatment of NASH in the human or animal body.

[0014] For example, in the treatment of diseases such as NAFLD, NASH, and T2DM, the compounds of the present invention may be administered before, after, or together with other pharmaceutically active substances.

[0015] The present invention further relates to pharmaceutically active salts of compounds of formula (I) with inorganic or organic acids or bases. It is expected that pharmaceutically acceptable salts of compounds of formula (I) and / or co-crystals of compounds of formula (I), preferably pharmaceutically acceptable co-crystals, may be formed.

[0016] It is expected that some of the compounds of formula (I) disclosed herein may form hydrates, solvates, polymorphs, metabolites, derivatives, isomers, or prodrugs. Some have chiral centers.

[0017] formulation Suitable formulations for administering the compounds of the present invention will be apparent to those skilled in the art and include, for example, tablets, pills, capsules, suppositories, lozenges, troches, liquids, elixirs, syrups, sachets, emulsions, inhalants, or dispersible powders. Preferred liquids are injection solutions (subcutaneous, intravenous, intramuscular) or infusion solutions (injectable).

[0018] The content of the pharmaceutically active compound should be sufficient to achieve the dosage range specified below, for example, 0.1 to 90% by weight, preferably 0.5 to 50% by weight, based on the total weight of the composition. The specified dosage can be given multiple times a day if necessary.

[0019] Suitable tablets can be obtained, for example, by mixing the active substance of the present invention with known excipients, such as inert diluents, carriers, disintegrants, adjuvants, surfactants, binders, and / or lubricants.

[0020] Coated tablets can be conveniently prepared by coating a core, manufactured in the same manner as a tablet, with a substance commonly used for tablet coating, such as Kollidon or shellac, gum arabic, talc, titanium dioxide, or sugar. The core can consist of multiple layers to achieve delayed release or to prevent incompatibilities. Similarly, tablet coatings can consist of multiple layers to achieve delayed release and may use excipients as described above for tablets.

[0021] Syrups and elixirs containing the active substances or combinations thereof according to the invention may further contain a sweetener such as saccharin, cyclamate, glycerol or sugar and a flavor enhancer, for example, vanillin or orange extract. They may also contain a suspending aid or thickener such as sodium carboxymethylcellulose, a humectant such as a condensation product of a fatty alcohol with ethylene oxide, or a preservative such as p-hydroxybenzoate.

[0022] Solutions for injection and infusion are prepared by conventional methods, for example, by adding an isotonicity agent, a preservative such as p-hydroxybenzoate, or a stabilizer such as an alkali metal salt of ethylenediaminetetraacetic acid, and optionally, an emulsifier and / or a dispersant. On the other hand, when water is used as a diluent, for example, an organic solvent may optionally be used to solvate the drug or dissolve the acid, and the solution may be transferred into an injection vial or ampule, or an infusion bottle.

[0023] Capsules can be prepared, for example, by mixing the active substance with an inert carrier such as lactose or sorbitol and packing them into gelatin capsules.

[0024] Suitable suppositories can be prepared by mixing with carriers provided for this purpose, such as, for example, neutral fats or polyethylene glycol or its derivatives.

[0025] Excipients that can be used include, for example, water; pharmaceutically acceptable organic solvents, such as paraffins (e.g., petroleum fractions), vegetable oils (e.g., peanut or sesame oil), mono- or polyfunctionalized alcohols (e.g., ethanol or glycerol); carriers, such as natural mineral powders (e.g., kaolin, clay, talc, or chalk), synthetic mineral powders (e.g., highly dispersed silicic acid and silicates), sugars (e.g., sucrose, lactose, and glucose), emulsifiers (e.g., lignin, spent sulfite pulp liquor, methylcellulose, starch, and polyvinylpyrrolidone); and lubricants (e.g., magnesium stearate, talc, stearic acid, sodium lauryl sulfate).

[0026] The preparation is administered by a conventional method, preferably orally or transdermally, most preferably orally.For oral administration, tablets can naturally contain additives such as sodium citrate, calcium carbonate, and dicalcium phosphate, in addition to the above-mentioned carriers, together with various additives such as starch, preferably potato starch, gelatin, etc.In addition, lubricants such as magnesium stearate, sodium lauryl sulfate, and talc can be used at the same time in the tableting process.In the case of aqueous suspension, the active substance can be combined with various flavor enhancers or colorants in addition to the above-mentioned excipients.

[0027] For parenteral use, a solution of the active substance and a suitable liquid carrier can be used.

[0028] The applicable daily dosage range of the compounds of formula (I) is usually from 1 mg to 2000 mg, preferably from 1 to 1000 mg.

[0029] For intravenous administration, the dosage ranges from 1 mg to 1000 mg at various infusion rates, preferably between 5 mg and 500 mg at various infusion rates.

[0030] It may sometimes be necessary to vary the specified amount depending on the body weight, age, route of administration, severity of the disease, individual response to the drug, nature of the formulation, and the time or interval at which the drug is administered (single or multiple doses per day, continuous or intermittent treatment). Thus, in some cases, a dose lower than the minimum dose mentioned above may be sufficient, while in other cases the upper limit may have to be exceeded. When administering large amounts, it may be advisable to divide them into several smaller doses throughout the day.

