Heterocyclic compounds and uses thereof

Abstract

The present invention relates to a novel hydrazinyl heterocyclic compound and, more specifically, to a novel hydrazinyl heterocyclic compound and uses thereof for treating diseases associated with various physiological functions through modulation of the histamine H4 receptor.

Description

Heterocyclic compounds and uses thereof

[0001] The present invention relates to a novel hydrazinyl heterocyclic compound, and more particularly, to a novel hydrazinyl heterocyclic compound and its use for treating diseases associated with various functions in a living body through its modulatory action on the histamine 4 receptor.

[0002] Histamine is a ubiquitous messenger molecule secreted by inflammatory and immune cells such as mast cells, T cells, enterochromosome-like cells, and neurons, exhibiting diverse physiological activities. Under normal or pathophysiological conditions, histamine, a bioactive small-molecular-weight amine, is secreted in response to external stimuli and binds to histamine receptors. Activated histamine receptors regulate diverse and important biological functions through intracellular mediation. In particular, histamine plays a central role in inflammatory and immune responses, as well as neurotransmitters and muscle contraction (W. Baumer et al., J. Dtsch. Dermatol. Ges. 2010, 8, 495-504).

[0003] Histamine receptors are reported to have four subtypes (histamine 1 receptor, histamine 2 receptor, histamine 3 receptor, and histamine 4 receptor) (De Backer et al., 1993. Biochem. Bioph. Res Commun. 197, 1601-1608; T. Oda et al., J. Biol. Chem. 2000, 275(47), 36781-36786; C. Liu et al., Mol. Pharmacol. 2001, 59(3), 420-426; T. Nguyen et al., Mol. Pharmacol. 2001, 59(3), 420-426; Y. Zhu et al., Mol. Pharmacol. 2001, 59(3), 434-441; KL Morse et al., J. Pharmaacol, Exp. Ther.2001, 296(3), 1058-1066; Igaz & Hegyesi.,Biology & Medical Aspects. 2004, 89~96 ME Parsons et al., Br. J. Pharmacol.2006,147, S127-S135).

[0004] The histamine H4 receptor (H4R) was discovered in 1994. This receptor protein consists of 390 amino acids (Fung-Leug, WP. et al., Curr. Opin. Invest. Drugs. 2004, 5(11), 1174-1183; IJP et al., Trends Pharmacol. Sci. 2005, 26(9), 462-469). It is a 7-transmembrane G-protein coupled receptor (Class A superfamily) that binds to Gi / o proteins in the inner part of the cell membrane. It has almost no amino acid homology with histamine 1 receptor and histamine 2 receptor, and shows ~35% homology with histamine 3 receptor (Lim. HD et al., Current Topics in Medi. Chem. 2006, 6, 1365-1373; RobertKiss, et al., Eur. J. Med. Chem., 2008, 43, 1059-1070).

[0005] Histamine 4 receptors are highly expressed mainly in immune tissues such as the thymus, bone marrow, and spleen, and are also observed in the brain (CNS), retina, lung, heart, and intestine. In terms of cells, high expression has been confirmed in immune cells such as eosinophils, basophils, T cells, mast cells, monocytes, and dendritic cells (RL Thurmond et al., Nat. Rev. Drug Discov. 2008, 7, 41-53; T. Nakamura et al., Biochem. Biophys. Res. Commun. 2000, 279, 615-620). The expression of histamine 4 receptors is significantly regulated in immune cells by various inflammatory stimuli, and plays a central role in inflammation and immune responses (Coge. F et al., Biochem. Biophys. Res Commun. 2002, 284(2), 301-309). The main mechanism is that histamine stimulation activates histamine 4 receptors in vivo, which increases the concentration of calcium, a second messenger, in cells or inhibits the secretion of cyclic adenosine monophosphate (cAMP) (M. Shahidet al., The Open Immunology Journal. 2009, 2, 9-41). Histamine 4 receptors play a central role in inflammation and immune responses, as well as influencing the activation, migration, infiltration, and production of cytokines and chemokines of various immune cells (R. Gutzmer et al., J. Immunol. 2005, 174, 5224-5232; CL Hofstra et al., J. Pharmacol. Exp. Ther. 2003, 305, 1212-1221; D. Dijkstra et al., J. Allergy Clin. Immunol. 2007, 120, 300-307; M. O'Reilly et al., J. Recept. Signal Transduct. Res.2002,22, 431-448).It exhibits physiological activity in various inflammatory responses such as neuropathic pain, inflammatory pain, arthritis, and nephritis by regulating the phosphorylation pathway of intracellular transcription factors and neurotransmitters (MD et al.,Pain. 2015, 156(12), 2492-2504).

[0006] As the effective roles and functions of histamine 4 receptor agonists in various disease areas including immunity and inflammation have been elucidated, various histamine 4 receptor ligands have been developed recently (WO 2013 / 048214 A2, WO 2021 / 066088 A1). Histamine 4 receptor antagonists are known to play an important role in inflammation, pain, and itch in various animal model experiments and clinical trials (SE et al., Eur J Pharmacol. 2011, 30, 667(1-3):383-8P; J. Dunford et al., J. Allergy Clin. Immunol. 2007, 119, 176-183; RL Thurmond et al., J. Pharmacol. Exp. Ther. 2004, 309, 404-413). In particular, histamine 4 receptor antagonists have been shown to effectively suppress itch induced by not only histamine but also other pruritic factors in animal experiments. This dual effect on inflammation and itch provides the basis for histamine 4 receptors as a good target for treating itch in a wide range of skin diseases accompanied by pruritus, such as atopic dermatitis, diabetic neuropathy associated pruritus, and chronic kidney disease associated pruritus (KK et al., J. Med. Chem. 2018, 61, 2949-2961; JM Cowden et al., J. Invest. Dermatol. 2010, 130, 1023-1033).

[0007] Recently, a study was published showing that histamine 4 receptors are specifically overexpressed in the eye, diabetic patients, and diabetic retina patients (Lee BJ et al., BMJ. Open. Diab. Res. Care. 2020, 8). Therefore, the development of treatments for diabetic neuropathy, allergic conjunctivitis, keratoconjunctivitis, ophthalmitis, dry eye, diabetic retinopathy, and age-related macular degeneration using these antagonists alone or in combination is highly anticipated. In addition, histamine 4 receptors are expressed in certain cancer cells (He, G.-H.et al.,(2018). J. Mol. Med. 96, 951-964), and research results related to the anticancer efficacy of histamine 4 receptor agonists alone or in combination have been published (WK Cai et al., Eur. J. Cancer. 2014, 50, 1195-1206; NA Massari et al., Oncotarget. 2017, 8, 26471-26491; AMB Abiuso et al., Eur. J. Cancer. 2018, 91, 125-135), so development as an anticancer agent is also expected.

[0008] An object of the present invention is to provide a heterocyclic compound that modulates histamine 4 receptors, or a pharmaceutically acceptable salt or isomer thereof.

