Inhibitors of nek7 kinase

Abstract

A compound is provided that has activity as an inhibitor of NEK7. The compound has structure (I): or a pharmaceutically acceptable salt, stereoisomer, or prodrug thereof, wherein A, X, Y, R 1 R 2 R 3 and R 4 As defined herein, methods for preparing and using such compounds, pharmaceutical compositions comprising such compounds, and methods for modulating NLRP3 inflammasome activity are also provided.

Description

background Technical Field

[0002] Embodiments of this disclosure generally relate to compounds, methods of their preparation, and their use as therapeutic or preventative agents, for example, for treating inflammation.

[0003] Description of related technologies

[0004] Inflammatory bodies are multiprotein complexes whose activation plays a central role in innate immunity and inflammation. To date, four inflammatory bodies have been described: NLRP1, NLRC4, NLRP3, and AIM2. The NLRP3 inflammatory body is composed of NLRP3, ASC, and caspase-1. Its activation leads to the activation of caspase-1, which promotes the secretion of IL-1β and IL-18, cytokines that mediate inflammation in animal disease models of several autoimmune diseases, myocardial infarction, metabolic syndrome, inflammatory bowel disease, and macrophage activation syndrome.

[0005] NEK7 is a member of the NIMA-associated kinase (NEK) family and acts as an NLRP3-binding protein that regulates its oligomerization and activation. NEK7 is a serine / threonine kinase essential for mitotic entry, cell cycle progression, cell division, and the process of mitosis. It is expressed in various tissues, such as the brain, heart, lung, liver, and spleen. Overexpression of NEK7 induces the generation of abnormal cells, which are closely associated with tumors such as retinoblastoma, gallbladder cancer, and head and neck cancer.

[0006] Numerous inhibitors have been widely used to interfere with effector signaling pathways involving IL-1β or IL-18 without eliminating the inflammatory response. Inhibitors that block NLRP3 inflammasome activation through NLRP3-NEK7 interaction have therapeutic or preventative activity in several human diseases, such as type 2 diabetes (T2D), atherosclerosis, gout, and neurodegenerative diseases. However, the exact mechanisms of NLRP3-NEK7 interaction are not fully understood.

[0007] Therefore, there is a need to develop inhibitors that directly target NEK7 to influence the inflammatory response regulated by the NLRP3 inflammasome in several pathological diseases, such as gout, atherosclerosis, type 2 diabetes, metabolic syndrome, macular degeneration, Alzheimer's disease, multiple sclerosis, and inflammatory bowel disease. Embodiments of this disclosure meet this need and offer other related advantages. Summary of the Invention

[0008] In short, embodiments of this disclosure provide compounds capable of modulating the activity of the NLRP3 inflammasome, including pharmaceutically acceptable salts, stereoisomers, and prodrugs thereof.

[0009] In one aspect, the present invention provides compounds of structure (I):

[0010]

[0011] Its acceptable salts, stereoisomers, or prodrugs, wherein A, X, Y, R 1 R 2 R 3 and R 4 Each is defined as follows.

[0012] On the other hand, pharmaceutical compositions comprising the disclosed compounds and methods of use for treating inflammation are also provided. Detailed Implementation

[0013] In the following description, certain specific details are set forth to provide a thorough understanding of various embodiments of this disclosure. However, those skilled in the art will understand that this disclosure can be practiced without these details.

[0014] Unless the context otherwise requires, throughout the specification and claims, the word “comprise” and its variations (e.g., “comprises” and “comprising”) are interpreted as having an open-ended, inclusive meaning, that is, as “including, but not limited to”.

[0015] In this specification, any concentration range, percentage range, ratio range, or integer range shall be understood to be any integer value included within the range, and, where appropriate, to include fractions (e.g., one-tenth and one-hundredth of an integer), unless otherwise stated. As used herein, the terms “about” and “approximately” mean ±20%, ±10%, ±5%, or ±1% of the range, value, or structure indicated, unless otherwise stated. It should be understood that the terms “a” and “an” as used herein refer to “one or more” of the enumerated components. The use of alternatives (e.g., “or”) shall be understood to mean one or both of the alternatives or any combination thereof.

[0016] Throughout this specification, the phrase "in one embodiment" or "in an embodiment" means that a specific feature, structure, or characteristic described with respect to that embodiment is included in at least one embodiment of the invention. Therefore, the phrases "in one embodiment" or "in an embodiment" appearing in various places throughout this specification do not necessarily all refer to the same embodiment. Furthermore, specific features, structures, or characteristics may be combined in one or more embodiments in any suitable manner.

[0017] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. As used in this specification and claims, the singular forms “a,” “an,” and “the” include a plurality of indicators unless the context clearly specifies otherwise.

[0018] "Amino" refers to the -NH2 group.

[0019] "Carboxy" refers to the -CO2H group.

[0020] "Cyano" refers to the -CN group.

[0021] "Hydroxy" (hydroxy / hydroxyl) refers to the -OH group.

[0022] "Nitro" refers to the -NO2 group.

[0023] "Oxyto" refers to the =O substituent.

[0024] "Thiol" refers to the -SH substituent.

[0025] "Thio" refers to the =S substituent.

[0026] "Alkyl" refers to a saturated straight-chain or branched hydrocarbon chain group consisting only of carbon and hydrogen atoms, having one to twelve carbon atoms (C1-C2). 12 Alkyl groups (C1-C8 alkyl), consisting of one to eight carbon atoms (C1-C6 alkyl) or one to six carbon atoms (C1-C6 alkyl), or any value within these ranges (e.g., C4-C6 alkyl, etc.), and connected to the remainder of the molecule by a single bond, such as methyl, ethyl, n-propyl, 1-methylethyl (isopropyl), n-butyl, n-pentyl, 1,1-dimethylethyl (tert-butyl), 3-methylhexyl, 2-methylhexyl, etc. The number of carbons mentioned refers to both main-chain and branched carbons, but excludes carbon atoms belonging to any substituent. Unless otherwise specified in the specification, alkyl groups may optionally be substituted.

[0027] "Alkenyl" refers to an unsaturated straight-chain or branched hydrocarbon chain group composed only of carbon and hydrogen atoms, containing one or more carbon-carbon double bonds and having two to twelve carbon atoms (C2-C4). 12The alkenyl group consists of two to eight carbon atoms (C2-C8 alkenyl) or two to six carbon atoms (C2-C6 alkenyl), or any value within these ranges, and is connected to the rest of the molecule by a single bond, such as vinyl, propenyl, butenyl, pentenyl, pent-1,4-dienyl, etc. The number of carbons mentioned refers to both main-chain and branched carbons, but excludes carbon atoms belonging to any substituent. Unless otherwise specifically stated in this specification, the alkenyl group may optionally be substituted.

[0028] The term "alkynyl" refers to an unsaturated straight-chain or branched hydrocarbon group having 2 to 12 carbon atoms (C2-C4). 12 The alkynyl group has 2 to 9 carbon atoms (C2-C9 alkynyl) or 2 to 6 carbon atoms (C2-C6 alkynyl) or any value within these ranges, and has at least one carbon-carbon triple bond. Examples of alkynyl groups may be selected from ethynyl, propynyl, but-1-alkynyl, but-2-alkynyl, etc. The number of carbons mentioned refers to both main-chain and branched carbons, but does not include carbon atoms belonging to any substituent. Unless otherwise specifically stated in this specification, the alkynyl group may optionally be substituted.

[0029] "Alkoxy" refers to the formula -OR a The group, wherein R a It is an alkyl group as defined above, containing one to twelve carbon atoms (C1-C2). 12 Alkoxy groups, consisting of one to eight carbon atoms (C1-C8 alkoxy groups) or one to six carbon atoms (C1-C6 alkoxy groups), or any values ​​within these ranges. Unless otherwise specified in the specification, the alkoxy group may optionally be substituted.

[0030] "Amino group" refers to the formula -NR a R b The group, wherein R a It is H or C1-C6 alkyl, and R b It is a C1-C6 alkyl group as defined above. Unless otherwise stated, the C1-C6 alkyl portion of the amino group may optionally be substituted.

[0031] "Aminoalkylcycloalkyl group" refers to the formula -R a R b NR c R d The group, wherein R a It is a cyclic hydrocarbon group as defined in this article, R b It is a C1-C6 alkyl group, R c It is H or C1-C6 alkyl, and R d It is a C1-C6 alkyl group as defined above. Unless otherwise stated, the cyclic hydrocarbon group of the aminoalkyl cyclic hydrocarbon group and each C1-C6 alkyl moiety may optionally be substituted.

[0032] "Aromatic ring" refers to a ring-shaped planar molecule or part of a molecule (i.e., a group) with resonant bonds that exhibits increased stability relative to other arrangements of atoms with the same atomic set. Typically, an aromatic ring contains a set of covalently bonded coplanar atoms and includes a large number of π-electrons (e.g., alternating double and single bonds), said π-electrons being an even number but not a multiple of 4 (i.e., 4n+2 π-electrons, where n = 0, 1, 2, 3, etc.). Aromatic rings include, but are not limited to, phenyl, naphthyl, imidazolyl, pyrrolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridinyl, pyridazinyl, and pyrimidinyl groups. Unless otherwise specifically stated in the specification, "aromatic ring" includes all optionally substituted groups.

[0033] "Aryl" refers to a group containing 6 to 18 carbon atoms (e.g., 6 to 10 carbon atoms (C6-C)). 10 A carbocyclic system consisting of an aryl group and at least one carbocyclic aromatic ring. For the purposes of embodiments of the invention, the aryl group is a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which may include fused or bridged ring systems. Aryl groups include, but are not limited to, those derived from: anthracene, acenaphthene, phenanthrene, anthracene, azulene, benzene, etc. Fluoranthene, fluorene, asymmetric indole, symmetric indole, indene, indene, naphthalene, phenanthracene, heptane, pyrene, and benzo[a]phenanthrene. Unless otherwise specified in the specification, the aryl group may optionally be substituted.

[0034] "Cyanoalkyl" means an alkyl group containing at least one cyano substituent. The -CN substituent may be on a primary, secondary, or tertiary carbon. Unless otherwise specified in the specification, the cyanoalkyl group may optionally be substituted.

[0035] "Carbocyclic" refers to a ring system in which each atom in the ring is carbon.

[0036] "Cyclic hydrocarbon group" refers to a non-aromatic monocyclic or polycyclic carbocyclic group consisting only of carbon and hydrogen atoms. It may include fused or bridged ring systems and has three to fifteen ring carbon atoms (C3-C4). 15 Cyclic hydrocarbon group), three to ten cyclic carbon atoms (C3-C) 10 The cyclic group consists of a cycloalkyl group (C3-C8 cycloalkyl group) or three to eight carbon atoms (C3-C4 cycloalkyl group) or any value within these ranges, such as three to four carbon atoms (C3-C4 cycloalkyl group), and is saturated or partially unsaturated and connected to the rest of the molecule by a single bond. Monocyclic groups include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Polycyclic groups include, for example, adamantyl, norbornyl, decahydronaphthyl, 7,7-dimethyl-bicyclo[2.2.1]heptyl, etc. Unless otherwise specifically stated in the specification, the cyclic alkyl group may optionally be substituted.

[0037] "alkyl cyclohydride group" refers to the formula -R a R b The group, wherein R a It is a cyclic hydrocarbon group and R b It is an alkyl group as defined above. Unless otherwise specified in the specification, the alkyl cyclic hydrocarbon group may optionally be substituted.

[0038] "Fusing" refers to any ring structure fused to another ring structure as described in this article.

[0039] "Halogen" refers to bromine, chlorine, fluorine, or iodine.

[0040] "Halogenated alkyl" refers to an alkyl group as defined above that is substituted with one or more halogen groups as defined above, such as trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl, 1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, etc. Unless otherwise specified in the specification, the halogenated alkyl group may optionally be substituted.

[0041] "Halogenated cyclic hydrocarbon group" refers to a cyclic hydrocarbon group as defined above, which is substituted with one or more halogen groups as defined above, such as trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl, 1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, etc. Unless otherwise specified in the specification, the halogenated cyclic hydrocarbon group may optionally be substituted.

[0042] "Halogenated alkyl cycloalkyl group" refers to the formula -R a R b The group, wherein R a It is a cyclic hydrocarbon group and R b It is a haloalkyl group as defined above. Unless otherwise specified in the specification, the haloalkyl cyclic hydrocarbon group may optionally be substituted.

[0043] "Halogenated cycloalkyl group" refers to the formula -R a R b The group, wherein R a It is an alkyl group and R b It is a halocyclic alkyl group as defined above. Unless otherwise specified in the specification, the halocyclic alkyl group may optionally be substituted.

[0044] "Heterocyclic cyclic hydrocarbon group" refers to the formula -R a R b The group, wherein R a It is a cyclic hydrocarbon group and R b It is a heterocyclic group as defined herein. Unless otherwise specified in the specification, the heterocyclic cyclic hydrocarbon group may optionally be substituted.

[0045] "Hydroxyalkyl" refers to an alkyl group as defined above, which is substituted with one or more hydroxyl groups. The hydroxyalkyl group is attached to the main chain via an alkyl carbon atom. Unless otherwise specified in the specification, the hydroxyalkyl group may optionally be substituted.

[0046] "Heterocyclic group" refers to a 3-18 membered, such as 3-10 or 3-8 membered, non-aromatic cyclic group having one to ten cyclic carbon atoms (e.g., two to ten) and one to six cyclic heteroatoms selected from nitrogen, oxygen, and sulfur. Unless otherwise specifically stated in this specification, a heterocyclic group is a partially or fully saturated and monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which may include fused, spirocyclic, and / or bridged ring systems. The nitrogen, carbon, and sulfur atoms in the heterocyclic group are optionally oxidized, and the nitrogen atom is optionally quaternized. Examples of such heterocyclic groups include, but are not limited to, dioxacyclopentyl, thienyl[1,3]dithiaalkyl, decahydroisoquinolinyl, furanoneyl, imidazolinyl, imidazoalkyl, isothiazolinyl, isoxazolinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, hexahydro-1H-pyrrolizine, 2-oxopiperidinyl, 2-oxopiperidinyl, 2-oxopiperidylalkyl, oxazolinyl, ethylene oxide, piperidinyl, piperazinyl, 4-piperidinoneyl, azacyclic butyl, pyrrolylalkyl, pyrazolyl, quininecycloyl, thiazoalkyl, tetrahydrofuranyl, trithiaalkyl, tetrahydropyranyl, thiomorpholinyl, thio-morpholinyl, 1-oxo-thiomorpholinyl, and 1,1-dioxo-thiomorpholinyl. Unless otherwise specifically stated in the specification, the heterocyclic group may optionally be substituted.