[0031] As used herein, "pharmaceutically acceptable salts" refers to derivatives of the disclosed compounds where the parent compound is modified by making acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like.

[0032] For example, such salts include salts from benzenesulfonic acid, benzoic acid, citric acid, ethanesulfonic acid, fumaric acid, gentisic acid, hydrobromic acid, hydrochloric acid, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, 4-methylbenzoic acid, phosphoric acid, salicylic acid, succinic acid, sulfuric acid, and tartaric acid.

[0033] Additionally, pharmaceutically acceptable salts can be formed with cations derived from ammonia, L-arginine, calcium, 2,2'-iminobisethanol, L-lysine, magnesium, N-methyl-D-glucamine, potassium, sodium, and tris(hydroxymethyl)-aminomethane.

[0034] The pharmaceutically acceptable salts of the present invention can be synthesized from a parent compound that contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or free base form of these compounds with a sufficient amount of the appropriate base or acid in water or an organic diluent such as ether, ethyl acetate, ethanol, isopropanol, or acetonitrile, or a mixture thereof.

[0035] Abbreviation [Table 1]

[0036] The features and advantages of the present invention will become apparent from the following detailed examples which illustrate the basis of the invention by way of example, without limiting its scope.

[0037] Preparation of Compounds of the Invention General Unless otherwise stated, all reactions are carried out using commercially available equipment and methods commonly used in chemical laboratories. Air- and / or moisture-sensitive starting materials are stored under protective gas, and the corresponding reactions and accompanying manipulations are carried out under protective gas (nitrogen or argon).

[0038] The compounds of the present invention are named according to IUPAC rules using MarvinSketch software (Chemaxon). In the event of a discrepancy between the structure and the name given to a compound, the structure takes precedence.

[0039] Chromatography Unless otherwise indicated, intermediates and final compounds were purified by preparative normal phase chromatography on silica gel using appropriate organic solvents.

[0040] NMR spectra were recorded on a Bruker BBFO ULTRASHIELD™ 300 AVANCE III 300 MHz or a Bruker BBFO ASCEND™ 400 AVANCE III 400 MHz and were recorded in the solvents indicated.

[0041] Analytical LCMS method: HPLC method A: Column: Sunfire C18 3.0x30mm 2.5μm Column producer: Water

[0042] [Table 2]

[0043] HPLC method B: Column: Sunfire C18 3.0x30mm 2.5μm Column producer: Water [Table 3]

[0044] HPLC method C: Column: Sunfire C18_2.1x30mm_2.5μm Column producer: Water [Table 4]

[0045] HPLC method D: Column: XBridge C18_3.0x30mm_2.5μm Column producer: Water [Table 5]

[0046] HPLC method E: Column: Sunfire C18 3.0x30mm 2.5μm Column producer: Water [Table 6]

[0047] HPLC method F: Column: XBridge C18_3.0x30mm_2.5μm Column producer: Water [Table 7]

[0048] HPLC method G: Column: Sunfire C18 3.0x30mm 2.5μm Column producer: Water [Table 8]

[0049] HPLC method H: Column: XBridge C18_3.0x30mm_2.5μm Column producer: Water [Table 9]

[0050] HPLC method I: Column: Sunfire C18 3.0x30mm 2.5μm Column producer: Water [Table 10]

[0051] The compounds of the present invention are prepared by the synthetic methods described below, and the substituents of the general formula have the meanings mentioned above.These methods are intended to be illustrative of the present invention, and the scope of the subject matter of the present invention and the compounds described in the claims is not limited to these examples.If the preparation of starting compounds is not described, they are commercially available or can be prepared in analogy with known compounds or methods described herein.Substances described in the literature are prepared according to published synthetic methods.

[0052] The syntheses described below may produce each compound in the salt form shown (eg, free base or acid), or in a different salt form (eg, TFA salt).

[0053] A. General Methods for Preparing Compounds of Formula (I) Scheme 1: General synthetic route to compound (I) [ka]

[0054] Compounds of general formula (I) can be prepared by reacting chloropyridine compounds of general formula (II) with anilines of general formula (III) under standard cross-coupling conditions, using a palladium catalyst, such as XPhos Pd G2, and a base, such as cesium carbonate, in a solvent, such as dioxane. The substituents R3 and R4 of aniline (III) may be outside the scope of the present invention and may subsequently be converted (e.g., by removing protecting groups) to substituents within the scope of the present invention. Compounds of general formula (II) can be prepared from dichloropyridine compounds of general formula (IV) through reaction with a suitable primary or secondary amine (V) in the presence of a base, such as N,N-diisopropylethylamine, in a solvent, such as ethanol. The structure of the applied amine (V) may be outside the scope of the present invention and may subsequently be converted (e.g., by removing protecting groups) to structure R2 within the scope of the present invention. Dichloropyridine compounds of general formula (IV) can be prepared by methods known in the literature or by the methods described below.