[0009] Another object of the present invention is to provide a pharmaceutical composition for preventing or treating a disease associated with histamine 4 receptor, comprising a therapeutically effective amount of the heterocyclic compound, or a pharmaceutically acceptable salt or isomer thereof, as an active ingredient, together with a pharmaceutically acceptable carrier.

[0010] Another object of the present invention is to provide a use of the heterocyclic compound, or a pharmaceutically acceptable salt or isomer thereof, in the prevention or treatment of diseases associated with histamine 4 receptors.

[0011] Another object of the present invention is to provide a method for preventing or treating a disease associated with histamine 4 receptors by administering the heterocyclic compound, or a pharmaceutically acceptable salt or isomer thereof, to a subject in need thereof.

[0012] According to one aspect of the present invention, a compound of the following chemical formula 1, or a pharmaceutically acceptable salt or isomer thereof, is provided:

[0013] [Chemical Formula 1]

[0014]

[0015]

[0016] In the above chemical formula 1,

[0017] X is CH or N;

[0018] R1 and R4 are each independently -H, cycloalkyl-alkyl, alkylamino-alkyl, dialkylamino-alkyl, aryl-alkyl or saturated heterocycle-alkyl, provided that R1 and R4 are not both -H; or may be linked to each other together with the N atom to which they are bonded to form a saturated or unsaturated heterocycle; wherein said aryl and heterocycle may be optionally substituted with one or more substituents selected from the group consisting of halo, alkyl, alkoxy, haloalkyl, cycloalkyl, alkylamino, dialkylamino and (alkyl)(alkoxycarbonyl)amino;

[0019] R2 and R3 are each independently -H, alkyl, haloalkyl or aryl, provided that R2 and R3 are not both -H; wherein the aryl may be optionally substituted with one or more substituents selected from the group consisting of hydroxy, halo, amino, alkyl, alkylamino and dialkylamino;

[0020] R5 is -H, hydroxy, halo or amino;

[0021] The above heterocycle has one or more heteroatoms selected from N, O and S.

[0022]

[0023] Unless otherwise specified, the following terms used in the present invention have the meanings set forth below. Any undefined term has the meaning understood in the art.

[0024] The term “halo” or “halogen” as used herein, either alone or in combination with other additional terms (e.g., haloalkyl), means fluorine (F), chlorine (Cl), bromine (Br), or iodine (I).

[0025] The term “hydroxy” as used herein means -OH.

[0026] The term “carbonyl” as used herein means -C(=O)-.

[0027] The term “amino” group as used herein may mean a primary, secondary or tertiary amino group bonded through a nitrogen atom, either alone or in combination.

[0028] As used herein, the term “alkyl,” when used alone or in combination with other additional terms (e.g., haloalkyl), means a radical of a straight or branched saturated aliphatic hydrocarbon group having, for example, 1 to 7 or 1 to 5 carbon atoms. Typical examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, 1-ethylpropyl, hexyl, isohexyl, 1,1-dimethylbutyl, 2,2-dimethylbutyl, 3,3-dimethylbutyl, and 2-ethylbutyl.

[0029] The term “alkoxy” as used herein means an alkyloxy (-O-alkyl group), for example, an alkyloxy having 1 to 7 or 1 to 5 carbon atoms.

[0030] The term "cycloalkyl" as used herein refers to a saturated aliphatic hydrocarbon radical having, for example, 3 to 10 carbon atoms or 3 to 8 carbon atoms in a ring shape. Typical examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and the like.

[0031] The term “aryl” as used herein means an aromatic hydrocarbon having, for example, 6 to 10 carbon atoms, including but not limited to, phenyl and naphthyl.

[0032]

[0033] According to one specific example of the present invention, in the chemical formula 1

[0034] X is CH or N;

[0035] R1 and R4 are each independently -H, C3-C 10Cycloalkyl-C1-C7 alkyl, C1-C7 alkylamino-C1-C7 alkyl, di(C1-C7 alkyl)amino-C1-C7 alkyl, C6-C 10 Aryl-C1-C7 alkyl or 4 to 10 membered saturated heterocycle-C1-C7 alkyl, provided that R1 and R4 are not both -H; or may be linked to each other together with the N atom to which they are attached to form a 4 to 10 membered saturated or unsaturated heterocycle; wherein said aryl and heterocycle are halo, C1-C7 alkyl, C1-C7 alkoxy, halo-C1-C7 alkyl, C3-C 10 which may be optionally substituted with 1 to 3 substituents selected from the group consisting of cycloalkyl, C1-C7 alkylamino, di(C1-C7 alkyl)amino and (C1-C7 alkyl)(C1-C7 alkoxycarbonyl)amino;

[0036] R2 and R3 are each independently -H, C1-C7 alkyl, halo-C1-C7 alkyl or C6-C 10 Aryl, provided that R2 and R3 are not both -H; wherein the aryl may be optionally substituted with 1 to 3 substituents selected from the group consisting of hydroxy, halo, amino, C1-C7 alkyl, C1-C7 alkylamino, and di(C1-C7 alkyl)amino;

[0037] R5 is -H, hydroxy, halo or amino;

[0038] The above heterocycle may have 1 to 3 heteroatoms selected from N, O and S.

[0039] According to another specific example of the present invention, in the chemical formula 1, X may be N.

[0040] According to another specific example of the present invention, in the above formula 1, R1 and R4 are each independently -H, C3-C8 cycloalkyl-C1-C5 alkyl, C1-C5 alkylamino-C1-C5 alkyl, di(C1-C5 alkyl)amino-C1-C5 alkyl, C6-C 10Aryl-C1-C5 alkyl or 4 to 8 membered saturated heterocycle-C1-C5 alkyl, provided that R1 and R4 are not both -H; or may be linked to each other together with the N atom to which they are bonded to form a 4 to 8 membered saturated or unsaturated heterocycle; wherein said aryl and heterocycle may be optionally substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C5 alkyl, halo-C1-C5 alkyl, C3-C8 cycloalkyl, C1-C5 alkylamino, di(C1-C5 alkyl)amino and (C1-C5 alkyl)(C1-C5 alkoxycarbonyl)amino.

[0041] According to another specific example of the present invention, in the above formula 1, R2 and R3 are each independently -H, C1-C5 alkyl, halo-C1-C5 alkyl or C6-C 10 Aryl, provided that R2 and R3 are not both -H; wherein the aryl may be optionally substituted with 1 to 3 substituents selected from the group consisting of hydroxy, halo, amino, and C1-C7 alkyl.

[0042] According to another specific example of the present invention, R5 in the above chemical formula 1 may be amino.