[0047] "Halogenated heterocyclic group" refers to a heterocyclic group containing at least one halogen substituent. The halogen substituent may be located on a primary, secondary, or tertiary carbon. Unless otherwise specified in the specification, the halogenated heterocyclic group may be optionally substituted.

[0048] "Halogenated heterocyclic alkyl" refers to formula -R a R b The group, wherein R a It is an alkyl group and R b It is a haloheterocyclic group as defined herein. Unless otherwise specified in the specification, the alkyl group of the haloheterocyclic group may optionally be substituted.

[0049] "Heterocyclic alkyl" refers to formula -R a R b The group, wherein R a It is an alkyl group and R b It is a heterocyclic group as defined herein. Unless otherwise specified in the specification, the heterocyclic alkyl group may optionally be substituted.

[0050] "Heteroaryl" refers to a 5-18 membered, such as a 5-6 membered ring system group, which contains one to thirteen ring carbon atoms, one to six cyclic heteroatoms selected from nitrogen, oxygen, and sulfur, and at least one aromatic ring. Heteroaryl can be a monocyclic, bicyclic, tricyclic, or tetracyclic system, which may include fused or bridged ring systems; and the nitrogen, carbon, or sulfur atom in the heteroaryl may optionally be oxidized; the nitrogen atom may optionally be quaternized. Examples include, but are not limited to, azirzoyl, acridineyl, benzimidazolyl, benzothiazoyl, benzoindolyl, benzodioxacyclopentenyl, benzofuranyl, benzooxazolyl, benzothiazoyl, benzothiadiazolyl, benzo[b][1,4]dioxazolyl, 1,4-benzodioxane, benzonaphthuryl, benzooxazolyl, benzodioxacyclopentenyl, benzodioxacyclohexenyl, benzopiperanyl, benzopiperanone, benzofuranyl, benzofuranone, benzothienyl / benzothiophenyl, benzotriazolyl, benzo[4,6]imidazo[1,2-a]pyridyl, carbazole, cenylyl, and dibenzofuran. The following are listed: yl, dibenzothiophene, furanyl, isothiazolyl, imidazolyl, indazole, indolyl, indazole, isoindolyl, indololinyl, isoindololinyl, isoquinolinyl, indolazinyl, isoxazolyl, naphridinyl, oxadiazolyl, 2-oxozadiazolyl, oxazolyl, 1-pyridinyl oxide, 1-pyridazinyl oxide, 1-phenyl-1H-pyrroleyl, phenazinyl, phenothiazinyl, phenotoxazinyl, phthalazinyl, pteridinyl, purine, pyrroleyl, pyrazolyl, pyridinyl, pyridinyl, pyridinyl, pyrimidinyl, pyridazinyl, quinazolinyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl, triazinyl, and thiophene (i.e., thiacyclopentadienyl). Unless otherwise specified in this specification, heteroaryl groups may optionally be substituted.

[0051] The terms oxazolyl, isoxazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, and 1,3,4-thiadiazolyl refer to the following structures respectively:

[0052]

[0053] Among them, oxazolyl, isoxazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl and 1,3,4-thiadiazolyl are derived from oxazolyl, isoxazolyl, 1, One of the carbon atoms in the rings of 2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, and 1,3,4-thiadiazolyl is covalently bonded to the rest of the molecule.

[0054] As used herein, the term "substituted" means a group (e.g., alkyl, alkenyl, alkylene, alkylcarbonyl, alkoxy, alkoxyalkyl, aminoalkyl, aryl, cyanoalkyl, cycloalkyl, haloalkyl, heterocyclic, heterocyclic alkenyl, heterocyclic alkyl, heteroaryl, heteroarylalkyl, and / or hydroxyalkyl) in which at least one hydrogen atom (e.g., 1, 2, 3, or all hydrogen atoms) is replaced by a bond with a non-hydrogen substituent. Examples of non-hydrogen substituents include, but are not limited to, amino, carboxyl, cyano, hydroxy, halogen, nitro, oxo, thiol, thio, alkyl, alkenyl, alkylcarbonyl, alkoxy, aryl, cyanoalkyl, cycloalkyl, haloalkyl, heterocyclic, heterocyclic alkyl, heteroaryl, heteroarylalkyl, and / or hydroxyalkyl substituents, each of which may also be optionally substituted by one or more of the aforementioned substituents.

[0055] In some specific embodiments, the optional substituents are independently selected from halogens, hydroxyl groups, cyano groups, amino groups, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cyclic hydrocarbon groups, C3-C8 halocyclic hydrocarbon groups, C6-C 10 Aryl, 5- or 6-membered heteroaryl, C1-C6 alkoxy and 3-8-membered heterocyclic groups.

[0056] The term "effective amount" or "therapeutic effective amount" refers to the amount of the compound described herein sufficient to achieve the intended application, including but not limited to the treatment of diseases as defined below. The therapeutic effective amount can vary depending on the intended therapeutic application (in vivo), or the individual being treated and the disease condition, such as the individual's weight and age, the severity of the disease condition, the method of administration, etc., which can be readily determined by one of ordinary skill in the art. The term also applies to doses that induce specific responses in target cells, such as a reduction in platelet adhesion and / or cell migration. Specific doses will vary depending on the specific compound selected, the dosing regimen followed, whether it is administered in combination with other compounds, the timing of administration, the tissue to which it is administered, and the physical delivery system it carries.

[0057] As used herein, “treatment” or “therapeutic effect” means a method for obtaining a beneficial or desired outcome (including, but not limited to, therapeutic and / or preventative effects) in relation to a disease, condition, or medical condition. A therapeutic benefit means the eradication or improvement of the underlying condition being treated. Furthermore, a therapeutic benefit is achieved by eradicating or improving one or more physiological symptoms associated with the underlying condition, thereby observing improvement in an individual, although the individual may still suffer from the underlying condition. Preventative effects include delaying or eliminating the onset of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, stopping, or reversing the progression of a disease or condition, or any combination thereof. In some embodiments, for a preventative benefit, the composition is administered to an individual at risk of having a particular disease, or to an individual reporting one or more physiological symptoms of a disease, even if the disease may not yet have been diagnosed.

[0058] As used herein, the terms “co-administration,” “combination administration,” and their grammatical equivalents cover the administration of two or more agents, including humans, such that the two agents and / or their metabolites are simultaneously present in the individual. Co-administration includes simultaneous administration of a single composition, administration of a single composition at different times, or administration of a composition in which both agents are present.

[0059] "Drug-acceptable salts" include both acid addition salts and base addition salts.

[0060] "Pharmaceutical-acceptable acid addition salts" refer to those salts that retain the biological efficacy of the free base, are biologically tolerable, or otherwise biologically suitable for administration to an individual. Generally, see SMBerge et al., "Pharmaceutical Salts," J. Pharm. Sci., 1977, 66:1-19, and Handbook of Pharmaceutical Salts, Properties, Selection, and Use, Stahl and Wermuth, eds., Wiley-VCH and VHCA, Zurich, 2002. Preferred pharmaceutical-acceptable acid addition salts are those that are pharmacologically effective and suitable for contact with patient tissues without adverse toxicity, irritation, or anaphylactic reactions. The acceptable acid addition salts for pharmaceuticals are formed from: inorganic acids, such as, but not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; and organic acids, such as, but not limited to, acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid, ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid, 4-acetamidobenzoic acid, camphoric acid, camphor-10-sulfonic acid, decanoic acid, hexanoic acid, caprylic acid, carbonic acid, cinnamic acid, citric acid, cyclohexanesulfonic acid, dodecanesulfonic acid, ethane-1,2-disulfonic acid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, formic acid, fumaric acid, galactosic acid, gentianic acid, glucosamine. Acids, including glucuronic acid, glucuronic acid, glutamic acid, glutamate, 2-oxoglutamate, glycerophosphate, glycolic acid, hippuric acid, isobutyric acid, lactic acid, lactobionic acid, lauric acid, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, mucoic acid, naphthalene-1,5-disulfonic acid, naphthalene-2-sulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinic acid, oleic acid, orotic acid, oxalic acid, palmitic acid, dihydroxynaphthalic acid, propionic acid, pyroglutamic acid, pyruvic acid, salicylic acid, 4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid, tartaric acid, thiocyanate, p-toluenesulfonic acid, trifluoroacetic acid, and undecenoic acid, etc.

[0061] "Pharmaceutical-acceptable base addition salts" refer to those salts that retain the biological efficacy of the free base, are biologically tolerable, or otherwise biologically suitable for administration to an individual. Generally, see SMBerge et al., "Pharmaceutical Salts," J. Pharm. Sci., 1977, 66:1-19, and Handbook of Pharmaceutical Salts, Properties, Selection, and Use, Stahl and Wermuth, eds., Wiley-VCH and VHCA, Zurich, 2002. Preferred pharmaceutical-acceptable base addition salts are those that are pharmacologically effective and suitable for contact with patient tissues without adverse toxicity, irritation, or anaphylactic reactions. Pharmaceutical-acceptable base addition salts are prepared by adding an inorganic or organic base to a free acid. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Preferred inorganic salts are ammonium, sodium, potassium, calcium, and magnesium salts. Salts derived from organic bases include, but are not limited to, salts of the following substances: primary amines, secondary amines, tertiary amines, substituted amines, including naturally occurring substituted amines, cyclic amines, and base ion exchange resins, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, diethanolamine, ethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-ethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, phenethylbenzylamine, benzathine penicillin, ethylenediamine, glucosamine, methylglucosamine, theobromine, triethanolamine, tromethamine, purine, piperazine, piperidine, N-ethylpiperidine, polyamine resins, etc. Particularly preferred organic bases are isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline, and caffeine.

[0062] In some implementations, pharmaceutically acceptable salts include quaternary ammonium salts, such as quaternary ammonium alkyl halides (e.g., methyl bromide).

[0063] The terms “antagonist” and “inhibitor” are used interchangeably, and they refer to compounds that have the ability to inhibit the biological function of a target protein, whether by inhibiting the activity or expression of the protein (e.g., the NLRP3 inflammasome or NEK7) or by inhibiting the association between the NLRP3 inflammasome and NEK7. Therefore, the terms “antagonist” and “inhibitor” are defined in the context of the biological action of the target protein. While preferred antagonists herein interact specifically with the target (e.g., bind), compounds that inhibit the biological activity of a target protein by interacting with other members of a signal transduction pathway in which the target protein is a member are also included in this definition. Preferred biological activities inhibited by antagonists are associated with tumor development, growth, or spread.

[0064] As used herein, the term "agonist" refers to a compound that has the ability to initiate or enhance the biological function of a target protein, whether or not by inhibiting the activity or expression of the target protein. Therefore, the term "agonist" is defined in the context of the biological action of the target peptide. While preferred agonists herein interact particularly with the target (e.g., bind), compounds that initiate or enhance the biological activity of the target peptide by interacting with other members of the signal transduction pathway in which the target peptide is a member are also included in this definition.

[0065] "Signal transduction" is the process by which stimulating or inhibitory signals are transmitted into and within cells to trigger intracellular responses.

[0066] The term "selective inhibition" or "selective inhibition" refers to the ability of a bioactive agent to preferentially reduce the target signal transduction activity compared to off-target signal transduction activity through direct or indirect interaction with the target.

[0067] "Individual" refers to an animal, such as a mammal, like a human. The methods described herein can be used for human treatment and veterinary applications. In some embodiments, the individual is a mammal, and in some embodiments, the individual is a human.

[0068] "Mammals" include humans and livestock as well as non-livestock animals, such as laboratory animals and domestic pets (e.g., cats, dogs, pigs, cattle, sheep, goats, horses, rabbits), and non-livestock animals such as wild animals.

[0069] The term “prodrug” is intended to refer to a compound (e.g., a compound of structure (I)) that can be converted into the bioactive compound described herein under physiological conditions or by solvent degradation. Thus, the term “prodrug” refers to a precursor of a pharmaceutically acceptable bioactive compound. In some respects, prodrugs are inert when administered to a subject but are converted into the active compound in vivo, for example, by hydrolysis. Prodrug compounds often offer advantages such as solubility, tissue compatibility, or delayed release in mammalian organisms (see, for example, Bundgard, H., Design of Prodrugs (1985), pp. 7–9, 21–24 (Elsevier, Amsterdam). In Higuchi, T., et al., “Pro-drugs as Novel Delivery Systems,” ACSSymposium Series, Vol. 14, and in Bioreversible Carriers in Drug Design, edited by Edward B. Roche, American Pharmaceutical Association and Pergamon). A discussion of prodrugs is provided in Press, 1987, and both are incorporated herein by reference in their entirety. The term "prodrug" also means any covalently bonded carrier that releases the active compound in vivo when such a prodrug is administered to a mammalian individual. Prodrugs of the active compounds described herein are typically prepared by modifying functional groups present in the active compound in such a way that the modification cleaves into the parent active compound either in a conventional manner or in vivo. Prodrugs include compounds in which a hydroxyl, amino, or thiol group is attached to any group that cleaves, respectively, to form a free hydroxyl, free amino, or free thiol group when the prodrug of the active compound is administered to a mammalian individual. Examples of prodrugs include, but are not limited to, acetic acid, formic acid, and benzoic acid derivatives with hydroxyl functional groups in the active compound, or acetamide, formamide, and benzamide derivatives with amine functional groups.

[0070] The term "in vivo" refers to events that occur within an individual's body.

[0071] The implementation schemes disclosed herein also refer to compounds covering all pharmaceutically acceptable structures (I).

[0072] Some embodiments also refer to in vivo metabolites of the disclosed compounds. Such products can be generated, for example, by oxidation, reduction, hydrolysis, amidation, esterification, etc., of the applied compound, primarily due to enzymatic processes. Therefore, embodiments include compounds produced by methods comprising administering the compound of this disclosure to mammals for a duration sufficient to produce its metabolites. Such products are typically identified by administering a detectable dose of the radiolabeled compound of this disclosure to animals (e.g., rats, mice, guinea pigs, monkeys) or to humans, allowing sufficient time for metabolism, and by isolating the metabolites from urine, blood, or other biological samples.