[0055] B. Synthesis of Intermediates B.1 Synthesis of dichloropyridine intermediates Intermediate I.1: 2,6-dichloro-4-(fluoroethyl)pyridine-3-carbonitrile [ka]

[0056] Step 1: Lithium diisopropylamide (100 ml; 2 mol / l in THF; 200 mmol) was cooled to -70°C. Acetic acid ethyl ester (19.6 ml; 200 mmol) was added, and the mixture was stirred at -70°C for 1 hour. Ethyl fluoroacetate (17.5 ml; 180 mmol) was added dropwise, and the mixture was stirred for a further 45 minutes, after which it was allowed to warm to ambient temperature. Ethyl acetate (100 ml) was added, and the mixture was acidified by the addition of aqueous HCl (4 mol / l). The organic layer was separated, washed with brine, dried over magnesium sulfate, and evaporated to dryness to give the crude product, ethyl 4-fluoro-3-oxobutanoate, as a brownish oil (24.7 g; 93%), which was carried on to the next step without further purification.

[0057] Step 2: To a mixture of the crude product from step 1 (24.7 g) and cyanoacetamide (14.0 g; 167 mmol) in ethanol (100 ml) was added KOH (9.36 g; 167 mmol). The mixture was refluxed overnight, after which the precipitate that formed was filtered off with suction. The solid was dissolved in warm water (30 ml) and acidified by adding aqueous HCl (4 mol / l). The precipitate was filtered off with suction, washed with water, and dried at 50 °C to give 2,6-dihydroxy-4-(fluoromethyl)pyridine-3-carbonitrile (18.7 g; 111 mmol; 67%) as a slightly brownish solid. MS (ESI pos.+neg. Loop-Inj.) m / z: 169 [M + H] +

[0058] Step 3: In a closed vessel, a mixture of 2,6-dihydroxy-4-(fluoromethyl)pyridine-3-carbonitrile (6.0 g; 35.7 mmol) from step 2 and phosphoryl chloride (50 ml; 546 mmol) was heated to 140° C. overnight and then cooled to ambient temperature. The mixture was added dropwise to warm water (400 ml), the temperature of which did not exceed 50° C. Caution! Exothermic! The resulting mixture was cooled to 0° C. and the precipitate that formed was filtered off with suction, washed with water and dried at ambient temperature to give 2,6-dichloro-4-(fluoromethyl)pyridine-3-carbonitrile (4.81 g; 23.5 mmol; 66%) as a brownish powder. 1 H NMR (400 MHz, DMSO-d6) δ ppm 7.87 (s, 1 H), 5.72 (d, J=45.73 Hz, 2 H) MS (ESI pos.+neg. Loop-Inj.) m / z: 203 [M + H] -

[0059] Intermediate I.2: 2,6-Dichloro-4-(difluoromethyl)pyridine-3-carbonitrile [ka] Prepared analogously to the procedure described for the synthesis of intermediate I.1, starting from 4-difluoro-3-oxobutanoate and applying steps 2 and 3. 1 H NMR (400 MHz, DMSO-d6) δ ppm 8.09 (s, 1 H), 7.29 (t, J=53.05 Hz, 1 H) MS (ESI pos.+neg. Loop-Inj.) m / z: 221 [M + H] - m / z: 203 [M+HO-HCl-H] -

[0060] B.2 Synthesis of aniline intermediates Intermediate II.1: Methyl 2-[3-amino-4-(methylsulfanyl)phenyl]acetate [ka]

[0061] Step 1: To a stirred mixture of 2-(4-fluoro-3-nitrophenyl)acetic acid (6.00 g; 30 mmol) in DMF (30 mL) was added NaOH (1.33 g; 33 mmol) and then water (8.0 mL) at room temperature. Methyl mercaptan sodium salt (15% in water; 2.32 g; 33 mmol) was added dropwise to the reaction mixture, and the temperature was raised to 50 °C. After 30 minutes, 20 mL of cold water and then 20 mL of 4 M HCl solution were added to the reaction mixture, and the resulting yellow solid was collected by filtration, washed with water, and dried under reduced pressure at 50 °C to give 2-[4-(methylsulfanyl)-3-nitrophenyl]acetic acid as a yellow solid (6.00 g; 22 mmol; 88%).

[0062] Step 2: To a solution of the product obtained in step 1 in methanol (100 ml) was added concentrated sulfuric acid (1.2 ml) at room temperature. The mixture was stirred at reflux temperature for 6 hours, after which the volatiles were evaporated. The resulting mixture was diluted with ethyl acetate and washed first with saturated sodium bicarbonate solution, then with brine, and dried over magnesium sulfate. The filtered organic layer was evaporated to dryness to give crude methyl 2-[4-(methylsulfanyl)-3-nitrophenyl]acetate (7.0 g), which was carried on to the next step without purification.