[0043]

[0044] According to another specific embodiment of the present invention, representative examples of the compound of the above chemical formula 1 may include, but are not limited to, the following compounds:

[0045] 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylpiperazin-1-yl)-1,3,5-triazin-2-amine;

[0046] 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine;

[0047] 4-(2-(tert-butyl)hydrazinyl)-6-(4-isopropylpiperidin-1-yl)-1,3,5-triazin-2-amine;

[0048] 4-(2-(tert-butyl)hydrazinyl)-6-(4-isopropylpiperazin-1-yl)-1,3,5-triazin-2-amine;

[0049] 6-(2-(tert-butyl)hydrazinyl)-N 2 -(4-Fluorobenzyl-1-yl)-1,3,5-triazine-2,4-diamine;

[0050] tert-Butyl(1-(4-amino-6-(2-(3-chlorophenyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidine-3yl)(methyl)carbamate;

[0051] 4-(2-(3-chlorophenyl)hydrazinyl)-6-(3-methylamino)azetidin-1-yl)-1,3,5-triazin-2-amine;

[0052] 4-(2-(tert-butyl)hydrazinyl)-6-(4-cyclohexylpiperidin-1-yl)-1,3,5-triazin-2-amine;

[0053] 6-(2-(tert-butyl)hydrazinyl)-N 2 -(cyclopropylmethyl)-1,3,5-triazin-2-amine;

[0054] 6-(2-(tert-butyl)hydrazinyl)-N 2 -(2-dimethylamino)ethyl)-1,3,5-triazine-2,4-diamine;

[0055] 6-(2-(tert-butyl)hydrazinyl)-N 2 -(4-dimethylamino)benzyl)-1,3,5-triazine-2,4-diamine;

[0056] 4-(2-isopropylhydrazinyl)-6-(3-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine; and

[0057] 6-(2-(tert-butyl)hydrazinyl)-N2-(2-morpholineethyl)-1,3,5-triazine-2,4-diamine.

[0058]

[0059] The names of the above compounds were written according to the nomenclature provided by PerkinElmer's ChemDraw Professional (Version 15.0.0.106).

[0060]

[0061] The compound of chemical formula 1 according to the present invention may have an asymmetric carbon center and an asymmetric axis or asymmetric plane, and therefore may exist as stereoisomers such as E or Z isomers, R or S isomers, and racemates, and all of these isomers and mixtures are included in the scope of the present invention.

[0062] When the compound of chemical formula 1 according to the present invention is a racemate, the racemate can be separated into each isomer by a conventional separation method, for example, by mixing a corresponding developing solvent, preferably hexane, ethyl acetate, dichloromethane, methanol, etc. in the normal phase, or water and acetonitrile, etc. in the reverse phase, using chiral column chromatography in normal or reverse phase.

[0063]

[0064] The compound of formula 1 according to the present invention can also form pharmaceutically acceptable salts. Representative acids that can be used in the preparation of such pharmaceutically acceptable salts include, but are not limited to, the following. Hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrobromic acid, hydroiodic acid, formic acid, citric acid, acetic acid, trichloroacetic acid or trifluoroacetic acid, benzoic acid, fumaric acid, maleic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, 2,2-dichloroacetic acid, acylated amino acids, adipic acid, alginic acid, ascorbic acid, L-aspartic acid, 4-acetamidobenzoic acid, (+)-camphoric acid, camphorsulfonic acid, (+)-(1S)-camphorsulfonic acid, capric acid, caproic acid, caprylic acid, cinnamic acid, cyclamic acid, dodecylsulfuric acid, ethane-1,2-disulfonic acid, ethanesulfonic acid, 2-hydroxy-ethanesulfonic acid, galactaric acid, gentisic acid, Acid addition salts formed by glucoheptanoic acid, D-gluconic acid, D-glucuronic acid, L-glutamic acid, α-oxo-glutaric acid, glycolic acid, hippuric acid, (+)-L-lactic acid, (±)-DL-lactic acid, lactobionic acid, (-)-L-malic acid, malonic acid, (±)-DL-mandelic acid, methanesulfonic acid, naphthalene-2-sulfonic acid, naphthalene-1,5-disulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinic acid, oleic acid, orthotonic acid, oxalic acid, palmitic acid, pamoic acid, L-pyroglutamic acid, salicylic acid, 4-amino-salicylic acid, sebacic acid, stearic acid, succinic acid, tannic acid, (+)-L-tartaric acid, thiocyanic acid, and undecylenic acid are included. In addition, amine derivatives may include other acid salts known and used in the art. These are prepared by commonly known processes.

[0065]

[0066] The compound of the above chemical formula 1 can be prepared according to the method described in the examples below, but is not limited thereto.

[0067]

[0068] The compound of formula 1 according to the present invention has excellent activity capable of modulating the human histamine 4 receptor (hH4R). Therefore, according to another aspect of the present invention, a pharmaceutical composition is provided comprising the compound of formula 1, or a pharmaceutically acceptable salt or isomer thereof, as an active ingredient, together with a pharmaceutically acceptable carrier.

[0069] According to another embodiment of the present invention, the pharmaceutical composition can be used to prevent or treat diseases associated with human histamine 4 receptor (hH4R).

[0070] According to another embodiment of the present invention, the disease associated with the human histamine 4 receptor (hH4R) may be selected from the group consisting of, but is not limited to, nasal polyps, allergic rhinitis, non-allergic rhinitis, viral rhinitis, nasal itching, sinusitis, nasal congestion (stuffy nose), asthma, chronic obstructive pulmonary disease (COPD), rheumatoid arthritis, atopic dermatitis, psoriasis, eczema, pruritus, skin itching, urticaria, idiopathic chronic urticaria, scleroderma, allergic conjunctivitis, keratoconjunctivitis, ophthalmitis, dry eye, diabetic retinopathy, heart failure, arrhythmia, atherosclerosis, multiple sclerosis, inflammatory bowel disease, inflammatory pain, neuropathic pain, osteoarthritis pain, thyroid autoimmune disease, immune-mediated diabetes, lupus, post-surgical adhesion formation, cancer, and vestibular disorder.

[0071] According to another embodiment of the present invention, the inflammatory bowel disease may be, but is not limited to, colitis, Crohn's disease or ulcerative colitis.

[0072]

[0073] The pharmaceutical composition according to the present invention can be prepared by mixing a therapeutically effective amount of a compound of formula 1, a pharmaceutically acceptable salt or isomer thereof, as an active ingredient, with a pharmaceutically acceptable carrier, receptor, binder, stabilizer, and / or diluent. In addition, when the pharmaceutical composition according to the present invention is prepared in the form of an injection solution, a pharmaceutically acceptable buffer, solubilizer, and / or isotonic agent can be mixed with the compound of formula 1 of the present invention, a pharmaceutically acceptable salt or isomer thereof.

[0074] The pharmaceutical composition according to the present invention can be prepared in a delivery form comprising one or more dosage units of a pharmaceutical agent using any preparation technique available or known to those skilled in the art and suitable pharmaceutical excipients. In the method of the present invention, the composition can be administered by any suitable delivery route, for example, oral or parenteral administration, parenteral, rectal, topical, or ophthalmic routes, or by inhalation. The pharmaceutical preparation can be in the form of a tablet, capsule, sachet, dragee, powder, granule, lozenge, powder for reconstitution, liquid preparation, or suppository. For example, the composition is formulated for intravenous infusion, spray, topical administration, or oral administration.