[0073] "Stable compound" and "stable structure" mean a compound that is stable enough to be isolated from the reaction mixture to a useful purity and formulated into an effective therapeutic agent.

[0074] Typically, crystallization produces solvates of the compounds disclosed herein. As used herein, the term "solvate" refers to an aggregate comprising one or more compound molecules of this disclosure and one or more solvent molecules. In some embodiments, the solvent is water, in which case the solvate is a hydrate. Alternatively, in other embodiments, the solvent is an organic solvent. Thus, the compounds of this disclosure can exist as hydrates, including monohydrates, dihydrates, hemihydrates, sesquihydrates, trihydrates, tetrahydrates, etc., and corresponding solvated forms. The compounds of this disclosure can be true solvates, while in other cases, the compounds of this disclosure retain only the external water or are a mixture of water and some external solvent.

[0075] "Optional" or "optionally" means that the event described below may or may not occur, and that the description includes examples of both the occurrence and non-occurrence of said event or situation. For example, "optionally substituted aryl" means that the aryl group may or may not be substituted, and that the description includes substituted aryl groups and unsubstituted aryl groups.

[0076] "Pharmaceutical composition" refers to a formulation of the compounds of this disclosure with a medium generally accepted in the art for delivering the compounds of this disclosure to mammals (e.g., humans). Such media include all pharmaceutically acceptable carriers, diluents, or excipients used therein.

[0077] "Drug-acceptable carriers, diluents, or excipients" include, but are not limited to, any adjuvants, carriers, excipients, glidants, sweeteners, diluents, preservatives, dyes / colorants, flavor enhancers, surfactants, wetting agents, dispersants, suspending agents, stabilizers, isotonic agents, solvents, or emulsifiers.

[0078] "Stereoisomers" are compounds composed of atoms bonded by the same bonds but with different three-dimensional structures, and these different three-dimensional structures are not interchangeable. This disclosure contemplates various stereoisomers and mixtures thereof and includes "enantiomers," which are two stereoisomers whose molecules are mirror images of each other and are not superimposed on each other.

[0079] The compounds of this disclosure (i.e., compounds of structure (I)) or pharmaceutically acceptable salts thereof may contain one or more geometrically asymmetric centers and thus may produce stereoisomers, such as enantiomers, diastereomers, and other stereoisomeric forms defined in absolute stereochemistry as (R)- or (S)-, or (D)- or (L)- of an amino acid. Embodiments therefore include all such possible isomers, as well as their racemic and optically pure forms. Optional active (+) and (-), (R)- and (S)- or (D)- and (L)- isomers may be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques such as chromatography and fractional crystallization. Conventional techniques for preparing / separating individual enantiomers include chiral synthesis from suitable optically pure precursors or resolution of racemic mixtures (or racemic mixtures of salts or derivatives) using, for example, chiral high-performance liquid chromatography (HPLC). When the compounds described herein contain olefinic double bonds or other geometrically asymmetric centers, this means that the compounds include E and Z geometric isomers, unless otherwise specified. Similarly, all tautomers are intended to be included.

[0080] Embodiments of this disclosure include all modes and morphologically restricted states of rotational isomers of the compounds of the invention. It also includes transisomers, which are stereoisomers resulting from steric rotation around a single bond, wherein energy differences caused by stereostrain or other contributing factors create rotational barriers high enough to allow the separation of individual conformational isomers. As an example, certain compounds of this disclosure may exist as mixtures of transisomers, or may be purified or enriched to contain only one transisomer.

[0081] In some embodiments, the compound of structure (I) is a mixture of enantiomers or diastereomers. In other embodiments, the compound of structure (I) is substantially an enantiomer or diastereomer.

[0082] "Tautomerism" refers to the proton migration from one atom of a molecule to another atom of the same molecule. Therefore, the embodiments include tautomerisms of the disclosed compounds.

[0083] The chemical nomenclature and structural formulas used in this paper are modified versions of the IUPAC nomenclature system, implemented using ACD Nomenclature version 9.07 and / or ChemDraw Professional version 17.0.0.206 (CambridgeSoft). For complex chemical names used in this paper, substituents are typically named before the groups they are attached to. For example, cyclopropylethyl comprises an ethyl backbone with cyclopropyl substituents. Unless otherwise described below, all bonds are identified in the chemical structural formulas used in this paper, but it is assumed that all bonds on some carbon atoms are bonded to a sufficient number of hydrogen atoms to complete the valence.

[0084] compound

[0085] This disclosure provides compounds capable of modulating the activity of the NLRP3 inflammasome, including its pharmaceutically acceptable salts, stereoisomers, and prodrugs.

[0086] The embodiments of this disclosure provide compounds having the following structure (I):

[0087]

[0088]

[0089] Or an acceptable salt, stereoisomer, or prodrug thereof, wherein:

[0090] A is each arbitrarily assigned to one or more R. 5 Replacement C6-C 10 Aryl, C3-C 10 Cyclic hydrocarbon groups, 3-10 membered heterocyclic groups, or 5-6 membered monocyclic heteroaryl groups;

[0091] X is N or CH;

[0092] Y is CHOH or NH;

[0093] R 1 It is an H or C1-C6 alkyl group;

[0094] R 2 It is a C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, 3-8 membered heterocyclic, or 5 or 6 membered heteroaryl group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy and 3-8 membered heterocyclic;

[0095] R 3It is selected from the following heteroaryl groups: oxazolyl, isoxazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, and 1,3,4-thiadiazolyl, wherein the substituents are selected from: amino, halogen, cyano, C1-C6 alkyl, ... C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C3-C8 cycloalkyl, C3-C8 alkylcycloalkyl, C3-C8 haloalkylcycloalkyl, C3-C8 aminoalkylcycloalkyl, C1-C6 cyanoalkyl, C1-C6 amino, C1-C6 hydroxyalkyl, 3-8 membered heterocyclic, 3-8 membered heterocyclic alkyl, 3-8 membered heterocyclic cycloalkyl, 3-8 membered halocyclic, 3-8 membered halocyclic alkyl, C3-C8 halocyclic and C3-C8 halocyclic alkyl and combinations thereof;

[0096] R 4 H is a C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cyclic hydrocarbon, 3-8 membered heterocyclic group, or C6-C alkyl group, each optionally substituted with one or more substituents. 10 aryl, or 5- or 6-membered heteroaryl, wherein the substituent is selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, and C1-C6 alkoxy; and

[0097] R 5 Each time it appears, it is independently a halogen, cyano, C1-C6 alkyl, C1-C6 hydroxyalkyl, or C1-C6 haloalkyl.

[0098] In some implementations of structure (I), A is optionally defined by one or more R. 5 Replacement C6-C 10 Aryl, C3-C 10 Cyclic hydrocarbon groups, 3-10 membered heterocyclic groups, or 5-6 membered monocyclic heteroaryl groups;

[0099] X is N or CH;

[0100] Y is CHOH or NH;

[0101] R 1 It is an H or C1-C6 alkyl group;

[0102] R 2It is a C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, 3-8 membered heterocyclic, or 5 or 6 membered heteroaryl group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy and 3-8 membered heterocyclic;

[0103] R 3 It is selected from the following heteroaryl groups: oxazolyl, isoxazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, and 1,3,4-thiadiazolyl, wherein the substituent is selected from: halogen, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C3-C8 cycloalkyl, and C3-C8 halocycloalkyl;

[0104] R 4 H is a C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cyclic hydrocarbon, 3-8 membered heterocyclic group, or C6-C alkyl group, each optionally substituted with one or more substituents. 10 aryl, or 5- or 6-membered heteroaryl, wherein the substituent is selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, and C1-C6 alkoxy; and

[0105] R 5 Each time it appears, it is independently a halogen, cyano, C1-C6 alkyl, C1-C6 alkyl, or C1-C6 haloalkyl.

[0106] One embodiment provides a compound of structure (I) or a pharmaceutically acceptable salt, stereoisomer, or prodrug thereof, wherein:

[0107] A is each arbitrarily assigned to one or more R. 5 Replacement C6-C 10 Aryl, C3-C 10 Cyclic hydrocarbon groups, 3-10 membered heterocyclic groups, or 5-6 membered monocyclic heteroaryl groups;

[0108] X is N or CH;

[0109] Y is CHOH or NH;

[0110] R 1 It is an H or C1-C6 alkyl group;

[0111] R 2H is a C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, 3-8 membered heterocyclic, or 5 or 6 membered heteroaryl group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy and 3-8 membered heterocyclic;

[0112] R 3 It is selected from the following heteroaryl groups: oxazolyl, isoxazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, and 1,3,4-thiadiazolyl, wherein the substituents are selected from: amino, halogen, cyano, C1-C6 alkyl, ... C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C3-C8 cycloalkyl, C3-C8 alkylcycloalkyl, C3-C8 haloalkylcycloalkyl, C3-C8 aminoalkylcycloalkyl, C1-C6 cyanoalkyl, C1-C6 amino, C1-C6 hydroxyalkyl, 3-8 membered heterocyclic, 3-8 membered heterocyclic alkyl, 3-8 membered heterocyclic cycloalkyl, 3-8 membered halocyclic, 3-8 membered halocyclic alkyl, C3-C8 halocyclic and C3-C8 halocyclic alkyl and combinations thereof;

[0113] R 4 H is a C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cyclic hydrocarbon, 3-8 membered heterocyclic group, or C6-C alkyl group, each optionally substituted with one or more substituents. 10 aryl, or 5- or 6-membered heteroaryl, wherein the substituent is selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, and C1-C6 alkoxy; and

[0114] R 5 Each time it appears, it is independently a halogen, cyano, C1-C6 alkyl, C1-C6 hydroxyalkyl, C1-C6 alkoxy, or C1-C6 haloalkyl.

[0115] In one implementation scheme, R 1 It is H. In other implementations, R 1 It is a C1-C6 alkyl group, such as methyl.

[0116] In one embodiment, a compound of structure (I) is provided, wherein R 2It is a branched C4-C6 alkyl, C3-C4 cyclic hydrocarbon, C3-C8 heterocyclic, or 5- or 6-membered heteroaryl group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, and 3-8-membered heterocyclic groups.

[0117] In another embodiment, a compound with structure (I) is provided, wherein R 2 It is a branched C4-C6 alkyl, C3-C4 cyclic hydrocarbon or C3-C8 heterocyclic group, each optionally substituted by one or more substituents, wherein the substituents are selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy and 3-8 membered heterocyclic groups.

[0118] In the specific implementation plan, R 2 It is a cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl group, each optionally substituted with one or more substituents selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, and 3-8 membered heterocyclic groups.

[0119] In different implementation schemes, R 2 It is a methyl, isopropyl, 2-methylpropyl, or allyl group, each optionally substituted with one or more substituents selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, and 3-8 membered heterocyclic groups.

[0120] In different implementation schemes, R 2 It is a methyl, ethyl, isopropyl, 2-methylpropyl, or allyl group, each optionally substituted with one or more substituents selected from halogens, hydroxyl groups, cyano groups, amino groups, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 alkoxy groups, and 3-8 membered heterocyclic groups.

[0121] In other implementation schemes, R 2 It is an oxocyclic butyl, tetrahydrofuranyl, tetrahydropyranyl, piperidinyl or tetrahydrothiophene group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy and 3-8 membered heterocyclic groups.

[0122] In other implementation schemes, R 2It is an oxetyl, tetrahydrofuranyl, tetrahydropyranyl, piperidinyl, aziridine, or tetrahydrothiophene group, each optionally substituted with one or more substituents, wherein the substituents are selected from halogens, hydroxyl groups, cyano groups, amino groups, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 alkoxy groups, and 3-8 membered heterocyclic groups.

[0123] In other implementation schemes, R 2 It is an oxocyclic butyl group, tetrahydrofuranyl group, tetrahydropyranyl group, piperidinyl group, azacyclic butyl group, pyrrolidinyl group, or tetrahydrothiophenyl group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, and 3-8 membered heterocyclic groups.

[0124] In a further implementation scheme, R 2 It is a pyridyl group optionally substituted with one or more substituents selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy and 3-8 membered heterocyclic groups.

[0125] In any of the foregoing embodiments, R 2 It is not replaced. In other of the aforementioned embodiments, R 2 It is substituted by one or more of hydroxyl and fluorine.

[0126] In any of the foregoing embodiments, R 2 It is not replaced. In other of the aforementioned embodiments, R 2 It is substituted by one or more of hydroxyl, methyl, methoxy and fluorine.

[0127] In a more specific implementation plan, R 2 It has one of the following structures:

[0128]

[0129] In a more specific implementation plan, R 2 It has one of the following structures:

[0130]

[0131] In a more specific implementation plan, R 2 It has one of the following structures:

[0132]

[0133] In some implementation schemes, R 2Optionally substituted with one or more substituents selected from: halogen, hydroxyl, cyano, amino, C1-C6 alkyl and 3-8 membered heterocyclic groups.

[0134] In some implementation schemes, R 2 It does not have the following structure:

[0135]

[0136] In any of the foregoing embodiments, R 3 It is an oxazolyl, isoxazolyl, 1,2,3-oxadiazolyl, or 1,3,4-oxadiazolyl group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogens, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C3-C8 cycloalkyl, and C3-C8 halocycloalkyl. For example, in some embodiments, R 3 It is an isoxazolyl group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C3-C8 cycloalkyl, and C3-C8 halocycloalkyl. In other specific embodiments, R 3 It is substituted by C1-C6 alkyl, C1-C6 haloalkyl, C3-C8 cyclic hydrocarbon or C3-C8 halocyclic hydrocarbon.

[0137] In other implementation schemes, R 3 It is an oxazolyl, isoxazolyl, 1,2,3-oxadiazolyl, 1,3,4-oxadiazolyl, thiazolyl, isothiazolyl, 1,2,4-thiadiazolyl, 1,3,4-thiadiazolyl or 1,2,4-triazolyl group, each optionally substituted with one or more substituents, wherein the substituents are selected from: halogens, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 haloalkyl, C3-C8 cycloalkyl, cyano, C1-C6 amino, C1-C6 hydroxyalkyl, 3-8 membered heterocyclic groups and C3-C8 halocyclic groups or combinations thereof.

[0138] In some implementations, R 3 It is an isoxazolyl group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cyclic hydrocarbon groups, cyano groups, C1-C6 amino groups, C1-C6 hydroxyalkyl groups, 3-8 membered heterocyclic groups, and C3-C8 halocyclic hydrocarbon groups or combinations thereof.