[0063] Step 3: To a degassed solution of the crude product from step 2 (6.0 g; 25 mmol) in methanol (40 ml) was added 10% palladium on carbon (2.65 g). The mixture was stirred in a Parr shaker under 50 psi hydrogen pressure for 12 hours. The reaction mixture was filtered through Celite and concentrated under reduced pressure. The crude product was purified by silica gel chromatography (DCM / methanol) to give methyl 2-[3-amino-4-(methylsulfanyl)phenyl]acetate (1.50 g; 7.10 mmol; 29%) as a colorless liquid. MS (ESI pos.+neg. Loop-Inj.) m / z: 212 M + H] +

[0064] Intermediate II.2: Ethyl 3-[3-amino-4-(methylsulfanyl)phenyl]-2-methylpropanoate [ka]

[0065] Step 1: To a solution of 4-chloro-3-nitrobenzyl alcohol (11.0 g; 58.6 mmol) in DMF (250 ml) was slowly added sodium methanethiolate (5% in water; 49.8 ml; 117 mmol). The mixture was stirred overnight, after which water was added and the mixture was acidified by the addition of aqueous HCl. The precipitate that formed was filtered off with suction and dried at 50 °C to give [3-amino-4-(methylsulfanyl)phenyl]methanol (9.50 g; 81%) as a yellow solid. MS (ESI pos.+neg. Loop-Inj.) m / z: 200 [M + H] + RT=0.51min (HPLC method C)

[0066] Step 2: The product of step 1 (9.50 g; 37.7 mmol) was dissolved in thionyl chloride (100 ml). The mixture was stirred at ambient temperature for 3 hours, then evaporated to dryness and co-evaporated with diethyl ether to give 5-(chloromethyl)-2-(methylsulfanyl)aniline (10.0 g; 96%) as a yellow solid. RT=0.77min (HPLC method C)

[0067] Step 3: Under argon, to a solution of sodium hydride (60% in mineral oil; 4.04 g; 101 mmol) in DMF (50 ml) was added dropwise a solution of diethyl methylmalonate (15.7 ml; 92 mmol) in DMF (30 ml) while cooling in an ice bath. After stirring for a further 10 min, a solution of the product of step 2 (10.0 g; 46 mmol) in DMF (20 ml) was added dropwise. The mixture was stirred at ambient temperature for 1 h, then poured into ice water and extracted with DCM. The organic layer was separated, washed with brine, dried over magnesium sulfate, filtered, and evaporated to give 1,3-diethyl 2-methyl-2-{[4-(methylsulfanyl)-3-nitrophenyl]methyl}propanedioate (16.6 g; 86%) as a dark brown oil. 1 H NMR (400 MHz, DMSO-d6) δ ppm 8.00 (d, J=1.27 Hz, 1 H), 7.49 - 7.56 (m, 2 H), 4.10 - 4.19 (m, 4 H), 3.22 (s, 2 H), 2.52 (s, 3 H), 1.27 (s, 3 H), 1.19 (t, J=7.10 Hz, 6 H) MS (ESI pos.+neg. Loop-Inj.) m / z: 356 [M + H] + RT=1.13min (HPLC method A)

[0068] Step 4: To the product of step 3 (3.24 g; 9.11 mmol) in THF (40 ml) was added Raney nickel (400 mg). The mixture was shaken under 50 psi of hydrogen until TLC showed complete consumption of the starting material. The catalyst was removed by filtration and the filtrate was evaporated to give 1,3-diethyl 2-{[3-amino-4-(methylsulfanyl)phenyl]methyl}-2-methylpropanedioate (2.79 g; 94%). 1H NMR (400 MHz, DMSO-d6) δ ppm 7.07 (d, J=7.83 Hz, 1 H), 6.43 (d, J=1.52 Hz, 1 H), 6.29 (dd, J=7.83, 1.77 Hz, 1 H), 5.11 (s, 2 H), 4.13 (q, J=7.07 Hz, 4 H), 2.95 (s, 2 H), 2.28 (s, 3 H), 1.21 (s, 3 H), 1.18 (t, J=7.07 Hz, 6 H) MS (ESI pos.+neg. Loop-Inj.) m / z: 326 [M + H] + RT=1.02 min (HPLC Method A)

[0069] Step 5: A mixture of the product of step 4 (2.72 g; 8.58 mmol), dioxane (30 mL), and aqueous HCl (4 mol / L; 15 mL) was refluxed for 24 h. The mixture was evaporated and co-evaporated with added toluene to give crude 3-[3-amino-4-(methylsulfanyl)phenyl]-2-methylpropionic acid (2.34 g; 121%), which was carried on to the next step without purification.

[0070] Step 6: To a solution of the crude product from step 5 (3.14 g; 13.9 mmol) in ethanol (40 ml) was added thionyl chloride (1.98 g; 1.21 ml; 16.7 mmol) dropwise. Caution: Exothermic reaction! The mixture was stirred overnight, evaporated, dissolved in ethyl acetate, and extracted with potassium carbonate (10% aqueous solution). The aqueous layer was re-extracted with ethyl acetate. The combined organic layers were extracted with water, then brine, separated, dried over magnesium sulfate, filtered, and evaporated. The crude product was purified by silica gel chromatography (petroleum ether / ethyl acetate 5%->25%) to give ethyl 3-[3-amino-4-(methylsulfanyl)phenyl]-2-methylpropanoate (1.08 g; 31%) as a brownish oil. 1H NMR (400 MHz, DMSO-d6) δ ppm 7.10 (d, J=7.86 Hz, 1 H), 6.51 (d, J=1.77 Hz, 1 H), 6.36 (dd, J=7.86, 1.90 Hz, 1 H), 5.11 (s, 2 H), 4.02 (q, J=7.05 Hz, 2 H), 2.69 - 2.77 (m, 1 H), 2.57 - 2.68 (m, 1 H), 2.45 - 2.51 (m, 1 H), 2.26 (s, 3 H), 1.12 (t, J=7.10 Hz, 3 H), 1.05 (d, J=6.84Hz, 3H) MS (ESI pos.+neg. Loop-Inj.) m / z: 254 [M + H] + RT=0.97 min (HPLC Method A)