[0075] When preparing oral preparations, conventional pharmaceutical carriers can be used. For example, for oral liquid preparations such as suspensions, syrups, elixirs, and solutions, carriers such as water, glycols, oils, and alcohols can be used; for solid preparations such as powders, pills, capsules, and tablets, starch, sugar, kaolin, lubricants, binders, and disintegrating agents can be used. Tablets and capsules are the most convenient dosage forms due to their ease of administration, and tablets and pills are preferably formulated as enteric coatings.

[0076] For parenteral preparations, sterile water is typically used as the carrier, and other ingredients such as solubilizers may also be included. Injectable preparations, such as sterile injectable aqueous or oily suspensions, can be prepared using suitable dispersing, wetting, or suspending agents according to known techniques. Suitable solvents include water, Ringer's solution, and isotonic NaCl solution. Sterile fixed oils are also commonly used as solvents or suspending media. Any non-irritating fixed oil, including mono- and di-glycerides, can be used for this purpose, and fatty acids such as oleic acid can also be used in injectable preparations.

[0077] For transdermal formulations, penetration enhancers and / or suitable wetting agents may be used as carriers, optionally along with suitable non-irritating additives. The additives are selected to facilitate transdermal administration and / or aid in the preparation of the desired composition. Transdermal formulations are administered in various ways, such as through transdermal patches, drops, or ointments.

[0078] The dosage and administration time of the pharmaceutical composition according to the present invention may vary depending on the patient's disease, condition, age, weight, and dosage form, and the composition may be administered to adults at a dose of 0.1 to 2,000 mg, preferably 1 to 200 mg, once or in several divided doses, but is not limited thereto.

[0079] The compound of chemical formula 1 according to the present invention, or a pharmaceutically acceptable salt or isomer thereof, has an excellent effect of activating or inhibiting histamine 4 receptors, and therefore, a pharmaceutical composition containing the compound is useful for preventing or treating related diseases that require modulation of histamine 4 receptors.

[0080] Hereinafter, the present invention will be described in more detail with reference to the following examples and experimental examples. However, these examples and experimental examples are merely illustrative and do not limit the scope of the present invention in any way.

[0081]

[0082] The definitions of abbreviations used in the examples below are as follows.

[0083]

[0084]

[0085] Example 1: Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylpiperazin-1-yl)-1,3,5-triazin-2-amine

[0086]

[0087]

[0088] (a) Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine

[0089] 2-Amino-4,6-dichlorotriazine (500 mg, 3.03 mmol) and tert-butylhydrazine HCl (453 mg, 3.64 mmol) were dissolved in EtOH (12 mL), and DIPEA (1.58 mL, 9.09 mmol) was slowly added at room temperature. The reaction solution was stirred at 90°C for 1.5 h, then H2O (50 mL) was added and extracted with EtOAc (100 mL). The organic layer was washed with brine, dried over anhydrous MgSO4, filtered through Celite, and then distilled under reduced pressure. The residue was purified by column chromatography on amine silica (DCM: MeOH = 20:1), and the fractions containing the product were combined and evaporated to obtain a white solid compound 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (420 mg, 63%).

[0090] LC / MS ESI (+): 218 (M+1)

[0091]

[0092] (b) Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylpiperazin-1-yl)-1,3,5-triazin-2-amine

[0093] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and 1-methylpiperazine (88 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (120 uL, 0.69 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. H2O (10 mL) was poured in, and the mixture was extracted with EtOAc (20 mL). The organic layer was washed with brine, dried over anhydrous MgSO4, filtered through Celite, and then distilled under reduced pressure. After dissolving the concentrate in methanol (1 mL), hexane was added, trituration was performed, and the solid was filtered and evaporated under reduced pressure to obtain a light beige solid compound 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylpiperazin-1-yl)-1,3,5-triazin-2-amine (55 mg, 85%).

[0094] LC / MS ESI (+): 281 (M+1)

[0095] 1 H NMR (400 MHz, DMSO-d6) δ= 7.96 (br s, 1H), 6.20 (br s, 2H), 3.61 (m, 4H), 3.03 (s, 1H), 2.32 (m, 4H), 2.22 (s, 3H), 1.00 (s, 9H)

[0096]

[0097] Example 2: Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine

[0098]

[0099]

[0100] (a) Synthesis of tert-butyl(1-(4-amino-6-chloro-1,3,5-2-yl)azetidin-3-yl)(methyl)carbamate

[0101] 2-Amino-4,6-dichlorotriazine (500 mg, 3.03 mmol) and tert-butyl azetidin-3-yl(methyl)carbamate (677 mg, 3.64 mmol) were dissolved in EtOH (12 mL), and DIPEA (1.06 mL, 6.06 mmol) was slowly added at room temperature. The reaction solution was stirred at 90°C for 1.5 h, then H2O (20 mL) was poured into it, and extracted with EtOAc (50 mL). The organic layer was washed with brine, dried over anhydrous MgSO4, filtered through Celite, and then distilled under reduced pressure. The residue was purified by column chromatography on amine silica (n-Hex: EtOAc = 2:1), and the fractions containing the product were combined and evaporated to obtain a white solid compound tert-butyl (1-(4-amino-6-chloro-1,3,5-2-yl)azetidin-3-yl)(methyl)carbamate (610 mg, 63%).

[0102] LC / MS ESI (+): 316 (M+1)

[0103]

[0104] (b) Synthesis of tert-butyl(1-(4-amino-6-(2-(tert-butyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate

[0105] tert-Butyl(1-(4-amino-6-chloro-1,3,5-2-yl)azetidin-3-yl)(methyl)carbamate (50 mg, 0.16 mmol) and tert-butylhydrazine HCl (59 mg, 0.48 mmol) were dissolved in 1,4-dioxane (1 mL) in a sealed tube, and then DIPEA (0.831 mL, 0.48 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 72 h, followed by the addition of H2O (10 mL) and extraction with EtOAc (20 mL). The organic layer was washed with brine, dried over anhydrous MgSO4, filtered through Celite, and then distilled under reduced pressure. After dissolving the concentrate in ethanol (1 mL), hexane was added, trituration was performed, and the solid was filtered and evaporated under reduced pressure to obtain a bright white solid compound tert-butyl (1-(4-amino-6-(2-(tert-butyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate (45 mg, 77%).

[0106] LC / MS ESI (+): 367 (M+1)

[0107]

[0108] (c) Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine

[0109] Tert-Butyl(1-(4-amino-6-(2-(tert-butyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate (40 mg, 0.11 mmol) was dissolved in DCM (1 mL), and TFA (0.5 mL) was slowly added at 0°C. The reaction mixture was stirred at room temperature for 1 h and then distilled under reduced pressure. The residue was purified by column chromatography on amine silica (DCM:MeOH = 40:1), and the fractions containing the product were combined and lyophilized to give 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine (23 mg, 79%) as a white solid.