[0139] In some implementations, R 3It is a triazole group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cycloalkyl groups, cyano groups, C1-C6 amino groups, C1-C6 hydroxyalkyl groups, 3-8 membered heterocyclic groups, and C3-C8 halocyclic groups or combinations thereof.

[0140] In some implementations, R 3 It is an isothiazolyl group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cycloalkyl groups, cyano groups, C1-C6 amino groups, C1-C6 hydroxyalkyl groups, 3-8 membered heterocyclic groups, and C3-C8 halocyclic groups or combinations thereof.

[0141] In some implementations, R 3 It is a 1,2,4-thiadiazole group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cycloalkyl groups, cyano groups, C1-C6 amino groups, C1-C6 hydroxyalkyl groups, 3-8 membered heterocyclic groups, and C3-C8 halocyclic groups or combinations thereof.

[0142] In some implementations, R 3 It is a 1,3,4-thiadiazole group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cycloalkyl groups, cyano groups, C1-C6 amino groups, C1-C6 hydroxyalkyl groups, 3-8 membered heterocyclic groups, and C3-C8 halocyclic groups or combinations thereof.

[0143] In some implementations, R 3 It is a 1,3,4-oxadiazolyl group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cycloalkyl groups, cyano groups, C1-C6 amino groups, C1-C6 hydroxyalkyl groups, 3-8 membered heterocyclic groups, and C3-C8 halocyclic groups or combinations thereof.

[0144] In some implementations, R 3 It is a 1,2,4-triazolyl group optionally substituted with one or more substituents selected from: halogens, C1-C6 alkyl groups, C2-C6 alkenyl groups, C2-C6 alkynyl groups, C1-C6 haloalkyl groups, C3-C8 cycloalkyl groups, cyano groups, C1-C6 amino groups, C1-C6 hydroxyalkyl groups, 3-8 membered heterocyclic groups, and C3-C8 halocyclic groups or combinations thereof.

[0145] In other implementation schemes, R 3 It is substituted by C1-C6 alkyl, C1-C6 haloalkyl, C3-C8 cyclic hydrocarbon, cyano, C1-C6 amino, C1-C6 hydroxyalkyl, 3-8 membered heterocyclic group or C3-C8 halocyclic hydrocarbon or combination thereof.

[0146] In various implementation schemes, R 3 It has one of the following structures:

[0147]

[0148] In other implementation schemes, R 3 It has one of the following structures:

[0149]

[0150]

[0151] In other implementation schemes, R 3 It has one of the following structures:

[0152]

[0153]

[0154] In other implementation schemes, R 4 It is H. In other implementations, R 4 It is a C1-C6 alkyl group, such as methyl.

[0155] In some embodiments, Y is CHOH. In other embodiments, Y is NH.

[0156] In other embodiments, X is N. In even more embodiments, X is CH.

[0157] In various implementation schemes, A is optionally defined by one or more R. 6 Replacement C6-C 10 Aryl, C3-C 10 Cyclic hydrocarbon group or 5-6 membered monocyclic heteroaryl group. It should be understood that A is a divalent group.

[0158] In some implementations, A is a divalent, optionally substituted C. 6-10 Aryl group. In some embodiments, A is a divalent optionally substituted 3-8 membered saturated or partially unsaturated carbocyclic ring. In some embodiments, A is a divalent optionally substituted 3-10 membered heterocyclic ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. In some embodiments, A is a divalent optionally substituted 5-6 membered monocyclic heteroaryl ring having 1-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.

[0159] In some embodiments, A is a divalent group selected from phenyl, pyridyl, cyclohexyl and cyclohexenyl, which is optionally substituted.

[0160] In other embodiments, A is phenyl. In different embodiments, A is a saturated or unsaturated cyclohexane group. In many more embodiments, A is a pyridyl group.

[0161] In other embodiments, A is an optional substituted pyrimidinyl group.

[0162] In any of the foregoing embodiments, A is not substituted. In different foregoing embodiments, A is replaced by one or more R 5 Replacement. For example, in some implementations, R 5 It is a halogen. In other embodiments, R 5 It is fluorine. In other different implementations, R 5 It is chlorine.

[0163] In some implementation schemes, R 5 It is cyano. In some implementations, R 5 It is a C1-C6 alkyl group. In some embodiments, R 5 It is methyl. In some embodiments, R 5 It is a C1-C6 haloalkyl group. In some embodiments, R 5 It is difluoromethyl. In other embodiments, R 5 It is a C1-C6 hydroxyalkyl group. In some embodiments, R 5 It is -CH2OH.

[0164] In some embodiments, A is a divalent group selected from the following: phenyl, naphthyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, cycloheptyl, adamantyl, cyclooctyl, [3.3.0] dicyclooctyl, [4.3.0] dicyclononyl, [4.4.0] dicyclodecyl, [2.2.2] dicyclooctyl, fluorenyl, indanyl, tetrahydronaphthyl, acridineyl, acridineyl, benzimidazolyl, benzofuranyl, benzothiathiopheneyl, benzothiapheneyl, benzooxazolyl, benzothiazolyl, benzotriazolyl, benzotetrazoleyl, benzyl Isoxazolyl, benzoisothiazolyl, benzimidazolinyl, carbazole, NH-carbazole, carbolinyl, chromanyl, chromenyl, cenyl, decahydroquinolinyl, dithiazinyl, tetrahydrofuranyl, furanyl, furazanyl, imidazolyl, imidazolyl, imidazolyl, 1H-indazole, pseudoindolyl, indololinyl, indolazinyl, indolyl, 3-indolyl, isoindololinyl, isopyridine, isobenzofuranyl, isochoryl, isoindololinyl, isoindololinyl, isoindolyl, isoquinolinyl, isothiazolyl, isoxazolyl, morpholinyl, naphthidyl, octahydroisoquinolinyl, oxadiazole 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl; 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, oxazolylalkyl, oxazolyl, oxazolylalkyl, pyrimidinyl, phenanthridine, phenanthiazinyl, phenoxthiazinyl, phenoxazinyl, phthalazinyl, piperazinyl, piperidinyl, pteridinyl, purine, pyranyl, pyrazinyl, pyrazolylalkyl, pyrazolinyl, pyrazolyl, pyridazinyl, pyridoxazolyl, pyridothiazolyl, pyridothiazolyl, pyridinyl, pyridinyl, pyrrolylalkyl, pyrrololinyl, 2-pyrroleyl, pyrroleyl, quinazole Phinyl, quinolinyl, 4H-quinazinyl, quinoxalinyl, quininecycloyl, tetrahydrofuranyl, tetrahydroisoquinolinyl, tetrahydroquinolinyl, thiadiazinyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, thiaanthryl, thiazolyl, thienyl, thienothiazolyl, thienooxazolyl, thienoimidazolyl, thienyl, triazinyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,5-triazolyl, 1,3,4-triazolyl, oxacyclobutyl, aziroxacyclobutyl, and xanthonyl.

[0165] In a specific implementation plan, A has one of the following structures:

[0166] or

[0167]

[0168] In other specific implementations, A has one of the following structures:

[0169]

[0170] In some implementations, the compound of structure (I) is a regulator of the NLRP3 inflammasome.

[0171] In a specific implementation, the compound of structure (I) is an inhibitor of NEK7 in a patient or biological sample.

[0172] In various embodiments, the compounds have one of the structures set forth in Table 1 below, or a pharmaceutically acceptable salt, stereoisomer, or prodrug thereof. The compounds in Table 1 are prepared as described in the examples or by methods known in the art and measured by mass spectrometry and / or... 1 H NMR analysis.

[0173] Table 1. Representative compounds with structure (I)

[0174]

[0175]

[0176]

[0177]

[0178]

[0179]

[0180]

[0181]

[0182]

[0183]

[0184]

[0185]

[0186]

[0187]

[0188]

[0189]

[0190]

[0191]

[0192]

[0193]

[0194]

[0195]

[0196] It should be understood that, in this description, the combination of substituents and / or variables in the formulas is only permitted if such effects produce stable compounds.

[0197] In another embodiment, the various compounds of this disclosure, existing in free base or acid form, can be converted into their pharmaceutically acceptable salts by treatment with a suitable inorganic or organic base or acid using methods known to those skilled in the art. Salts of the compounds of this disclosure can be converted back to their free base or acid forms using standard techniques.

[0198] The following provides methods for producing the compounds described herein. Starting components can typically be obtained from sources such as Sigma-Aldrich, Lancaster Synthesis, Inc., Maybridge, Matrix Scientific, TCI, and Fluorochem USA, or synthesized from sources known to those skilled in the art (see, for example, Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 5th Edition, (Wiley, December 2000)) or prepared as described herein.

[0199] The following general reaction scheme illustrates an exemplary method for preparing a compound with structure (I):

[0200]

[0201] Or an acceptable salt, stereoisomer, or prodrug of the drug, wherein A, X, Y, R 1 R 2 R 3 and R 4 Each as defined in this article.

[0202] General Reaction Scheme 1

[0203] The following is a general reaction scheme (where X) 1 It is a halogen, and X, R 1 R 2 (A and A have the meanings described herein) An example of a method for preparing amine intermediate B is shown:

[0204]

[0205] As shown in general reaction scheme 1, pyrazolopyrimidine is functionalized with an alkyl or arylboronic acid or a suitable electrophile in the presence of a base to give intermediate A, which can then undergo palladium-catalyzed arylation (followed by a reduction step if necessary) to form amine intermediate B.

[0206] General Reaction Scheme 2

[0207] The following general reaction scheme illustrates an example of a method for preparing carbamate intermediate C:

[0208]

[0209] As shown in general reaction scheme 2, intermediate C can be reacted in the presence of a base via phenyl chloroformate and the indicated heteroarylamine (R). 3 It is prepared by the reaction of amine-substituted analogues. General reaction scheme 2 describes that R 4 It is the preparation of compounds containing H; however, R in which 4 Compounds that are not H can be prepared by setting R after intermediate I using a similar method. 4 It can be prepared by means of a suitable substituted heteroarylamine, or by means of a suitable substituted heteroarylamine.

[0210] General Reaction Scheme 3

[0211] The following general reaction scheme illustrates examples of methods for preparing compounds of structure (I):

[0212]

[0213] Intermediate B and intermediate C are treated with a base (e.g., trimethylamine) in THF to give the compound with structure (I).

[0214] General Reaction Scheme 4

[0215] The following general reaction scheme illustrates examples of methods for preparing compounds of structure (I):

[0216]

[0217] Intermediate B is reacted with phenyl chloroformate under appropriate conditions to give intermediate C. Then, intermediate C is coupled with an amine in THF using a suitable base (e.g., trimethylamine, DIPEA, DMAP, etc.) to give the compound of structure (I).

[0218] Any of the above reaction schemes can be modified at any step to add and / or modify substituents, or the order of steps can be appropriately changed at any stage of the entire synthesis of the desired compound. For example, those skilled in the art will readily understand that the carbamate analogue of intermediate B can be alternatively prepared and reacted with R. 3 The amine analogues were reacted to prepare compounds of structure (I).

[0219] Those skilled in the art will also understand that, in the methods for preparing the compounds described herein, the functional groups of intermediate compounds may require protection by suitable protecting groups. Such functional groups include, but are not limited to, hydroxyl, amino, mercapto, and carboxylic acids. Suitable protecting groups for hydroxyl groups include trialkylsilyl or diarylalkylsilyl (e.g., tert-butyldimethylsilyl, tert-butyldiphenylsilyl, or trimethylsilyl), tetrahydropyranyl, benzyl, etc. Suitable protecting groups for amino, amidine, and guanidine groups include tert-butoxycarbonyl, benzyloxycarbonyl, etc. Suitable protecting groups for mercapto groups include -C(O)-R" (where R" is alkyl, aryl, or arylalkyl), p-methoxybenzyl, triphenylmethyl, etc. Suitable protecting groups for carboxylic acids include alkyl esters, aryl esters, or arylalkyl esters. Protecting groups may optionally be added or removed according to standard techniques known to those skilled in the art and as described herein. The use of protecting groups is described in detail in Green, TW, and PGMUTZ, Protective Groups in Organic Synthesis (1999), 3rd edition, Wiley. As those skilled in the art will understand, protecting groups can also be polymeric resins, such as Wang resins, Rink resins, or 2-chlorotriphenylmethyl-chloride resins.

[0220] Those skilled in the art will also understand that although such protected derivatives of the compounds of this disclosure may not be pharmacologically active, they can similarly be administered to mammals and subsequently metabolized in vivo to form pharmacologically active compounds of this disclosure. Therefore, such derivatives can be described as “prodrugs.” Prodrugs of the compounds of this disclosure are included within the scope of embodiments of this disclosure.

[0221] Pharmaceutical Composition

[0222] Other embodiments involve pharmaceutical compositions. The pharmaceutical composition comprises any one (or more) of the aforementioned compounds and a pharmaceutically acceptable carrier. In some embodiments, the pharmaceutical composition is formulated for oral administration. In other embodiments, the pharmaceutical composition is formulated for injection. In numerous embodiments, the pharmaceutical composition comprises the compounds disclosed herein and additional therapeutic agents (e.g., anticancer agents). Non-limiting examples of such therapeutic agents are described below.

[0223] Suitable routes of administration include, but are not limited to, oral, intravenous, rectal, aerosol, parenteral, ocular, pulmonary, mucosal, transdermal, vaginal, ocular, nasal, and local administration. Additionally, by way of example only, parenteral delivery includes intramuscular, subcutaneous, intravenous, intramedullary injection, as well as intrathecal, direct intracardiac, intraperitoneal, intralymphatic, and intranasal injection.

[0224] In some embodiments, the compounds described herein are administered locally rather than systemically, for example, by direct injection into an organ, typically in the form of a reservoir formulation or a sustained-release formulation. In specific embodiments, long-acting formulations are administered via implantation (e.g., subcutaneously or intramuscularly) or by intramuscular injection. Furthermore, in other embodiments, the compounds are delivered in a targeted drug delivery system, such as in liposomes coated with organ-specific antibodies. In such embodiments, the liposomes target the organ and are selectively absorbed by the organ. In other embodiments, the compounds described herein are provided in the form of a rapid-release formulation, a prolonged-release formulation, or an intermediate-release formulation. In other embodiments, the compounds described herein are administered locally.