[0071] Intermediate III. 3-[3-amino-4-(methylsulfanyl)phenyl]propionic acid [ka]

[0072] Step 1: To a solution of 4-chloro-3-nitrocinnamic acid (1.00 g; 8.79 mmol) in DMF (30 ml) was added sodium hydroxide (387 mg; 9.67 mmol). The mixture was stirred until all the NaOH dissolved, then sodium methanethiolate (15% in water; 4.11 ml; 9.67 mmol) was added and the mixture was stirred for a further 2 h. Ice water (30 ml) was added, then the mixture was acidified by adding aqueous HCl (6 mol / l; 3.5 ml). The formed precipitate was filtered off with suction, washed with water and dried at 50 °C to give 3-[4-(methylsulfanyl)-3-nitrophenyl]prop-2-enoic acid (2.05 g; 98%) as a yellow solid. MS (ESI pos.+neg. Loop-Inj.) m / z: 238 [M - H] - RT=0.94min (HPLC method A)

[0073] Step 2: Palladium on carbon (10%) was added to a solution of the product of step 1 (0.300 g; 1.25 mmol) in methanol (10 ml). The mixture was shaken overnight under 50 psi hydrogen pressure. The catalyst was suction filtered and the filtrate was evaporated to dryness. The residue was dissolved in methanol (10 ml) and Raney nickel (100 mg) was added. The mixture was shaken for 3 days under 50 psi hydrogen pressure. The catalyst was suction filtered and the filtrate was evaporated to dryness. The crude product was purified by preparative RP-HPLC (column: C18; mobile phase: water / ACN / TFA) to give the title compound (85 mg; 22%) as a slightly brownish solid. MS (ESI pos.+neg. Loop-Inj.) m / z: 212 [M - H] - RT=0.82min (HPLC method C) [Example]

[0074] C. Synthesis of Example Compounds Example 1 2-[3-({6-[cis-octahydropyrrolo[3,4-c]pyrrol-2-yl]-3-cyano-4-(trifluoromethyl)pyridin-2-yl}amino)-4-(methylsulfanyl)phenyl]acetic acid [ka]

[0075] Step 1 (Procedure A) To a solution of the dichloropyridine reagent 2,6-dichloro-4-(trifluoromethyl)nicotinonitrile (1.70 g; 7.05 mmol) in ethanol (50 mL) was added the amine reagent cis-N-BOC-hexahydropyrrolo[3,4-c]pyrrole (1.50 g; 7.05 mmol) and N,N-diisopropylethylamine (2.44 mL; 14.1 mmol). The mixture was stirred overnight at ambient temperature and then evaporated to dryness. The crude product was purified by preparative RP-HPLC (column: C18 X-bridge; mobile phase: water-ACN + 0.1% aqueous ammonia; 55 °C) to give tert-butyl 5-[6-chloro-5-cyano-4-(trifluoromethyl)pyridin-2-yl]-cis-octahydropyrrolo[3,4-c]pyrrole-2-carboxylate as a colorless solid. MS (ESI pos.+neg. Loop-Inj.) m / z: 417 [M + H] + RT=1.22min (HPLC method A)

[0076] Step 2 (Procedure B) To a mixture of tert-butyl 5-[6-chloro-5-cyano-4-(trifluoromethyl)pyridin-2-yl]-cis-octahydropyrrolo[3,4-c]pyrrole-2-carboxylate (from Step 1; 50 mg; 0.12 mmol), catalyst XPhos G2 (20 mg; 0.025 mmol), and cesium carbonate (110 mg; 0.34 mmol) in dioxane (2.0 mL) was added the aniline reagent intermediate II.1 (methyl 2-[3-amino-4-(methylsulfanyl)phenyl]acetate; 30 mg; 0.14 mmol). The mixture was stirred at 100 °C for 4 h, cooled to ambient temperature, diluted with DMF, filtered, and evaporated to dryness. The crude product was purified by preparative RP-HPLC (column: C18; mobile phase: water-ACN+TFA; 60 °C).

[0077] Step 3 (Acidic Deprotection, Procedure C) The product from step 2 was dissolved in DCM / TFA (3:1) and stirred at ambient temperature for 2 h. The mixture was evaporated to dryness to give methyl 2-[3-({6-[cis-octahydropyrrolo[3,4-c]pyrrol-2-yl]-3-cyano-4-(trifluoromethyl)pyridin-2-yl}amino)-4-(methylsulfanyl)phenyl]acetate as the TFA salt (29 mg; 40% over two steps). MS (ESI pos.+neg. Loop-Inj.) m / z: 492 [M + H] + RT=0.96min (HPLC method A)