[0110] LC / MS ESI (+): 267 (M+1)

[0111] 1 H NMR (400 MHz, DMSO-d6) δ= 7.96 (br s, 1H), 6.29 (br s, 2H), 4.54 (br s, 1H), 4.00 (br t,J=7.8Hz, 2H), 3.59(dd,J=5.1, 8.7Hz, 2H), 3.51 - 3.38 (m, 1H), 2.20 (s, 3H), 0.98 (s, 9H)

[0112]

[0113] Example 3: Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-(4-isopropylpiperidin-1-yl)-1,3,5-triazin-2-amine

[0114]

[0115]

[0116] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and 4-isopropylpiperidine (67 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (120 μL, 0.69 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. Hexane was added to the reaction solution, triturated, and the solid was filtered and evaporated under reduced pressure to obtain a brown solid compound 4-(2-(tert-butyl)hydrazinyl)-6-(4-4-isopropylpiperidin-1-yl)-1,3,5-triazin-2-amine (68 mg, 96%).

[0117] LC / MS ESI (+): 308 (M+1)

[0118] 1 H NMR (400 MHz, DMSO-d6) δ= 8.70 (br s, 1H), 4.67 (br d, 1H), 3.25 (d, 2H), 2.76(t, 2H), 1.74 (br, d, 2H), 1.62 (br, d, 1H), 1.34 -1.31 (m, 3H), 1.01 (s, 6H), 0.86 (s, 9H)

[0119]

[0120] Example 4: Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-(4-isopropylpiperazin-1-yl)-1,3,5-triazin-2-amine

[0121]

[0122]

[0123] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and 4-isopropylpiperazine (88 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and DIPEA (120 μL, 0.69 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 h and then cooled to room temperature. Hexane was added to the reaction solution, triturated, and the solid was filtered and evaporated under reduced pressure. The residue was purified by column chromatography on amine silica (DCM: MeOH = 50:1), and the fractions containing the product were combined and lyophilized to obtain 4-(2-(tert-butyl)hydrazinyl)-6-(4-isopropylpiperazin-1-yl)-1,3,5-triazin-2-amine (63 mg, 89%) as a light beige solid.

[0124] LC / MS ESI (+): 309 (M+1)

[0125] 1 H NMR (400 MHz, DMSO-d6) δ = 10.4 (br s, 1H), 7.30 (br d, 1H), 4.72 (br s, 2H), 3.44 (br, 4H), 3.37 (br, 4H), 1.38 (s, 9H), 1.02 (s, 6H)

[0126]

[0127] Example 5: Synthesis of 6-(2-(tert-butyl)hydrazinyl)-N2-(4-fluorobenzyl-1-yl)-1,3,5-triazine-2,4-diamine

[0128]

[0129]

[0130] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and 4-fluorobenzylamine (87 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (120 uL, 0.69 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. Normal hexane was added to the reaction solution, triturated, and the solid was filtered and evaporated under reduced pressure. The residue was purified by column chromatography on amine silica (DCM: MeOH = 50:1), and the fractions containing the product were combined and lyophilized to give a beige solid compound 6-(2-(tert-butyl)hydrazinyl)-N 2 -(4-Fluorobenzyl-1-yl)-1,3,5-triazine-2,4-diamine (53 mg, 75%) was obtained.

[0131] LC / MS ESI (+): 306 (M+1)

[0132] 1 H NMR (400 MHz, MeOD-d4) δ= 7.34 (d, 2H), 7.04 (d, 2H), 4.52 (s, 2H), 1.11(s, 9H)

[0133]

[0134] Example 6: Synthesis of tert-butyl(1-(4-amino-6-(2-(3-chlorophenyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate

[0135]

[0136]

[0137] tert-Butyl(1-(4-amino-6-chloro-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate (50 mg, 0.16 mmol) and 3-chlorophenylhydrazine HCl (84.5 mg, 0.48 mmol) were dissolved in 1,4-dioxane (1 mL) in a sealed tube, and then DIPEA (125.4 μL, 0.72 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. The reaction solution was filtered and concentrated on amine silica gel (NH SiO2), and then solid crystals were obtained at low temperature under DCM-Hex, which were filtered and evaporated under reduced pressure to obtain a beige solid compound tert-butyl(1-(4-amino-6-(2-(3-chlorophenyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate (55 mg, 81%).

[0138] LC / MS ESI (+): 422 (M+1)

[0139]

[0140] Example 7: Synthesis of 4-(2-(3-chlorophenyl)hydrazinyl)-6-(3-methylamino)azetidin-1-yl)-1,3,5-triazin-2-amine

[0141]

[0142]

[0143] Tert-Butyl(1-(4-amino-6-(2-(3-chlorophenyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidin-3yl)(methyl)carbamate (40 mg, 0.095 mmol) was dissolved in DCM (1 mL), and TFA (0.5 mL) was slowly added at 0°C. The reaction mixture was stirred at room temperature for 1 hour and then distilled under reduced pressure. The residue was purified by column chromatography on reversed-phase silica (H2O:CH3CN = 70:30 containing 0.1% TFA), and the fractions containing the product were combined, concentrated by evaporation, and lyophilized to obtain a pale purple solid compound 4-(2-3-chlorophenyl)hydrazinyl)6-(3-methylamino)azetidin-1-yl)-1,3,5-triazin-2-amine (23 mg, 75%).

[0144] LC / MS ESI (+): 321 (M+1)

[0145] 1 H NMR (400 MHz, DMSO-d6) δ= 8.54 (br s, 1H), 8.22 (br s, 1H), 7.89 (br s, 1H), 7.10 (t, 1H), 6.62 (m, 2H), 6.43 (br s, 2H), 4.00 (m, 2H), 3.62 (m, 2H), 3.38 (m, 1H), 2.21 (s, 3H)

[0146]

[0147] Example 8: Synthesis of 4-(2-(tert-butyl)hydrazinyl)-6-(4-cyclohexylpiperidin-1-yl)-1,3,5-triazin-2-amine

[0148]

[0149]

[0150] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (35 mg, 0.16 mmol) and 4-cyclohexylpiperidine HCl (50 mg, 0.25 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (84 μL, 0.48 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. n-Hexane was added to the reaction solution, triturated, and the solid was filtered and evaporated under reduced pressure to obtain a beige solid compound 4-(2-(tert-butyl)hydrazinyl)-6-(4-cyclohexylpiperidin-1-yl)-1,3,5-triazin-2-amine (31 mg, 56%).

[0151] LC / MS ESI (+): 349 (M+1)

[0152] 1 H NMR (400 MHz, DMSO-d6) δ= 8.87 (br s, 1H), 7.02 (br d, 1H), 4.69 (br, 2H), 3.61 (m, 2H), 3.15(m, 2H), 1.68 (br, 4H), 1.30 - 1.23(m, 12H), 1.06(s, 9H)

[0153]

[0154] Example 9: Synthesis of 6-(2-(tert-butyl)hydrazinyl)-N2-(cyclopropylmethylamino)-1,3,5-triazin-2-amine

[0155]

[0156]

[0157] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and cyclopropylmethylamine (49 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (120 μL, 0.69 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 h and then cooled to room temperature. The residue was purified by column chromatography on amine silica (DCM: MeOH = 50:1), and the fractions containing the product were combined and lyophilized to give a beige solid compound 6-(2-(tert-butyl)hydrazinyl)-N 2 -(Cyclopropylmethyl)-1,3,5-triazin-2-amine (42 mg, 72%) was obtained.