[0225] In a treatment method according to an embodiment of the invention, an effective amount of a compound of at least one structure (I) is administered to an individual who has or has been diagnosed with such a disease, condition, or medical condition. The effective amount or dose may be determined by methods such as modeling, dose-escalation studies, or clinical trials, for example, the mode or route of administration or drug delivery, the pharmacokinetics of the agent, the severity and progression of the disease, condition, or medical condition, the individual's prior or ongoing treatments, the individual's health status and response to the drug, and the judgment of the treating physician.

[0226] The compounds according to this disclosure are effective over a wide dosage range. For example, in the treatment of adults, dosages of 10 to 5000 mg / day, 100 to 5000 mg / day, 1000 to 4000 mg / day, and 1000 to 3000 mg / day are examples of dosages used in some embodiments. The precise dosage depends on the route of administration, the form of the compound administered, the subject being treated, the weight of the subject being treated, and the preference and experience of the attending physician.

[0227] In some embodiments, the compounds of this disclosure are administered in a single dose. Typically, this administration is by injection, such as intravenous injection, to rapidly introduce the drug. However, other routes may be used as appropriate. A single dose of the compounds of this disclosure may also be used to treat acute conditions.

[0228] In some embodiments, the compounds of this disclosure are administered in multiple doses. In some embodiments, the administration is about once, twice, three times, four times, five times, six times, or more than six times daily. In other embodiments, the administration is about once a month, once every two weeks, once a week, or once every other day. In another embodiment, the compounds of this disclosure are administered together with another agent (e.g., an anticancer agent) about once a day to about six times a day. In another embodiment, the administration of the compounds and agents of this disclosure is sustained for less than about 7 days. In another embodiment, the administration is sustained for more than about 6 days, 10 days, 14 days, 28 days, two months, six months, or one year. In some cases, continuous administration is achieved and maintained, if necessary.

[0229] The compounds of this disclosure can be administered for as long as necessary. In some embodiments, the compounds of this disclosure are administered for more than 1, 2, 3, 4, 5, 6, 7, 14, or 28 days. In some embodiments, the compounds of this disclosure are administered for less than 28, 14, 7, 6, 5, 4, 3, 2, or 1 day. In some embodiments, the compounds of this disclosure are administered on a continuous basis for a long period, for example, to treat chronic effects.

[0230] In some embodiments, the compounds of this disclosure are administered in separate dosage forms. It is known in the art that individualization of dosing regimens is essential for optimal treatment due to inter-individual variability in the pharmacokinetics of compounds.

[0231] In some embodiments, the compounds described herein are formulated into pharmaceutical compositions. In specific embodiments, the pharmaceutical compositions are formulated in a conventional manner using one or more physiologically acceptable carriers, said carriers comprising excipients and adjuvants that facilitate the processing of the disclosed compounds into pharmaceutically acceptable formulations. A suitable formulation depends on the chosen route of administration. Any pharmaceutically acceptable technology, carrier, and excipient is suitable for formulating the pharmaceutical compositions described herein: Remington: The Science and Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack Publishing Company, 1995); Hoover, John E., Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania 1975; Liberman, H.A. and Lachman, L., eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, NY, 1980; and Pharmaceutical Dosage Forms and Drug Delivery Systems, Seventh Edition (Lippincott Williams & Wilkins 1999).

[0232] This article provides pharmaceutical compositions comprising a compound containing one or more structures (I) and a pharmaceutically acceptable carrier.

[0233] This document provides pharmaceutical compositions comprising one or more compounds selected from structure (I) and pharmaceutically acceptable diluents, excipients, and carriers. In some embodiments, the compounds are administered as a pharmaceutical composition, wherein one or more compounds selected from structure (I) are mixed with other active ingredients, as in combination therapy. This document covers all combinations of active substances described in the Combination Therapy section below, as well as all combinations of active substances described throughout this disclosure. In specific embodiments, the pharmaceutical composition comprises one or more compounds of structure (I).

[0234] In one embodiment, the pharmaceutical composition of the compound of structure (I) is a modulator of the NLRP3 inflammasome.

[0235] In a specific implementation, the pharmaceutical composition of the compound of structure (I) inhibits NEK7 when administered to a patient or biological sample.

[0236] As used herein, a pharmaceutical composition refers to a mixture of one or more compounds selected from structure (I) with other chemical components, such as carriers, stabilizers, diluents, dispersants, suspending agents, thickeners, and / or excipients. In some embodiments, the pharmaceutical composition facilitates the administration of the compound to an organism. In some embodiments, a therapeutically effective amount of one or more compounds selected from structure (I) provided herein is administered in the pharmaceutical composition to a mammal suffering from a disease, condition, or medical condition to be treated. In a specific embodiment, the mammal is a human. In some embodiments, the therapeutically effective amount varies depending on the severity of the disease, the individual's age and relative health status, the efficacy of the compound used, and other factors. The compounds described herein are used alone or in combination with one or more therapeutic agents as components of mixtures.

[0237] In one embodiment, one or more compounds selected from structure (I) are formulated into an aqueous solution. In a specific embodiment, by way of example only, the aqueous solution is selected from physiologically compatible buffers, such as Hank's solution, Ringer's solution, or physiological saline buffer. In other embodiments, one or more compounds selected from structure (I) are formulated for transmucosal administration. In a specific embodiment, the transmucosal formulation includes an exfoliant suitable for the barrier to be penetrated. In other embodiments, wherein the compounds described herein are formulated for other parenteral injections, suitable formulations include aqueous or non-aqueous solutions. In a specific embodiment, such solutions include physiologically compatible buffers and / or excipients.

[0238] In another embodiment, the compounds described herein are formulated for oral administration. The compounds described herein are formulated by combining the active compound with, for example, a pharmaceutically acceptable carrier or excipient. In various embodiments, the compounds described herein are formulated into oral dosage forms, including (by way of example only) tablets, powders, pills, sugar-coated pills, capsules, liquids, gels, syrups, elixirs, liquids, suspensions, etc.

[0239] In some embodiments, pharmaceutical formulations for oral use are obtained by mixing one or more solid excipients with one or more compounds described herein, optionally grinding the resulting mixture, and processing the granular mixture after adding suitable excipients (if desired) to obtain tablets or sugar-coated pellet cores. Suitable excipients are particularly fillers, such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose formulations, such as corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth gum, methylcellulose, microcrystalline cellulose, hydroxypropyl methylcellulose, sodium carboxymethylcellulose; or other substances, such as polyvinylpyrrolidone (PVP or povidone) or calcium phosphate. In specific embodiments, a disintegrant is optionally added. By way of example only, disintegrants include cross-linked croscarmellose sodium carboxymethylcellulose, polyvinylpyrrolidone, agar, or alginate or its salts such as sodium alginate.

[0240] In one embodiment, the dosage form, such as sugar-coated cores and tablets, is provided with one or more suitable coatings. In a specific embodiment, a concentrated sugar solution is used to coat the dosage form. The sugar solution optionally contains additional components, such as, by way of example only, gum arabic, talc, polyvinylpyrrolidone, carbomer gel, polyethylene glycol and / or titanium dioxide, lacquer solution, and suitable organic solvents or solvent mixtures. For identification purposes, dyes and / or pigments are also optionally added to the coating. Additionally, dyes and / or pigments are optionally used to characterize different combinations of active compound dosages.

[0241] In some embodiments, a therapeutically effective amount of at least one of the compounds described herein is formulated into other oral dosage forms. Oral dosage forms include push-fit capsules made of gelatin, and soft-sealable capsules made of gelatin and a plasticizer (e.g., glycerin or sorbitol). In specific embodiments, the push-fit capsule contains the active ingredient mixed with one or more fillers. Fillers include (by way of example only) lactose, binders (e.g., starch), and / or lubricants (e.g., talc or magnesium stearate), and optionally, stabilizers. In other embodiments, the soft capsule contains one or more active compounds dissolved or suspended in a suitable liquid. Suitable liquids include, by way of example only, one or more fatty oils, liquid paraffin, or liquid polyethylene glycol. Additionally, a stabilizer is optionally added.

[0242] In other embodiments, the compounds described herein are formulated for parenteral injection, including formulations suitable for rapid concentration or continuous infusion. In specific embodiments, the formulation for injection is provided in a unit dosage form (e.g., in an ampoule) or in a multi-dose container. Optionally, a preservative is added to the injectable formulation. In other embodiments, the pharmaceutical composition is formulated for parenteral injection as a sterile suspension, solution, or emulsion in an oily or aqueous medium. The parenteral injection formulation optionally contains a formulation, such as a suspending agent, stabilizer, and / or dispersant. In specific embodiments, the pharmaceutical formulation for parenteral administration comprises an aqueous solution of the active compound in a water-soluble form. In further embodiments, a suspension of one or more compounds selected from structure (I) is prepared as a suitable oily injectable suspension. Suitable lipophilic solvents or media for use in the pharmaceutical compositions described herein include (by way of example only) fatty oils (e.g., sesame oil) or synthetic fatty acid esters (e.g., ethyl oleate or triglycerides) or liposomes. In some specific embodiments, the aqueous injection suspension contains a substance that increases the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension contains a suitable stabilizer or a reagent that increases the solubility of the compound to allow for the preparation of a highly concentrated solution. Alternatively, in other embodiments, the active ingredient is in powder form and is prepared for use with a suitable medium (e.g., sterile, pyrogen-free water) prior to use.

[0243] The pharmaceutical composition comprises at least one pharmaceutically acceptable carrier, diluent, or excipient, and one or more compounds selected from the structure (I) described herein as the active ingredient. The active ingredient is in the form of a free acid or free base, or in the form of a pharmaceutically acceptable salt. Furthermore, the methods and pharmaceutical compositions described herein include the use of N-oxides, crystalline forms (also called polymorphs), and active metabolites of these compounds having the same type of activity. All tautomers of the compounds described herein are included within the scope of the compounds presented herein. In addition, the compounds described herein include unsolvable forms as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, etc. The solvated forms of the compounds presented herein are also considered to be disclosed herein. Furthermore, the pharmaceutical composition optionally includes other drugs or pharmaceutical agents, carriers, adjuvants, such as preservatives, stabilizers, wetting agents or emulsifiers, solubilizers, salts for adjusting osmotic pressure, buffers, and / or other substances of therapeutic value.

[0244] Methods for preparing compositions comprising the compounds described herein include formulating the compounds with one or more inert, pharmaceutically acceptable excipients or carriers to form a solid, semi-solid, or liquid. Solid compositions include, but are not limited to, powders, tablets, dispersible granules, capsules, pouches, and suppositories. Liquid compositions comprise solutions in which the compounds are dissolved, emulsions containing the compounds, or solutions containing liposomes, micelles, or nanoparticles containing the compounds disclosed herein. Semi-solid compositions include, but are not limited to, gels, suspensions, and creams. The pharmaceutical compositions described herein are in the form of liquid solutions or suspensions, solid forms suitable for dissolving or suspending in a liquid prior to use, or as emulsions. These compositions also optionally contain small amounts of non-toxic excipients, such as wetting agents or emulsifiers, pH buffers, etc.

[0245] In some embodiments, pharmaceutical compositions comprising one or more compounds selected from structure (I) are exemplary in liquid form, wherein the agent is present in a solution, a suspension, or both. Typically, when the composition is administered as a suspension, a first portion of the agent is present in solution, and a second portion of the agent is present as particles suspended in a liquid matrix. In some embodiments, the liquid composition comprises a gel formulation. In other embodiments, the liquid composition is aqueous.

[0246] In some embodiments, the aqueous suspension contains one or more polymers as suspending agents. Polymers include water-soluble polymers, such as cellulose polymers, such as hydroxypropyl methylcellulose; and water-insoluble polymers, such as cross-linked carboxyl-containing polymers. Certain pharmaceutical compositions described herein contain mucosal adhesion polymers selected from, for example, carboxymethyl cellulose, carbomer (acrylic polymer), poly(methyl methacrylate), polyacrylamide, polycarbofil, acrylate / butyl acrylate copolymer, sodium alginate, and dextran.

[0247] The pharmaceutical composition may also optionally include a solubilizer to improve the solubility of one or more compounds selected from structure (I). The term "sorcerer" generally includes an agent that results in the formation of a micelle solution or a real solution of the pharmaceutical agent. Certain acceptable nonionic surfactants, such as polysorbate 80, may be used as solubilizers, as well as ophthalmologically acceptable ethylene glycols, polyethylene glycols such as polyethylene glycol 400, and ethylene glycol ethers.

[0248] In addition, the pharmaceutical composition optionally includes one or more pH adjusters or buffers, including acids such as acetic acid, boric acid, citric acid, lactic acid, phosphoric acid, and hydrochloric acid; bases such as sodium hydroxide, sodium phosphate, sodium borate, sodium citrate, sodium acetate, sodium lactate, and tris(hydroxymethyl)aminomethane; and buffers such as citrate / dextran, sodium bicarbonate, and ammonium chloride. These acids, bases, and buffers are included in amounts necessary to maintain the pH of the composition within an acceptable range.

[0249] Optionally, the composition further includes one or more salts in an amount such that the osmotic pressure of the composition reaches an acceptable range. Such salts include salts having sodium, potassium, or ammonium cations and chloride, citrate, ascorbate, borate, phosphate, bicarbonate, sulfate, thiosulfate, or bisulfite anions; suitable salts include sodium chloride, potassium chloride, sodium thiosulfate, sodium bisulfite, and ammonium sulfate.

[0250] Other pharmaceutical compositions optionally include one or more preservatives to inhibit microbial activity. Suitable preservatives include mercury-containing substances, such as phenylmercuric nitrate and thimerosal; stable chlorine dioxide; and quaternary ammonium compounds, such as benzalkonium chloride, hexadecyltrimethylammonium bromide, and hexadecylpyridinium chloride.

[0251] The composition may contain one or more surfactants to enhance physical stability or for other purposes. Suitable nonionic surfactants include polyoxyethylene fatty acid glycerides and vegetable oils, such as polyoxyethylene (60) hydrogenated castor oil; and polyoxyethylene alkyl ethers and alkylphenyl ethers, such as octylphenyl polyol 10 and octylphenyl polyol 40.

[0252] The composition may contain one or more antioxidants to enhance chemical stability when needed. Suitable antioxidants include, by way of example only, ascorbic acid and sodium metabisulfite.