[0078] Step 4 (Basic Deprotection, Procedure D) To a solution of methyl 2-[3-({6-[cis-octahydropyrrolo[3,4-c]pyrrol-2-yl]-3-cyano-4-(trifluoromethyl)pyridin-2-yl}amino)-4-(methylsulfanyl)phenyl]acetate (from Step 3; 25 mg; 0.041 mmol) in methanol (1.0 ml) was added aqueous NaOH (1 mol / l; 150 μl; 0.15 mmol). The mixture was stirred at ambient temperature overnight and then acidified by the addition of aqueous HCl (1 mol / l; 150 μl). The product was purified by preparative RP-HPLC (column: C18; mobile phase: water-ACN+TFA; 60 °C) to give 2-[3-({6-[cis-octahydropyrrolo[3,4-c]pyrrol-2-yl]-3-cyano-4-(trifluoromethyl)pyridin-2-yl}amino)-4-(methylsulfanyl)phenyl]acetic acid as the TFA salt (21 mg; 86%). 1H NMR (400 MHz, DMSO-d6) δ ppm 8.86 (br s, 2 H), 8.59 (s, 1 H), 7.95 (d, J=1.65 Hz, 1 H), 7.43 (d, J=7.98 Hz, 1 H), 7.06 (dd, J=8.05, 1.84 Hz, 1 H), 6.35 (s, 1 H), 3.65 - 3.74 (m, 2 H), 3.57 (s, 2 H), 3.40 - 3.55 (m, 4 H), 3.07 - 3.16 (m, 4 H), 2.40 (s, 3 H) MS (ESI pos.+neg. Loop-Inj.) m / z: 478 [M + H] + RT=0.92min (HPLC method A)

[0079] Example 2 3-(3-{[3-cyano-4-(difluoromethyl)-6-[(3S)-3-(dimethylamino)pyrrolidin-1-yl]pyridin-2-yl]amino}-4-(methylsulfanyl)phenyl)-2-methylpropionic acid [ka]

[0080] Step 1 was carried out according to procedure A, applying the dichloropyridine reagent 2,6-dichloro-4-(difluoromethyl)pyridine-3-carbonitrile and the amine reagent (3S)-N,N-dimethylpyrrolidin-3-amine to give 2-chloro-4-(difluoromethyl)-6-[(3S)-3-(dimethylamino)pyrrolidin-1-yl]pyridine-3-carbonitrile as a light brown solid. MS (ESI pos.+neg. Loop-Inj.) m / z: 301 [M + H] + RT=0.97min (HPLC method D)

[0081] Step 2 was carried out according to procedure B (but without chromatographic purification), applying the aniline reagent ethyl 3-[3-amino-4-(methylsulfanyl)phenyl]-2-methylpropionate (intermediate II.1).

[0082] The crude product from step 2 was carried on to step 3 and carried out according to procedure D to give 3-(3-{[3-cyano-4-(difluoromethyl)-6-[(3S)-3-(dimethylamino)pyrrolidin-1-yl]pyridin-2-yl]amino}-4-(methylsulfanyl)phenyl)-2-methylpropionic acid as the TFA salt (slightly brownish solid). MS (ESI pos.+neg. Loop-Inj.) m / z: 490 [M + H] + RT=0.88min (HPLC method A)

[0083] Examples 3 and 4 Chiral Separation of Example 2 (3-(3-{[3-cyano-4-(difluoromethyl)-6-[(3S)-3-(dimethyl-amino)pyrrolidin-1-yl]pyridin-2-yl]amino}-4-(methylsulfanyl)phenyl)-2-methylpropionic acid) The diastereomeric mixture Example 2 was separated by applying chiral SFC. Column: CHIRAL ART® Cellulose-SC 20x250mm 5μm Solvent: scCO2 75%; 2-propanol + 20 mM NH3 25% Back pressure regulator 150bar Temperature 40℃ Flow rate 60ml / min Sample concentration: 20mg / ml Sample solvent: methanol Injection volume 200 μl

[0084] The two isolated crude diastereomers were further purified by preparative RP-HPLC (column: C18; mobile phase: water-ACN+TFA; 60°C) and lyophilized to give the title compound as a TFA salt (slightly yellowish solid).

[0085] Optical purity was determined by analytical SFC. Column: CHIRAL ART® Cellulose-SC; Solvent: scCO2 75%; 2-propanol + 10 mM NH3 25%

[0086] Example 3 (first eluting diastereomer; RT=3.16 min): [ka]

[0087] The absolute configuration of the methyl substituent at the alpha position of the carboxylic acid is arbitrarily assigned, and therefore the pure diastereomeric Example 3 may have the structure shown or may have the structure shown in Example 4. MS (ESI pos.+neg. Loop-Inj.) m / z: 490 [M + H] + RT=0.90min (HPLC method A)

[0088] Example 4 (second eluting diastereomer; RT=3.61 min): [ka]

[0089] The absolute configuration of the methyl substituent at the alpha position of the carboxylic acid is arbitrarily assigned, therefore, the pure diastereomeric Example 4 may have the structure shown or may have the structure shown in Example 3. MS (ESI pos.+neg. Loop-Inj.) m / z: 490 [M + H] + RT=0.89min (HPLC method A)