[0158] LC / MS ESI (+): 252 (M+1)

[0159] 1 H NMR (400 MHz, MeOD-d4) δ= 3.25 (t, 2H), 1.16 (s, 9H), 1.08 (br, 1H), 0.54 (m, 2H), 0.28 (m, 2H)

[0160]

[0161] Example 10: Synthesis of 6-(2-(tert-butyl)hydrazinyl)-N2-(2-dimethylamino)ethyl)-1,3,5-triazine-2,4-diamine

[0162]

[0163]

[0164] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and 4-dimethylethylenediamine (61 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (120 μL, 0.69 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. The residue was purified by column chromatography on reversed-phase silica (H2O:CH3CN = 70:30 containing 0.1% TFA). The fractions containing the product were combined, concentrated by evaporation, and lyophilized to obtain a white solid compound, 6-(2-(tert-butyl)hydrazinyl)-N 2 -(2-Dimethylamino)ethyl)-1,3,5-triazine-2,4-diamine (37 mg, 60%) was obtained

[0165] LC / MS ESI (+): 269 (M+1)

[0166] 1 H NMR (400 MHz, MeOD-d4) δ 3.80 (t, 2H), 3.40 - 3.35 (t, 2), 2.99 (s, 6H), 1.17 (s, 9H)

[0167]

[0168] Example 11: Synthesis of 6-(2-(tert-butyl)hydrazinyl)-N2-(4-dimethylamino)benzyl)-1,3,5-triazine-2,4-diamine

[0169]

[0170]

[0171] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and 4-dimethylaminobenzylamine HCl (129 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (120 μL, 0.69 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 h and then cooled to room temperature. Hexane was added to the reaction solution, triturated, and the solid was filtered and evaporated under reduced pressure to obtain a light beige solid compound, 6-(2-(tert-butyl)hydrazinyl)-N 2 -(4-dimethylamino)benzyl)-1,3,5-triazine-2,4-diamine (49 mg, 65%) was obtained

[0172] LC / MS ESI (+): 331 (M+1)

[0173] 1 H NMR (400 MHz, MeOD-d4) δ= 7.34 (d, 2H), 6.97 (d, 2H), 4.42 (s, 2H), 3.31 (s, 6H), 1.02(s, 9H)

[0174]

[0175] Example 12: Synthesis of 4-(2-isopropylhydrazinyl)-6-(3-(methylamino)azetidin-1-yl-1,3,5-triazin-2-amine

[0176]

[0177]

[0178] (a) Synthesis of tert-butyl(1-(4-amino-6-(2-(isopropylhydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate

[0179] tert-Butyl(1-(4-amino-6-chloro-1,3,5-2-yl)azetidin-3-yl)(methyl)carbamate (50 mg, 0.16 mmol) and isopropylhydrazine HCl (52.4 mg, 0.48 mmol) were dissolved in 1,4-dioxane (1 mL) in a sealed tube, and then DIPEA (125.4 μL, 0.72 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. The reaction solution was filtered and concentrated on amine silica gel (NH SiO2), and then solid crystals were obtained at low temperature under DCM-Hx, which were filtered and evaporated under reduced pressure to obtain a beige solid compound tert-butyl(1-(4-amino-6-(2-(isopropylhydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate (36 mg, 64%).

[0180] LC / MS ESI (+): 353 (M+1)

[0181]

[0182] (b) Synthesis of 4-(2-isopropylhydrazinyl)-6-(3-methylamino)azetidin-1-yl)-1,3,5-triazin-2-amine

[0183] Tert-Butyl(1-(4-amino-6-(2-(isopropylhydrazinyl)-1,3,5-triazin-2-yl)azetidin-3-yl)(methyl)carbamate (30 mg, 0.09 mmol) was dissolved in DCM (1.0 mL), and TFA (0.5 mL) was slowly added at 0°C. The reaction solution was stirred at room temperature for 1 h. After stirring at room temperature for 1 h, the reaction solution was distilled under reduced pressure. The residue was purified by column chromatography on reversed-phase silica (H2O:CH3CN = 70:30 containing 0.1% TFA), and the fractions containing the product were combined and lyophilized to give 4-(2-isopropylhydrazinyl)-6-(3-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine (16 mg, 71%) as a white solid.

[0184] LC / MS ESI (+): 253 (M+1)

[0185] 1 H NMR (400 MHz, DMSO-d6) δ= 9.32 (br, 1H), 8.32 (br, 1H), 4.60 - 4.11 (br, 1H), 3.63 (br t,J=7.8Hz, 2H), 3.16(dd,J=5.1, 8.7Hz, 2H), 3.14 (m, 1H), 2.50 (s, 3H), 1.15 (s, 9H)

[0186]

[0187] Example 13: Synthesis of 6-(2-(tert-butyl)hydrazinyl)-N2-(2-morpholineethyl)-1,3,5-triazine-2,4-diamine

[0188]

[0189]

[0190] 4-(2-(tert-butyl)hydrazinyl)-6-chloro-1,3,5-triazin-2-amine (50 mg, 0.23 mmol) and 2-morpholinoethane-1-amine (50 mg, 0.69 mmol) were dissolved in 1,4-dioxane (1 mL) in a seal tube, and then DIPEA (40 μL, 0.23 mmol) was slowly added at room temperature. The reaction solution was stirred at 100°C for 48 hours and then cooled to room temperature. The residue was purified by column chromatography on reversed-phase silica (H2O:CH3CN = 70:30 containing 0.1% TFA). The fractions containing the product were combined, concentrated by evaporation, and lyophilized to give the colorless amorphous compound 6-(2-(tert-butyl)hydrazinyl)-N 2 -(2-Morpholineethyl)-1,3,5-triazine-2,4-diamine (17 mg, 24%) was obtained.