[0253] In some embodiments, the aqueous suspension composition is encapsulated in a single-dose, non-resealable container. Alternatively, a multi-dose, resealable container is used, in which case a preservative is typically included in the composition.

[0254] In alternative embodiments, other delivery systems for hydrophobic pharmaceutical compounds are used. Liposomes and emulsions are examples of delivery media or carriers that can be used herein. In some embodiments, organic solvents, such as N-methylpyrrolidone, are also used. In other embodiments, sustained-release systems, such as a semi-permeable matrix of a solid hydrophobic polymer containing the therapeutic agent, are used to deliver the compounds described herein. Various sustained-release materials are useful herein. In some embodiments, sustained-release capsules release the compound for weeks up to more than 100 days. Additional protein stabilization strategies are employed depending on the chemical properties and biological stability of the therapeutic agent.

[0255] In some embodiments, the formulations described herein comprise one or more antioxidants, metal chelators, thiol-containing compounds, and / or other general stabilizers. Examples of such stabilizers include, but are not limited to: (a) about 0.5% to about 2% w / v glycerol, (b) about 0.1% to about 1% w / v methionine, (c) about 0.1% to about 2% w / v monothioglycerol, (d) about 1 mM to about 10 mM EDTA, (e) about 0.01% to about 2% w / v ascorbic acid, (f) 0.003% to about 0.02% w / v polysorbate 80, (g) 0.001% to about 0.05% w / v polysorbate 20, (h) arginine, (i) heparin, (j) dextran sulfate, (k) cyclodextrin, (l) pentosan polysulfates and other heparin-like substances, (m) divalent cations, such as magnesium and zinc; or (n) combinations thereof.

[0256] In some embodiments, the concentration of one or more compounds selected from structure (I) provided in the pharmaceutical composition of this disclosure is greater than 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 19.75%, 19.50%, 19.25%, 19%, 18.75%, 18.50%, 18.25%, 18%, 17.75%, 17.50%, 17.25%, 17%, 16.75%, 16.50%, 16.25%, 16%, 15.7%. 5%, 15.50%, 15.25% 15%, 14.75%, 14.50%, 14.25% 14%, 13.75%, 13.50%, 13.25% 13%, 12.75%, 12.50%, 12.25% 12%, 11.75%, 11.50%, 11.25% 11%, 10.75%, 10.50%, 10.25% 10%, 9.75%, 9.50%, 9.25% 9%, 8.75%, 8.50%, 8.25% 8%, 7.7 5%, 7.50%, 7.25% 7%, 6.75%, 6.50%, 6.25% 6%, 5.75%, 5.50%, 5.25% 5%, 4.75%, 4.50%, 4.25% 4%, 3.75%, 3.50%, 3.25% 3%, 2.75%, 2.50%, 2.25% 2%, 1.75%, 1.50% 125%, 1%, 0.5%, 0.4%, 0.3%, 0.2%, 0.1%, 0.09%, 0.08%, 0.07% 0.06%, 0.05%, 0.04%, 0.03%, 0.02%, 0.01%, 0.009%, 0.008%, 0.007%, 0.006%, 0.005%, 0.004%, 0.003%, 0.002%, 0.001%, 0.0009%, 0.0008%, 0.0007%, 0.0006%, 0.0005%, 0.0004%, 0.0003%, 0.0002%, or 0.0001% w / w, w / v, or v / v.

[0257] In some embodiments, the concentration of one or more compounds selected from structure (I) provided in the pharmaceutical composition of this disclosure is about 0.0001% to about 50%, about 0.001% to about 40%, about 0.01% to about 30%, about 0.02% to about 29%, about 0.03% to about 28%, about 0.04% to about 27%, about 0.05% to about 26%, about 0.06% to about 25%, or about 0. 0.07% to about 24%, about 0.08% to about 23%, about 0.09% to about 22%, about 0.1% to about 21%, about 0.2% to about 20%, about 0.3% to about 19%, about 0.4% to about 18%, about 0.5% to about 17%, about 0.6% to about 16%, about 0.7% to about 15%, about 0.8% to about 14%, about 0.9% to about 12%, about 1% to about 10% w / w, w / v or v / v.

[0258] In some embodiments, the amount of one or more compounds selected from structure (I) provided in the pharmaceutical composition of this disclosure is equal to or less than 10g, 9.5g, 9.0g, 8.5g, 8.0g, 7.5g, 7.0g, 6.5g, 6.0g, 5.5g, 5.0g, 4.5g, 4.0g, 3.5g, 3.0g, 2.5g, 2.0g, 1.5g, 1.0g, 0.95g, 0.9g, 0.85g, 0.8g, 0.75g, 0.7g, 0.65g, 0.6g, 0.55g, 0.5g, 0.45g, 0.4g, or 0.35g. , 0.3g, 0.25g, 0.2g, 0.15g, 0.1g, 0.09g, 0.08g, 0.07g, 0.06g, 0.05g, 0.04g, 0.03g, 0.02g, 0.01g, 0.009g, 0.008g, 0.007g, 0.006g , 0.005g, 0.004g, 0.003g, 0.002g, 0.001g, 0.0009g, 0.0008g, 0.0007g, 0.0006g, 0.0005g, 0.0004g, 0.0003g, 0.0002g, or 0.0001g.

[0259] In some embodiments, the amount of one or more compounds selected from structure (I) provided in the pharmaceutical composition of this disclosure is 0.0001-10g, 0.0005-9g, 0.001-8g, 0.005-7g, 0.01-6g, 0.05-5g, 0.1-4g, 0.5-4g, or 1-3g.

[0260] Packaging materials used to package the pharmaceutical compositions described herein include those found, for example, in U.S. Patents 5,323,907, 5,052,558, and 5,033,252. Examples of pharmaceutical packaging materials include (but are not limited to) blister packs, bottles, tubes, inhalers, pumps, bags, vials, containers, syringes, bottles, and any packaging material suitable for the selected formulation and the intended mode of administration and treatment. For example, a container may comprise one or more of the compounds described herein, optionally in the form of a composition or in combination with another pharmaceutical agent disclosed herein. The container may optionally have a sterile inlet (e.g., the container is an intravenous solution bag or a vial with a stopper that can be punctured by a hypodermic needle). Such kits may optionally contain compounds with an identification description or label or instructions relating to their use in the methods described herein.

[0261] For example, a kit typically includes one or more additional containers, each containing one or more materials (e.g., reagents, optionally in concentrated form, and / or devices) desired from a commercial and user perspective for the use of the compounds described herein. Non-limiting examples of such materials include, but are not limited to, buffers, diluents, filters, needles, syringes; carriers, packages, containers, vials, and / or tube labels listing the contents and / or instructions for use; and instruction manuals with instructions for use. A set of instructions will also typically be included. Labels may optionally be located on or associated with the container. For example, a label is located on the container when the letters, numbers, or other characters forming the label are attached, molded, or etched onto the container itself; and a label is associated with the container, such as as an instruction manual, when the label is present within a container or carrier that also holds the container. Furthermore, the label is used to indicate that the contents will be used for a specific therapeutic application. Additionally, the label indicates instructions for use of the contents, such as those described herein. In some embodiments, the pharmaceutical composition is contained in a package or dispenser device comprising one or more unit dosage forms containing the compounds provided herein. The package may contain, for example, metal or plastic foil, such as blister packs. Alternatively, the package or dispenser device may be accompanied by instructions for use. Alternatively, the packaging or dispenser may also include a notification associated with the container, provided by the government agency regulating the manufacture, use, or sale of the drug, reflecting the form of the drug approved by that agency for human or veterinary administration. Such notification may be, for example, a label for a prescription drug approved by the U.S. Food and Drug Administration, or an approved product label. In some embodiments, a composition containing the compounds provided herein formulated in a compatible drug carrier is prepared, placed in a suitable container, and labeled for the treatment of a specified condition.

[0262] method

[0263] Embodiments of this disclosure can be used as modulators of the NLRP3 inflammasome by inhibiting NEK7 in the host species. Therefore, compounds of structure (I) can also be used to treat conditions mediated by effector signaling molecules such as IL-1β and IL-18.

[0264] The host or patient can belong to any mammal species, such as primates, especially humans; rodents, including mice, rats, and hamsters; rabbits; horses, cattle, dogs, cats, etc. Animal models are of great interest for experimental research, providing models for treating human diseases.

[0265] In one embodiment, this disclosure can be used as an inhibitor of the NLRP3 inflammasome activation mechanism. Therefore, compounds of structure (I) can also be used to treat conditions caused by activation in the host species.

[0266] In another embodiment, the compound of structure (I) can be used as an inhibitor of NLRP3 (protein)-NEK7 (protein) interaction. Therefore, the compound can also be used to treat conditions caused by NLRP3-NEK7 association in a host species.

[0267] In some embodiments, the compound of structure (I) can be used to treat human conditions mediated by effectors selected from IL-1β, IL-18 and caspase-1.

[0268] Embodiments of this disclosure also relate to the use of compounds according to structure (I) and / or physiologically acceptable salts thereof for the preventive or therapeutic treatment and / or monitoring of diseases caused, mediated, and / or regulated by NLRP3 inflammasome activity. Furthermore, embodiments of the invention relate to the use of compounds according to structure (I) and / or physiologically acceptable salts thereof for the preparation of medicaments for the preventive or therapeutic treatment and / or monitoring of diseases caused, mediated, and / or regulated by NLRP3 inflammasome activity. In some embodiments, the invention provides the use of compounds according to structure I or physiologically acceptable salts thereof for the preparation of medicaments for the preventive or therapeutic treatment of NLRP3-mediated conditions.

[0269] In another embodiment, this disclosure relates to a method of treating inflammatory diseases or conditions mediated by the NLRP3 inflammasome by administering a therapeutically effective amount of the compound of structure (I) to a patient in need.

[0270] In some implementations, diseases that can be treated with compounds of structure (I) include type II diabetes, atherosclerosis, Alzheimer's disease, aging, fatty liver, metabolic syndrome, asthma, psoriasis, obesity, acute and chronic tissue damage caused by infection, gout, arthritis, enteritis, hepatitis, peritonitis, silicosis, UV-induced sunburn, contact hypersensitivity, sepsis, cancer, neurodegenerative diseases, multiple sclerosis, and Muckle-Wells syndrome.

[0271] In some other embodiments, in a method of treating a condition or disease with the compound of structure (I), said condition or disease is selected from autoimmune diseases, inflammatory conditions, cardiovascular diseases, neurodegenerative diseases, bacterial and viral infections, allergies, asthma, pancreatitis, multiple organ failure, kidney disease, platelet aggregation, cancer, transplantation, sperm motility, aplasia, transplant rejection, lung injury, respiratory diseases, ischemic conditions, and cancer.

[0272] In some implementations, the NEK7-related conditions that can be treated with compounds of structure (I) are selected from rheumatoid arthritis, psoriatic arthritis, osteoarthritis, systemic lupus erythematosus, lupus nephritis, ankylosing spondylitis, osteoporosis, systemic sclerosis, multiple sclerosis, psoriasis, type I diabetes, type II diabetes, inflammatory bowel disease (Crohn's disease and ulcerative colitis), hyperimmunoglobulinemia D and periodic fever syndrome, cryptothermal protein-associated periodic syndrome, Schnitzler syndrome, systemic juvenile idiopathic arthritis, adult-onset Still's disease, gout, pseudogout, SAPHO syndrome, Kassman disease, sepsis, stroke, atherosclerosis, celiac disease, DIRA (IL-1 receptor antagonist deficiency), Alzheimer's disease, Parkinson's disease, and cancer.

[0273] This article also includes treatment methods in which a compound of at least one structure (I) is administered in combination with an anti-inflammatory agent or therapeutic agent. Anti-inflammatory agents include, but are not limited to, NSAIDs, non-specific and COX-2-specific cyclooxygenase inhibitors, gold compounds, corticosteroids, methotrexate, tumor necrosis factor (TNF) antagonists, immunosuppressants, and methotrexate. Examples of NSAIDs include, but are not limited to, ibuprofen, flurbiprofen, naproxen and naproxen sodium, diclofenac, combinations of diclofenac sodium and misoprostol, sulindac, oxapazine, diflunisal, piroxicam, indomethacin, etodoxacin, fenofosine calcium, ketoprofen, nabumetone sodium, sulfasalazine, tometidine sodium, and hydroxychloroquine.

[0274] Examples of NSAIDs also include COX-2 specific inhibitors, such as celecoxib, vardecoxib, romecoxib, and / or etoricoxib.

[0275] In some implementations, the anti-inflammatory agent is a salicylate. Salicylates include, but are not limited to, acetylsalicylic acid or aspirin, sodium salicylate, choline, and magnesium salicylate.

[0276] Anti-inflammatory agents can also be corticosteroids. For example, corticosteroids can be cortisone, dexamethasone, methylprednisolone, prednisolone, prednisolone sodium phosphate, or prednisone.

[0277] In another embodiment, the anti-inflammatory agent is a gold compound, such as sodium gold thiomalate or auronoxine.

[0278] This disclosure also includes embodiments in which the anti-inflammatory agent is a metabolic inhibitor, such as a dihydrofolate reductase inhibitor, such as methotrexate, or a dihydroorotate dehydrogenase inhibitor, such as leflunomide.

[0279] Therapeutic agents may also include agents for pain and inflammation, such as histamine and histamine antagonists, bradykinin and bradykinin antagonists, serotonin (5-hydroxytryptamine), lipid substances produced by biotransformation of products of selective hydrolysis of membrane phospholipids, arachidic acid, prostaglandins, thromboxanes, leukotrienes, aspirin, nonsteroidal anti-inflammatory agents, analgesics and antipyretics, agents that inhibit the synthesis of prostaglandins and thromboxanes, selective inhibitors of inducible cyclooxygenase, selective inhibitors of inducible cyclooxygenase-2, endocrine substances, paracrine hormones, somatostatin, gastrin, cytokines that mediate interactions in humoral and cellular immune responses, lipid-derived endocrine substances, arachidic acid, β-adrenergic agonists, ipratropium, glucocorticoids, methylxanthine, sodium channel blockers, opioid receptor agonists, calcium channel blockers, membrane stabilizers, and leukotriene inhibitors.

[0280] Other embodiments of this disclosure involve a combination of at least one anti-inflammatory compound that is an anti-monoclonal antibody (e.g., eculizumab or pecrolimus) and a TNF antagonist (e.g., etanercept or infliximab, which is an anti-TNFα monoclonal antibody).