[0090] Preparation of Example 4 in Salt-Free (Zwitterionic) Form Example 4 (TFA salt; 2.4 g) prepared as described above was dissolved in water-ethanol (3:1; 800 ml). The mixture was heated to 90°C and filtered hot. The pH was adjusted to 6-7 by adding aqueous NaOH (1 mol / L), after which the solution was cooled to ambient temperature and allowed to stand overnight. The precipitate was filtered, washed with water, and dried at 60°C to give 1.27 g (69%) of the zwitterionic form as an off-white, amorphous solid. 1 H NMR (400 MHz, DMSO-d6) δ ppm 8.43 (s, 1 H), 8.14 (s, 1 H), 7.41 (d, J=7.98 Hz, 1 H), 7.00 (t, J=54.20 Hz, 1 H), 6.95 (dd, J=7.98, 1.52 Hz, 1 H), 6.21 - 6.36 (m, 1 H), 3.53 - 3.88 (m, 3 H), 3.13 - 3.49 (m, 3 H), 2.78 - 2.94 (m, 2 H), 2.54 - 2.69 (m, 2 H), 2.38 (s, 3 H), 2.20 (s, 6 H), 1.05 (d, J=6.59 Hz, 3 H)

[0091] The following examples in Table 1a were prepared by analogous procedures from the starting materials shown in Table 1b, applying the procedures described above (see synthesis of Examples 1 and 2), in the reaction sequence shown.

[0092] [Table 1a-1]

[0093] [Table 1a-2]

[0094] [Table 1a-3]

[0095] Table 1a-4

[0096] Table 1a-5

[0097] Table 1a-6

[0098] Table 1a-7

[0099] Table 1a-8

[0100] Table 1a-9

[0101] Table 1a-10

[0102] Table 1a-11

[0103] Table 1a-12

[0104] Table 1a-13

[0105] [Table 1b-1]

[0106] [Table 1b-2]

[0107] [Table 1b-3]

[0108] [Table 1b-4]

[0109] [Table 1b-5]

[0110] [Table 1b-6]

[0111] [Table 1b-7]

[0112] [Table 1b-8]

[0113] [Table 1b-9]

[0114] Example 50 ({[3-({6-[cis-octahydropyrrolo[3,4-c]pyrrol-2-yl]-3-cyano-4-(trifluoromethyl)pyridin-2-yl}amino)-4-methylsulfanyl)phenyl]methoxy}(methyl)phosphinic acid [ka]

[0115] A mixture of Example 31 (100 mg; 0.222 mmol), methylphosphonic acid (21.4 g; 0.222 mmol), N,N'-dicyclohexylcarbodiimide (55.1 mg; 0.267 mmol), and a catalytic amount of DMAP in chloroform (10 ml) was gently refluxed overnight. The mixture was evaporated to dryness, and the crude product was purified by preparative RP-HPLC (column: C8; mobile phase: water / ACN / ammonia) to give the ammonium salt of the title compound (21 mg; 18%) as a colorless solid. MS (ESI pos.+neg. Loop-Inj.) m / z: 528 [M + H] + RT=0.79min (HPLC method D)

[0116] Examples 51 and 52 Chiral separation of Example 41 (3-[3-({3-cyano-6-[(3S)-3-(dimethylamino)pyrrolidin-1-yl]-4-(trifluoromethyl)pyridin-2-yl}amino)-4-methylsulfanyl)phenyl]-2-methylpropionic acid) The diastereomeric mixture Example 41 was separated applying chiral SFC: Column: Lux® Cellulose-2 10x250mm 5μm Solvent: scCO2 70%; MeOH + 20 mM NH3 30% Back pressure regulator 120bar Temperature 40℃ Flow rate 10ml / min Sample concentration: 10mg / ml Sample solvent: 100% MeOH Injection volume 200 μl

[0117] The optical purity was determined by analytical SFC, applying the same column material and solvents.

[0118] Example 51 (first eluting diastereomer; RT=3.83 min): [ka]

[0119] The absolute configuration of the methyl substituent at the alpha position of the carboxylic acid is arbitrarily assigned, therefore the pure diastereomeric Example 51 may have the structure shown or may have the structure shown in Example 52. MS (ESI pos.+neg. Loop-Inj.) m / z: 508 [M + H] +

[0120] Example 52 (Second-eluting diastereomer; RT=4.57 min): [ka]

[0121] The absolute configuration of the methyl substituent at the alpha position of the carboxylic acid is arbitrarily assigned, therefore the pure diastereomeric Example 52 may have the structure shown or may have the structure shown in Example 51. MS (ESI pos.+neg. Loop-Inj.) m / z: 508 [M + H] +

[0122] biological methods Assay A: Human KHK-C inhibition assay: Kinase activity of recombinant His-tagged KHK isoforms The enzymatic activity of recombinant human KHK-C was determined using Promega's ADP-GLO™ Kinase Assay Kit as described in the instructions. Briefly, 1.25 μg / ml His-tagged human KHK-A was incubated with 15 mM D-fructose at room temperature for 60 minutes. His-tagged human KHK-C (1 μg / ml) and His-tagged mouse KHK-C (0.625 μg / ml) were incubated with 400 μM D-fructose and 200 μM ATP at room temperature for 60 minutes. His-tagged rat KHK-C (0.5 μg / 1.5 ml) was incubated with 100 μM D-fructose and 200 μM ATP at room temperature for 60 minutes.