[0191] LC / MS ESI (+): 311 (M+1)

[0192] 1H NMR (400 MHz, DMSO-d6) δ= 9.82 (br, 1H), 8.26 (br, 2H), 7.71 (br, 1H), 3.97(br, d, 2H), 3.67 (m, 6H), 3.29 (m, 2H), 3.11 (m, 2H), 1.12 (s, 9H)

[0193]

[0194] Experimental Example 1: Human Histamine 4 Receptor (hH4R) Binding Analysis

[0195] The compound prepared in the example was diluted 1,000-fold (v / w) in DMSO solution, and 1 μL of the diluted compound solution was mixed with 99 μL of assay buffer (50 mM Tris-HCl pH 7.4, 5.0 mM EDTA) to prepare concentrations of 0.01, 0.03, 0.1, 0.3, and 1 μM. 20 μL of the prepared compound solution was dispensed into each well of a 96-well plate, and 20 μL of 100 μM histamine diluted in assay buffer and 1% DMSO solution were also dispensed into each well to calculate nonspecific binding and total binding. 15 μg of cell membranes (Multispan) overexpressing human histamine 4 receptors were diluted in 160 μL of assay buffer and dispensed into each well. [ 3 H] Labeled histamine (PerkinElmer) was diluted to a concentration of 10 nM, dispensed 20 μL into each well, and left in a 27°C water bath for 30 minutes. After the reaction was completed, 100 μL of the mixture was dispensed onto a glass fiber plate pre-absorbed with 0.5% polyethyleneamine, and unbound [ 3 H] labeled histamine was removed by vacuum. After washing six times with 200 μL of washing buffer (50 mM Tris-HCl, pH 7.4), the plate was dried in an oven at 37°C for 18 hours. 100 μL of betaScint cocktail solution was dispensed into each well, and after 10 minutes, [ bound to human histamine 4 receptor3 H] The CPM (count per minute) value of labeled histamine was expressed as Micro beta2 TM ) was measured. The binding affinity (IC) of the compound of the example to the human histamine 4 receptor 50 ) was analyzed using the Excel program, and the results are shown in Table 1.

[0196] [Table 1]

[0197]

[0198]

[0199] Experimental Example 2: Solubility Test in Artificial Gastrointestinal Solution

[0200] [First amount]

[0201] 2.0 g of sodium chloride was diluted to 1 L with 7.0 mL of hydrochloric acid and water. The pH at this time was approximately 1.2. A small amount (20 mL) of this solution was taken, and 25 μL of Triton X-100 was added to make the total volume 25 mL.

[0202] [Second Amount]

[0203] Add 0.348 g of sodium hydroxide, 3.438 g of NaH2PO4, 6.186 g of sodium chloride, and water to make 1 L. The pH at this time was 6.5. Take a small amount (20 mL) of this solution, then add 41.25 mg of sodium taurocholate and 25 μL of 750 mM lecithin dissolved in ethanol, and make the total volume 25 mL.

[0204] [Experimental Method]

[0205] For the compounds manufactured in the examples, the first and second solutions were each prepared to have 2 mg / mL, and then shaken for 1 minute and sonicated for 1 minute, and then heated at 37°C for 1 hour for the first solution and for 2 hours for the second solution. After heating, the test solutions were filtered to remove any undissolved compounds in the test solutions. Finally, 100 μL of the fraction in which the compound was completely dissolved was added to prepare an analysis solution. The solubility of the analysis solutions for each test solution was measured using liquid chromatography. The results of the solubility evaluation in the first and second solutions are shown in Table 2, and high solubility was confirmed.

[0206] [Table 2]

[0207]

[0208]

[0209] Experimental Example 3: Metabolic Stability Experiment

[0210] [Preparation of microsome solution]

[0211] Microsomes (human, dog, rat, and mouse) with a protein concentration of 20 mg / mL were diluted with 0.1 M phosphate buffer (pH: 7.4) to prepare a concentration of 2 mg / mL. The compound initially prepared at 50 mM in DMSO solution was diluted to 1 mM using DMSO, and then diluted with distilled water to prepare a concentration of 4 μM. The microsome dilution prepared above was left in a 37°C water bath for 5 minutes, and then mixed 1:1 with the 4 μM prepared compound to prepare a microsome solution with a protein concentration of 1 mg / mL and a compound concentration of 2 μM.

[0212] [NADPH solution preparation]

[0213] A 2 mM NADPH solution was prepared by dissolving it in 0.1 M phosphate buffer.

[0214] [Experimental Method]

[0215] The reaction was initiated by mixing the microsome solution prepared above with 2 mM NADPH in a 1:1 ratio to obtain a final microsome protein concentration of 0.5 mg / mL, compound 1 μM, and NADPH 1 mM. Samples were taken after 0, 10, 30, and 60 minutes, mixed with acetonitrile in a 1:1 ratio to terminate the reaction, and centrifuged. The supernatant was analyzed using liquid chromatography (or, proteins, etc. are filtered through a centrifugal filter using a 96-well filter plate to separate and analyze the sample). The half-life was obtained using the peak area of ​​the compound remaining after 0, 10, 30, and 60 minutes, and CL, a parameter indicating metabolic stability, was calculated. int(liver) (hepatic intrinsic clearance) was calculated and shown in Table 3. It was confirmed that metabolic stability was very stable regardless of species.

[0216] [Table 3]

[0217]

[0218]

[0219] Experimental Example 4: Plasma Stability Experiment

[0220] After adding the drug to the plasma and reacting it at 37°C for 2 hours, the plasma was collected and the % remaining compared to the amount of drug before the reaction was measured using LC-MS / MS.

[0221] (1) Before the experiment, frozen plasma was thawed in a 37℃ water bath.

[0222] (2) Plasma was centrifuged at 13,000 rpm for 5 minutes to remove clots.

[0223] (3) 120 μl of acetonitrile containing 120 μl of internal standard was dispensed into a 96-well plate (on the ice).

[0224] (4) 693 μl of plasma was added to another 96-well plate.

[0225] (5) 7 μl of the compound prepared in the 0.1 mM example was added and mixed well.

[0226] (6) 60 μl was quickly dispensed into a new 96-well plate.

[0227] (7) Among the busy samples, 50 μl of the 0-minute sample was added to 120 μl of acetonitrile (including internal standard).

[0228] (8) 50 μl was taken from each sample at 10, 30, 60, and 120 minutes, placed in 120 μl of acetonitrile, and re-bleached.

[0229] (9) After 120 minutes, all samples were transferred to a 96-well filter plate and centrifuged.

[0230] (10) 5 μl of the sample was injected into LC / MSMS and analyzed.

[0231]

[0232] In this experiment, enalapril, bisacodyl, and procaine, which have been reported to be stable in rat, mouse, and human plasma, were used as positive controls (enalapril, <1% for rats; procaine, <1% for mice and humans; bisacodyl, <1% for dogs). The experimental results are shown in Table 4 below.

[0233] [Table 4]

[0234]

Claims

1. A compound of the following chemical formula 1, or a pharmaceutically acceptable salt or isomer thereof: [Chemical Formula 1] In the above chemical formula 1, X is CH or N; R1 and R4 are each independently -H, cycloalkyl-alkyl, alkylamino-alkyl, dialkylamino-alkyl, aryl-alkyl or saturated heterocycle-alkyl, provided that R1 and R4 are not both -H; or may be linked to each other together with the N atom to which they are bonded to form a saturated or unsaturated heterocycle; wherein said aryl and heterocycle may be optionally substituted with one or more substituents selected from the group consisting of halo, alkyl, alkoxy, haloalkyl, cycloalkyl, alkylamino, dialkylamino and (alkyl)(alkoxycarbonyl)amino; R2 and R3 are each independently -H, alkyl, haloalkyl or aryl, provided that R2 and R3 are not both -H; wherein the aryl may be optionally substituted with one or more substituents selected from the group consisting of hydroxy, halo, amino, alkyl, alkylamino and dialkylamino; R5 is -H, hydroxy, halo or amino; The above heterocycle has one or more heteroatoms selected from N, O and S.