[0281] Therapeutic agents used in combination with compounds of structure (I) may also include small molecule compounds that inhibit the activation of the NLRP3 inflammasome, such as MCC950, sulforaphane, isoliquiritigenin, β-hydroxybutyrate, flufenamic acid, mefenamic acid, 3,4-methylenedioxy-β-nitrostyrene (MNS), and ternolactone.

[0282] Other embodiments of this disclosure also relate to a combination of at least one active agent being an immunosuppressive compound, such as an immunosuppressive compound selected from methotrexate, leflunomide, cyclosporine, tacrolimus, azathioprine, and mycophenolate mofetil.

[0283] Compounds of the disclosed structure (I) can be administered in combination with other known therapeutic agents, including anticancer agents. As used herein, the term "anticancer agent" refers to any agent administered to a patient with cancer for the treatment of cancer.

[0284] In some implementation schemes, anticancer agents fall into the following categories:

[0285] Alkylating agents: such as hexamethylpyrimidine, bendamustine, busulfan, carmustine, chlorambucil, nitrogen mustard, cyclophosphamide, dacarbazine, ifosfamide, inprofen, toxalate, lomustine, melphalan, dibromomannitol, dibromoeusine, nimustine, ramustine, temozolomide, thiotepa, treoxazol, dichloromethyldiethylamine, carboquinone; apatziquinone, formustine, glutamine, ifosfamide, piperobromidine, trefophosphamide, uramustine, TH-3024, VAL-0834;

[0286] Platinum compounds: such as carboplatin, cisplatin, epplatin, miplatin hydrate, oxaliplatin, lobaplatin, nedaplatin, pyrazoplatin, saxaplatin; lobaplatin, nedaplatin, pyrazoplatin, saxaplatin;

[0287] DNA altering agents: such as amrubicin, bifenthrin, decitabine, mitoxantrone, procarbazine, trabectedin, clofarabine; acridine, bromocriptine, pixantrone, lamosine 1,3;

[0288] Topoisomerase inhibitors: such as etoposide, irinotecan, razorizine, sobuzosen, teniposide, toponotecan; aminonaphthylfenoxate, belotecone, eletidine, voriracin;

[0289] Microtubule modulators: such as cabazitaxel, docetaxel, elebulin, ixaprilone, paclitaxel, vincristine, vinorelbine, vindesine, vinflunine; forbrene, tistanoxel;

[0290] Antimetabolites: such as asparaginase 3, azacitidine, leucovorin calcium, capecitabine, cladribine, cytarabine, enoxabin, fluorouridine, fludarabine, fluorouracil, gemcitabine, mercaptopurine, methotrexate, nelabine, pemetrexed, pralatrexate, azathioprine, thioguanine, carmoflu; deoxyfluorouridine, esilaribine, raltitrexed, sapattabine, tegafur 2,3, trimethoprim;

[0291] Anticancer antibiotics: such as bleomycin, actinomycin D, doxorubicin, epirubicin, idarubicin, levamisole, mitofocin, mitomycin C, romidesin, streptozotocin, pentorubicin, fentoxin, fosfostatin, zolomatin, daunomycin, procainoxam; arubicin, pepromycin, pirarubicin;

[0292] Hormones / antagonists such as abalix, abiraterone, bicalutamide, busherin, carotestosterone, chlorestradiol, degarelix, dexamethasone, estradiol, flucloprone, flumethasone, flutamide, fulvestrant, goserelin, histidine, leuprorelin, medroxyprogesterone acetate, mitotane, naffarelin, nandrolone, nilumethoxazole, octreotide, prednisolone, raloxifene, tamoxifen, thyroid-stimulating hormone alpha, toremifene, triplostertan, triptorelin, diethylstilbestrol; acobifen, danazol, diloxacin, cyclothionol, oterolone, enzalutamide 1,3;

[0293] Aromatase inhibitors: such as ammoniaglutide, anastrozole, exemestane, fadroxil, letrozole, testosterone; formestan;

[0294] Small molecule kinase inhibitors: such as crizotinib, dasatinib, erlotinib, imatinib, lapatinib, nilotinib, pazopanib, regorafenib, ruxotinib, sorafenib, sunitinib, vandetanib, vemurafenib, besutinib, gefitinib, axitinib; afatinib, alisertib, dabrafenib, dacomitinib, dinaciclib, dovirtinib, enzastaurin, nintedanib, lenvatinib, linifanib, linstatinib (… Linsitinib, masitinib, midotutolin, motixab, neratinib, orantinib, perifol, panatinib, radotinib, rigoseltib, tipifamib, tivantinib, tivozanib, trametinib, pimasertib, brinibuturate, sildenafil.

[0295] In some embodiments, the drugs administered in combination with the compounds described herein include any suitable drug that is effectively delivered by inhalation, such as analgesics like codeine, dihydromorphine, ergotamine, fentanyl, or morphine; angina preparations like diltiazem; antihistamines like cromoglycate, ketotifen, or naldoromib; anti-infectives like cephalosporins, penicillin, streptomycin, sulfonamides, tetracycline, or pentanemidine; antihistamines like desapride; anti-inflammatory drugs like beclomethasone, flunisolone, budesonide, teprednisolone, triamcinolone, or fluticasone; antitussives like noscarpine; and bronchodilators like ephedrine, adrenaline, fenoterol, or folic acid. Moterol, isoproterenol, orsinol, phenylephrine, phenylpropanolamine, pibuterol, taproterol, limterol, salbutamol, salmeterol, terbutaline, isotharine, tuloterol, or (-)-4-amino-3,5-dichloro-α-[[[6-[2-(2-pyridyl)ethoxy]hexyl]-amino]methyl]benzyl alcohol; diuretics, such as amiloride; anticholinergics, such as ipratropium, atropine, or oxytropine; hormones, such as cortisone, hydrocortisone, or prednisolone; xanthines, such as aminophylline, choline, lysine theophylline, or theophylline; and therapeutic proteins and peptides, such as insulin or glucagon. Those skilled in the art will appreciate that, where appropriate, a drug may be used in the form of a salt (e.g., as an alkali metal or amine salt or as an acid addition salt), an ester (e.g., a lower alkyl ester), or a solvate (e.g., a hydrate) to optimize the drug’s activity and / or stability.

[0296] The agents disclosed herein, or other suitable agents, are administered according to the condition being treated. Therefore, in some embodiments, one or more compounds of this disclosure will be administered co-administered with other agents as described above. When used in combination therapy, the compounds described herein are administered simultaneously or separately from a second agent. Such combined administration may include simultaneous administration of two agents in the same dosage form, simultaneous administration in separate dosage forms, and separate administration. In other words, the compounds described herein and any of the aforementioned agents may be formulated together in the same dosage form and administered simultaneously. Alternatively, the compounds of this disclosure and any of the aforementioned agents may be administered simultaneously, wherein the two agents are present in separate formulations. In another alternative, the compounds of this disclosure may be administered only after any of the agents described above, or vice versa. In some embodiments of the single-administration regimen, the compounds of this disclosure and any of the aforementioned agents are administered at intervals of minutes, hours, or days.

[0297] In some implementations, the compound of structure (I) is administered as a single therapy.

[0298] To identify signal transduction or mechanistic pathways and to detect interactions between various signal transduction pathways, scientists have developed suitable models or model systems, such as cell culture models and transgenic animal models. To determine certain stages in a signal transduction cascade, interacting compounds can be used to modulate the signal. The compounds of embodiments of this disclosure can also be used as reagents for testing NEK7-dependent signal transduction pathways in animal and / or cell culture models or in the clinical diseases mentioned in this application.

[0299] The methods of the embodiments of the present invention can be performed in vitro or in vivo. The sensitivity of specific cells to treatment with the compound of structure (I) can be specifically determined by in vitro testing (whether in research or in clinical application). Typically, cell cultures are combined with various concentrations of the compound for a period of time sufficient for the active agent to inhibit NEK7 activity, typically from about 1 hour to 1 week. Cultured cells from biopsy samples or cell lines can be used for in vitro treatment.

[0300] In some implementations, the compound of structure (I) inhibits the IC50 of NEK7. 50 The concentration of the compound required to inhibit 50% of NEK kinase activity was determined. Compounds with structure (I) showed IC50. 50 The potency value is less than about 5 mM, preferably less than about 1 mM, and even more preferably less than about 0.100 mM, as further described in the embodiments.

[0301] The examples and formulations provided below further illustrate and demonstrate the compounds of this disclosure and methods for preparing and testing such compounds. It should be understood that the scope of this disclosure is not limited in any way by the scope of the examples and formulations provided below. In the examples below, and throughout the specification and claims, unless otherwise stated, molecules having a single stereocenter exist as a racemic mixture. Unless otherwise stated, those molecules having two or more stereocenters exist as a racemic mixture of diastereomers. Single enantiomers / diastereomers can be obtained by methods known to those skilled in the art.

[0302] Example

[0303] The following embodiments are provided for illustrative purposes.

[0304] General Procedure

[0305] All proton NMR experiments were recorded at 400 MHz on a Bruker NEO spectrometer equipped with a BBFO probe. The deuterated solvent contained less than 0.05% v / v tetramethylsilane, which was used as a reference signal (set to 0.00 ppm). When the deuterated solvent did not contain tetramethylsilane, the residual undeuterated solvent peaks were used as reference signals according to published guidelines (J. Org. Chem. 1997, 62(21), 7512-7515). Chemical shifts are expressed in parts per million (ppm, δ units). Coupling constants are in Hertz (Hz). The split patterns depicted distinct multimodality and were designated as s (singleton), d (doublet), t (triplet), q (quartet), m (multiplet), qt (quintet), or brs (broad singlet).

[0306] LC / MS analysis was performed on an Agilent Technologies UHPLC 1290 Infinity II equipped with a G6125 MS detector.

[0307] Microwave reaction was performed using a Monowave 300 via Anton Paar GmbH using a standard protocol.

[0308] NEK7 enzyme assay

[0309] Casein substrate (a mixture of hydrolyzed and partially dephosphorylated α, β, and κ casein from bovine milk, obtained from Sigma-Aldrich, catalogue #C4765, diluted to a final concentration of 1 mg / mL in distilled water) and full-length recombinant human NEK7 (expressed by baculovirus in Sf9 insect cells using an N-terminal GST tag, obtained from SignalChem, catalogue #N09-10 G, 0.1 μg / μL) were mixed in assay buffer (20 mM Hepes pH 7.5, 10 mM MgCl2, 1 mM EGTA, 0.02% Brij35, 0.02 mg / mL BSA, 0.1 mM Na3VO4, 2 mM DTT, 1% DMSO). The compound of interest (serially 3-fold diluted in DMSO from 10 μM to 0.5 nM) or the mediator (1% DMSO) was partitioned into the kinase reaction mixture using an acoustic technique (Echo 550; nanoliter range). After incubating at room temperature for 20 minutes, add [ 33The kinase reaction was initiated by P-ATP (specific activity 10 μCi / μl), and the mixture was incubated at room temperature for 2 hours. The reaction was then stopped by spotting the reaction mixture onto cellulose phosphate P81 paper. After washing, the radioactivity of the P81 paper was measured, and the kinase activity data are expressed as the percentage of remaining kinase activity in the test sample compared to the reaction with the medium. IC50 was obtained using Prism (GraphPad Software). 50 Values ​​and curve fitting.

[0310] IL-1β release assay

[0311] Approximately 1.5 million THP-1 cells were seeded into each well of a 6-well TC plate and incubated with 40 nM PMA in RPMI (10% FBS, 1% Penstrep) for 24 h. The medium was then removed, and the cells were left to stand in RPMI (10% FBS, 1% Penstrep) for 24 h. Afterward, the medium was removed, and the cells were pretreated for 2 h with various concentrations of the compound of interest in RPMI (5% FBS) (typically serially diluted 3-fold in RPMI + 5% FBS, ranging from 1 μM to 0.5 nM). The medium was removed again, and the cells were incubated with 250 ng / mL LPS and the compound of interest (at the above concentrations) in RMPI (5% FBS) for 2 h. Finally, the medium was removed, and the cells were incubated with 20 μM Nigerianin and the compound of interest (at the above concentrations) in Opti-MEM for 30 min. The cell culture medium was then collected, and the amount of lysed IL-1β was determined using a JESS instrument (ProteinSimple) and standard protocols. The cleavage antibody Il-1β was obtained from Cell Signaling (catalog number #83186S) and used at a 1:20 dilution in antibody diluent 2. Protein Simple 1x anti-rabbit HRP secondary antibody was used with Protein Simple luminol and peroxide for chemiluminescent detection. The incubation time for the primary antibody was increased from 30 minutes to 60 minutes.

[0312] abbreviation:

[0313] ℃ (degrees Celsius); 11H NMR (proton nuclear magnetic resonance); ACN (acetonitrile); Boc (tert-butyloxycarbonyl); DCM (dichloromethane); DIPEA (N,N-diisopropylethylamine); DMAP (4-dimethylaminopyridine); DMF (N,N-dimethylformamide); DMSO-d6 (deuterated dimethyl sulfoxide); eq (equivalent); EtOAc (ethyl acetate); g (gram); (g gas); h (hour); HPLC (high performance liquid chromatography); LCMS (liquid chromatography-mass spectrometry); MeOH (methanol); mg (milligram) ); min (minutes); mL (milliliters); mmol (millimoles); N (normal); n-BuOH (1-butanol); Pd(PPh3)4 (palladium-tetra(triphenylphosphine)); PdCl2(dppf)([1,1'-bis(diphenylphosphino)ferrocene]palladium(II) chloride); sec- (secondary); TBAF (tetra-n-butylammonium fluoride); tert- (tertiary); TFA (trifluoroacetic acid); THF (tetrahydrofuran); TLC (thin-layer chromatography); UPLC (ultra-high performance liquid chromatography).