[0123] All incubations used the following assay buffer: 50 mM HEPES pH 7.4, 4 mM MgCl, 20 mM KCl, 0.01% Tween 20, 1 mM DTT. Enzyme reactions were stopped and developed using the ADP-GLO™ Kinase Kit according to the manufacturer's instructions, and results were analyzed using luminescence signals determined with an EnVision multiplate reader. Signals from enzyme and substrate-only samples were reported as 100%, and signals from enzyme-only samples were reported as 0%.

[0124] Assay B: Inhibition of fructose-1-phosphate production in HepG2 cells: Quantitative determination of fructose-1-phosphate in HepG2 cells. HepG2 cells (BB Knowles, Wistar Institute) were incubated with test compounds or vehicle (DMSO) in medium (EMEM, 10 mM NEAA, 8 mM glutamine, 10% FCS) at 37°C for 30 minutes under 5% CO2. D-fructose was added to a final concentration of 15 mM, and the cells were incubated for an additional 60 minutes under the same conditions. Cells were placed on ice, washed with phosphate-buffered saline, and lysed in 10 mM ammonium acetate. Cellular proteins were precipitated with acetonitrile, and an aliquot of the supernatant was analyzed for fructose-1-phosphate using RapidFire-MS / MS (RIAS) technology. Fructose-6-phosphate (0.1 μM) in the samples was used as an internal standard for quantification.

[0125] The following table shows the IC of the example compounds as determined using Assay A. 50 Indicates the value.

[0126] [Table 11] JPEG2025539698000065.jpg18779

[0127] The following table shows the IC of the example compounds as determined using Assay B. 50 Indicates the value.

[0128] [Table 12]

[0129] Comparison of the example compound with the prior art compound 6-[(3S,4S)-3,4-dihydroxy-1-pyrrolidinyl]-2-[(3R)-3-hydroxy-3-methyl-1-pyrrolidinyl]-4-(trifluoromethyl)-3-pyridinecarbonitrile (CAS registration number: 2711012-28-9).

[0130] [ka] This prior art compound (Huard et al., J. Med. Chem. 2017, 60, 7835-7849) was synthesized and then tested in Assay A under the same conditions as the example compounds of the present invention. The IC 50 The value was measured to be 187 nM.

Claims

1. A compound having the formula (I) or a salt thereof: 【Chemistry 1】 (In the formula, R1 is 【Chemistry 2】 and R2 is 【Transformation 3】 and where: 【Chemistry 4】 teeth, 【Transformation 5】 NH in etc. 2 ; 【Transformation 6】 NHMe in, etc.; and 【Transformation 7】 NMe in etc. 2 ; 【Transformation 8】 Hydroxymethyl in, etc.; 【Chemistry 9】 Hydroxy in and is substituted with one or two substituents selected from R3 is, 【Chemistry 10】 and Here, R5 is 【Chemistry 11】 Me or Et, optionally substituted with up to three F, such as R4 is, 【Chemistry 12】 It is.)

2. R1 is CHF 2 or CF 3 2. The compound of claim 1, wherein:

3. R2 is, 【Chemistry 13】 The compound of claim 1, which is (3-dimethylamino-pyrrolidin-1-yl).

4. R3 is, 【Chemistry 14】 or cyclopropyl.

5. R3 is SO 2 The compound or salt thereof according to claim 4, wherein Me is

6. R4 is, 【Chemistry 15】 2. The compound according to claim 1, or a salt thereof,

7. The compound according to claim 1, which is selected from the group consisting of compounds 1-52 of formula (I) or a salt thereof.

8. 10. A salt of a compound according to any one of claims 1 to 7 for use as a pharmaceutical.

9. A pharmaceutical preparation prepared using the compound or salt thereof according to any one of claims 1 to 7.

10. 10. A process for the preparation of a compound according to claim 1, comprising: General formula (II) 【Chemistry 16】 and a chloropyridine compound of the general formula (III) 【Chemistry 17】 under cross-coupling conditions applying a palladium catalyst such as XPhos G2 and a base such as cesium carbonate in a solvent such as dioxane. [Chemistry 18] The method comprising:

11. The compound of general formula (II) is a compound of general formula (IV) 【Chemistry 19】 by reaction with a suitable primary or secondary amine (V) in the presence of a base such as N,N-diisopropylethylamine in a solvent such as ethanol.

12. 10. A pharmaceutical composition comprising at least one compound according to claim 1 or a pharmaceutically acceptable salt thereof, together with one or more pharmaceutically acceptable carriers.

13. A pharmaceutical composition for use in the treatment of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH) and type 2 diabetes mellitus (T2DM).

14. 13. The pharmaceutical composition of claim 12, comprising a therapeutically effective amount of a compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 7 in the range of 0.1 to 90% by weight of the total composition, preferably in the range of 0.5 to 50% by weight of the total composition.

15. 13. A compound according to at least one of claims 1 to 7 or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 12, for use in the treatment or prevention of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH) and type 2 diabetes mellitus (T2DM).

16. 8. The compound or salt thereof according to any one of claims 1 to 7, for use in the treatment and / or prevention of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH) and type 2 diabetes mellitus (T2DM), wherein said compound is administered before, after or together with at least one other pharmaceutically active substance.

Images