2. In paragraph 1, X is CH or N; R1 and R4 are each independently -H, C3-C 10 Cycloalkyl-C1-C7 alkyl, C1-C7 alkylamino-C1-C7 alkyl, di(C1-C7 alkyl)amino-C1-C7 alkyl, C6-C 10 Aryl-C1-C7 alkyl or 4 to 10 membered saturated heterocycle-C1-C7 alkyl, provided that R1 and R4 are not both -H; or may be linked to each other together with the N atom to which they are attached to form a 4 to 10 membered saturated or unsaturated heterocycle; wherein said aryl and heterocycle are halo, C1-C7 alkyl, C1-C7 alkoxy, halo-C1-C7 alkyl, C3-C 10 which may be optionally substituted with 1 to 3 substituents selected from the group consisting of cycloalkyl, C1-C7 alkylamino, di(C1-C7 alkyl)amino and (C1-C7 alkyl)(C1-C7 alkoxycarbonyl)amino; R2 and R3 are each independently -H, C1-C7 alkyl, halo-C1-C7 alkyl or C6-C 10 Aryl, provided that R2 and R3 are not both -H; wherein the aryl may be optionally substituted with 1 to 3 substituents selected from the group consisting of hydroxy, halo, amino, C1-C7 alkyl, C1-C7 alkylamino, and di(C1-C7 alkyl)amino; R5 is -H, hydroxy, halo or amino; A compound or a pharmaceutically acceptable salt or isomer thereof, wherein the heterocycle has 1 to 3 heteroatoms selected from N, O and S.

3. A compound or a pharmaceutically acceptable salt or isomer thereof, characterized in that X is N in the first paragraph.

4. In paragraph 1, R1 and R4 are each independently -H, C3-C8 cycloalkyl-C1-C5 alkyl, C1-C5 alkylamino-C1-C5 alkyl, di(C1-C5 alkyl)amino-C1-C5 alkyl, C6-C 10 A compound or a pharmaceutically acceptable salt or isomer thereof, characterized in that the compound is aryl-C1-C5 alkyl or 4 to 8 membered saturated heterocycle-C1-C5 alkyl, provided that R1 and R4 are not both -H; or may be linked to each other together with the N atom to which they are bonded to form a 4 to 8 membered saturated or unsaturated heterocycle; wherein the aryl and heterocycle may be optionally substituted with 1 to 3 substituents selected from the group consisting of halo, C1-C5 alkyl, halo-C1-C5 alkyl, C3-C8 cycloalkyl, C1-C5 alkylamino, di(C1-C5 alkyl)amino, and (C1-C5 alkyl)(C1-C5 alkoxycarbonyl)amino.

5. In paragraph 1, R2 and R3 are each independently -H, C1-C5 alkyl, halo-C1-C5 alkyl or C6-C 10 A compound or a pharmaceutically acceptable salt or isomer thereof, characterized in that it is aryl, provided that R2 and R3 are not both -H; wherein the aryl can be optionally substituted with one to three substituents selected from the group consisting of hydroxy, halo, amino, and C1-C7 alkyl.

6. A compound or a pharmaceutically acceptable salt or isomer thereof, characterized in that R5 is amino in the first paragraph.

7. In the first paragraph, a compound characterized in that the compound of the chemical formula 1 is selected from the group consisting of the following compounds or a pharmaceutically acceptable salt or isomer thereof: 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylpiperazin-1-yl)-1,3,5-triazin-2-amine; 4-(2-(tert-butyl)hydrazinyl)-6-(4-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine; 4-(2-(tert-butyl)hydrazinyl)-6-(4-isopropylpiperidin-1-yl)-1,3,5-triazin-2-amine; 4-(2-(tert-butyl)hydrazinyl)-6-(4-isopropylpiperazin-1-yl)-1,3,5-triazin-2-amine; 6-(2-(tert-butyl)hydrazinyl)-N 2 -(4-Fluorobenzyl-1-yl)-1,3,5-triazine-2,4-diamine; tert-Butyl(1-(4-amino-6-(2-(3-chlorophenyl)hydrazinyl)-1,3,5-triazin-2-yl)azetidine-3yl)(methyl)carbamate; 4-(2-(3-chlorophenyl)hydrazinyl)-6-(3-methylamino)azetidin-1-yl)-1,3,5-triazin-2-amine; 4-(2-(tert-butyl)hydrazinyl)-6-(4-cyclohexylpiperidin-1-yl)-1,3,5-triazin-2-amine; 6-(2-(tert-butyl)hydrazinyl)- N 2 -(cyclopropylmethyl)-1,3,5-triazin-2-amine; 6-(2-(tert-butyl)hydrazinyl)- N 2 -(2-dimethylamino)ethyl)-1,3,5-triazine-2,4-diamine; 6-(2-(tert-butyl)hydrazinyl)- N 2 -(4-dimethylamino)benzyl)-1,3,5-triazine-2,4-diamine; 4-(2-isopropylhydrazinyl)-6-(3-methylamino)azetidin-1-yl-1,3,5-triazin-2-amine; and 6-(2-(tert-butyl)hydrazinyl)-N 2 -(2-morpholineethyl)-1,3,5-triazine-2,4-diamine.

8. A pharmaceutical composition comprising a therapeutically effective amount of a compound according to any one of claims 1 to 7, or a pharmaceutically acceptable salt or isomer thereof, as an active ingredient, together with a pharmaceutically acceptable carrier.

9. A pharmaceutical composition for preventing or treating a disease associated with human histamine 4 receptor (hH4R) according to claim 8.

10. A pharmaceutical composition according to claim 9, characterized in that the disease associated with the human histamine 4 receptor (hH4R) is selected from the group consisting of nasal polyps, allergic rhinitis, non-allergic rhinitis, viral rhinitis, nasal itching, sinusitis, nasal congestion (stuffy nose), asthma, chronic obstructive pulmonary disease (COPD), rheumatoid arthritis, atopic dermatitis, psoriasis, eczema, pruritus, skin itching, urticaria, idiopathic chronic urticaria, scleroderma, allergic conjunctivitis, keratoconjunctivitis, ophthalmitis, dry eye, diabetic retinopathy, heart failure, arrhythmia, atherosclerosis, multiple sclerosis, inflammatory bowel disease, inflammatory pain, neuropathic pain, osteoarthritis pain, thyroid autoimmune disease, immune-mediated diabetes, lupus, post-surgical adhesion formation, cancer, and vestibular disorder.

11. A pharmaceutical composition according to claim 10, characterized in that the inflammatory bowel disease is colitis, Crohn's disease or ulcerative colitis.

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