[0314] Preparation of synthetic intermediates

[0315] Intermediate A1

[0316] 1-Cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0317]

[0318] Copper(II) acetate (0.348 g, 1.916 mmol), 2,2'-bipyridine (0.299 g, 1.916 mmol), and sodium bicarbonate (0.322 g, 3.830 mmol) were added to a stirred solution of 3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.500 g, 1.916 mmol) and cyclopropylboronic acid (0.329 g, 3.830 mmol) in dichloroethane (10 mL). The resulting mixture was stirred at 70 °C under an oxygen atmosphere for 12 hours. After the reaction was complete (as shown by TLC), the reaction mixture was filtered through a diatomaceous earth pad, and the pad was then rinsed with DCM (20 mL × 2). The combined filtrate was washed with water (20 mL) and brine (25 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to obtain crude material, which was purified by rapid chromatography (silica gel 230-400 mesh, eluted with 20% EtOAc / petroleum ether) to give the title compound as a grayish-white solid (0.24 g, 36% yield). 1HNMR (400MHz, DMSO-d6) δ = 8.21 (s, 1H), 3.74-3.79 (m, 1H), 1.11-1.15 (m, 2H), 1.04-1.09 (m, 2H). LCMS: 301.8[M+H].

[0319] Intermediate A2

[0320] 3-Iodo-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0321]

[0322] In a sealed 25 mL tube, Cs₂CO₃ (12.38 g, 38.31 mmol) and 2-iodopropane (3.60 g, 21.16 mmol) were added to a stirred solution of 3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine (5.00 g, 19.15 mmol) in DMF (25 mL). The reaction mixture was stirred at 90 °C for 16 h, and after the reaction was complete (as indicated by TLC), it was poured into crushed ice (50 g) and stirred for 15 min. The resulting solid was filtered, washed with water (2 × 5 mL), and dried to give the title compound (3.25 g, 56% yield) as an off-white solid. 1 H NMR (400MHz, DMSO-d6) δ = 8.18 (s, 1H), 4.93-4.99 (m, 1H), 1.42 (d, J = 6.8Hz, 6H). LCMS: 303.8[M+H].

[0323] Intermediate A3 to Intermediate A12

[0324] The following intermediates are prepared via a similar procedure to that described for intermediate A2, using a suitable reagent (alkyl halide or toluenesulfonate) instead of 2-iodopropane as shown below:

[0325]

[0326]

[0327]

[0328] Intermediate A13

[0329] 1-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol

[0330]

[0331] NaH₂PO₄ (0.044 g, 0.372 mmol) was added to a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.100 g, 0.380 mmol), 2,2-dimethylethylene oxide (0.055 g, 0.760 mmol), and K₂CO₃ (0.050 g, 0.372 mmol) in acetonitrile (3 mL) and water (1 mL), and the resulting solution was microwave-treated at 150 °C for 1 hour. After the reaction was complete (as shown by TLC), the solvent was removed under reduced pressure to obtain a crude material, which was purified by rapid chromatography (silica gel 230-400 mesh, eluted with 25% EtOAc / petroleum ether) to give the title compound (0.064 g, 51% yield) as a pale brown solid. 1 H NMR (400MHz, DMSO-d6) δ = 8.20 (s, 1H), 4.19 (s, 2H), 1.09 (s, 6H). LCMS: 334.0[M+H].

[0332] Intermediate A14

[0333] 3-Iodo-1-(pyridin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0334]

[0335] The title compound was prepared from 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (1.00 g, 3.83 mmol) and pyridin-4-ylboronic acid (0.94 g, 7.66 mmol) via a procedure similar to that described for intermediate A1, and was obtained as a pale brown solid (0.27 g, 21% yield). LCMS: 338.8 [M+H].

[0336] Intermediate A15

[0337] 3-Iodo-1-methyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0338]

[0339] At 0 °C, in a sealed 25 mL tube, Cs₂CO₃ (0.780 g, 2.394 mmol) and methyl iodide (0.138 mL, 2.203 mmol) were added to a solution of 3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.500 g, 1.916 mmol) in DMF (3 mL). The reaction mixture was stirred at 25 °C for 1 hour, and after the reaction was complete (as indicated by TLC), it was poured into crushed ice (50 g) and stirred for 30 minutes. The resulting solid was filtered, washed with water (2 × 5 mL), and dried to give the title compound (0.380 g, 67% yield) as a yellow solid. 1 H NMR (400MHz, DMSO-d6) δ = 8.22 (s, 1H), 6.79 (s, 2H), 3.89 (s, 3H). LCMS: 276.0[M+H].

[0340] Intermediate A16

[0341] 1-Cyclopropyl-3-iodo-1H-pyrazolo[4,3-c]pyridine-4-amine

[0342] Step 1: Synthesis of 4-chloro-3-iodo-1H-pyrazolo[4,3-c]pyridine

[0343]

[0344] KOH (1.320 g, 23.0 mmol) and iodine (1.620 g, 12.8 mmol) were added to a solution of 4-chloro-1H-pyrazolo[4,3-c]pyridine (1.000 g, 6.4 mmol) in dioxane (10 mL), and the resulting mixture was stirred at 75 °C for 4 hours. After the reaction was complete (as shown by TLC), the reaction mixture was filtered through a diatomaceous earth pad, and the filtrate was concentrated under reduced pressure to give a crude material, which was purified by reversed-phase column chromatography to give the title compound (0.633 g, 63% yield) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ=14.12 (bs, 1H), 8.14 (d, J=6.0Hz, 1H), 7.66 (d, J=5.6Hz, 1H). LCMS: 279.9[M+H].

[0345] Step 2: Synthesis of 4-chloro-1-cyclopropyl-3-iodo-1H-pyrazolo[4,3-c]pyridine

[0346]

[0347] The title compound was prepared from 4-chloro-3-iodo-1H-pyrazolo[4,3-c]pyridine (0.630 g, 2.20 mmol) and cyclopropylboronic acid (0.329 g, 3.83 mmol) via a procedure similar to that described for intermediate A1, and was obtained as a white solid (0.430 g, 60% yield). 1 H NMR (400MHz, DMSO-d6) δ=8.21 (d, J=6.0Hz, 1H), 7.81 (d, J=5.6Hz, 1H), 3.84-3.89 (m, 1H), 1.14-1.17 (m, 4H). LCMS: 319.7[M+H].

[0348] Step 3: Synthesis of 1-cyclopropyl-3-iodo-1H-pyrazolo[4,3-c]pyridine-4-amine

[0349]

[0350] A mixture of 0.20 g of 4-chloro-1-cyclopropyl-3-iodo-1H-pyrazolo[4,3-c]pyridine and an aqueous solution of ammonium hydroxide (25% aqueous solution, 8 mL) was microwave-irradiated at 150 °C for 2 h. After the reaction was complete (as shown by TLC), the reaction mixture was concentrated under reduced pressure to give the title compound (0.19 g, quantitative yield) as a grayish-white solid. 1 H NMR (400MHz, DMSO-d6) δ = 8.11 (s, 1H), 7.38 (s, 1H), 6.68 (bs, 2H), 3.48-3.54 (m, 1H), 0.97-0.99 (m, 4H). LCMS: 301.0[M+H].

[0351] Intermediate A17

[0352] 1-(3-(benzyloxy)cyclobutyl)-3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0353]

[0354] 3-(benzyloxy)cyclobutyl methanesulfonate (prepared as described in PCT No. WO2019 / 092170, 0.491 g, 1.916 mmol) and Cs₂CO₃ (0.624 g, 1.916 mmol) were added to a solution of 3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.250 g, 0.958 mmol) in DMF (5 mL), and the resulting mixture was stirred at 90 °C for 12 h. After the reaction was complete (as shown by TLC), the reaction mixture was poured into ice water (50 mL) and extracted with ethyl acetate (2 × 30 mL). The combined organic layers were dried over Na₂SO₄, filtered, and evaporated under reduced pressure to give the title compound (0.400 g, 75% LCMS purity) as a light brown gel, which was used without further purification. LCMS: 422.0 [M+H].

[0355] Intermediate A18

[0356] 3-Iodo-1-(1-methylazacyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0357]

[0358] The title compound was prepared as described in PCT Publication No. WO 2002 / 076986.

[0359] Intermediate A19

[0360] 1-Cyclopropyl-N-(2,4-Dimethoxybenzyl)-3-iodo-1H-pyrazolo[4,3-c]pyridine-4-amine

[0361]

[0362] A mixture of 4-chloro-1-cyclopropyl-3-iodo-1H-pyrazolo[4,3-c]pyridine (A16, 0.880 g, 2.75 mmol) and (2,5-dimethoxyphenyl)methylamine (1.245 mL, 8.26 mmol) in n-BuOH (10 mL) was stirred at 110 °C for 12 h. After the reaction was complete (as shown in LCMS), the reaction mixture was concentrated under reduced pressure to give a crude material, which was purified by Isolera (silica gel 230-400 mesh, eluted with 40% EtOAc / petroleum ether) to give the title product (1.0 g, 80% yield) as a yellow gel. 1H NMR (400MHz, DMSO-d6) δ=7.81 (d, J=6.4Hz, 1H), 7.19 (d, J=8.4Hz, 1H), 6.87 (d, J=6.0Hz, 1H), 6.60 (d, J=2.4Hz, 1H), 6.53 -6.56 (m, 1H), 6.45-6.47 (m, 1H), 4.62 (d, J=5.6Hz, 2H), 3.88 (s, 3H), 3.73 (s, 3H), 3.66-3.70 (m, 1H), 1.01-1.10 (m, 4H). LCMS: 451.0[M+H].

[0363] Intermediate A20

[0364] 2-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethane-1-ol

[0365]

[0366] The title compound was prepared as described in PCT Publication No. WO 2011 / 119663.

[0367] Intermediate A21

[0368] 3-Iodo-1-(2-methoxyethyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0369]

[0370] K₂CO₃ (0.397 g, 2.870 mmol) and 1-bromo-2-methoxyethane (0.319 g, 2.299 mmol) were added to a solution of 3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.500 g, 1.916 mmol) in DMF (6 mL), and the resulting mixture was stirred in a sealed tube at 80 °C for 12 h. After the reaction was complete (as shown by TLC), the reaction mixture was poured into crushed ice (25 g) and extracted with EtOAc (2 × 50 mL). The combined organic extracts were dried over Na₂SO₄, filtered, and concentrated under reduced pressure to give the title product (0.500 g, crude), which was used without further purification. 1 H NMR (400MHz, DMSO-d6) δ = 8.20 (s, 1H), 4.43 (t, J = 5.6Hz, 2H), 3.75 (t, J = 5.6Hz, 2H), 3.20 (s, 3H). LCMS: 319.8[M+H].

[0371] Intermediate A22

[0372] 3-Iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0373]

[0374] The title compound was prepared as described in USPTO Publication No. WO 2015 / 165279.

[0375] Intermediate A23

[0376] 3-Iodo-1-(1-methylpiperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0377]

[0378] The title compound was prepared as described in USPTO Publication No. WO 2018 / 121228.

[0379] Intermediate B1

[0380] 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0381]

[0382] A mixture of 1-cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (Al, 0.500 g, 1.66 mmol), 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.433 g, 1.82 mmol) and K₂CO₃ (0.688 g, 4.98 mmol) in 1,4-dioxane (25 mL) and water (2.5 mL) was purged with N₂ for 10 min. Pd(PPh₃)₄ (0.092 g, 0.08 mmol) was then added, and the reaction mixture was stirred at 100 °C for 16 h. After the reaction was complete (as shown by TLC), the mixture was filtered through a diatomaceous earth mat, which was then rinsed with EtOAc (2 × 10 mL). The combined filtrates were concentrated under reduced pressure to obtain crude material, which was purified by rapid chromatography (silica gel 230-400 mesh, eluted with 2% MeOH / DCM) to give the title compound as a yellow solid (0.46 g, 98% yield). 1H NMR (400MHz, DMSO-d6) δ = 8.23 ​​(s, 1H), 7.15-7.24 (m, 2H), 6.87-6.91 (m, 1H) ), 5.47(bs, 2H), 3.80-3.84(m, 1H), 1.18-1.19(m, 2H), 1.05-1.08(m, 2H). LCMS: 285.0[M+H].

[0383] Intermediate B2

[0384] 3-(4-aminophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0385] Step 1: Synthesis of 1-isopropyl-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0386]

[0387] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A2, 1.887 g, 6.23 mmol) and (4-nitrophenyl)boronic acid (1.56 g, 9.34 mmol), and was obtained as a yellow solid (1.242 g, 67% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.38-8.40 (m, 2H), 8.28 (s, 1H), 7.92-7.95 (m, 2H), 5.07-5.14 (m, 1H), 1.51 (d, J = 6.8Hz, 6H). LCMS: 299.1[M+H].

[0388] Step 2: Synthesis of 3-(4-aminophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0389]

[0390] Iron powder (2.320 g, 41.60 mmol) and ammonium chloride (2.220 g, 41.60 mmol) were added to a stirred solution of 1-isopropyl-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (1.242 g, 4.16 mmol) in ethanol (50 mL) and water (20 mL), and the resulting mixture was heated to 80 °C for 3 hours. After the reaction was complete (as shown by TLC), the mixture was filtered through a diatomaceous earth mat, and the mat was then rinsed with EtOAc (2 × 25 mL). The combined filtrates were concentrated under reduced pressure, and the residue was dissolved in EtOAc (100 mL), washed with brine (25 mL), dried over Na₂SO₄, filtered, and evaporated under reduced pressure to give the title compound (1.042 g, quantitative yield) as a pale yellow solid, which was used without further purification.

[0391] Intermediate B3

[0392] 3-(4-amino-3-fluorophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0393] Step 1: Synthesis of 3-(3-fluoro-4-nitrophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0394]

[0395] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A2, 0.10 g, 0.32 mmol) and (3-fluoro-4-nitrophenyl)boronic acid (0.71 g, 0.39 mmol), and was obtained as a yellow solid (0.07 g, 67% yield). LCMS: 315.1 [MH].

[0396] Step 2: Synthesis of 3-(4-amino-3-fluorophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0397]

[0398] The title compound was prepared from 3-(3-fluoro-4-nitrophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.07 g, 0.22 mmol) and Fe / NH4Cl via a procedure similar to that described in step 2 of intermediate B2, and was obtained as a pale yellow solid (0.09 g, quantitative yield), which was used without further purification. LCMS: 287.1 [M+H].

[0399] intermediate B4

[0400] 3-(4-aminophenyl)-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0401] Step 1: Synthesis of 3-(4-nitrophenyl)-1-(oxetane-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0402]

[0403] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-(oxetane-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A3, 0.800 g, 2.522 mmol) and (4-nitrophenyl)boronic acid (0.632 g, 3.78 mmol), and was obtained as a yellow solid (0.596 g, 76% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.41-8.43 (m, 2H), 8.30 (s, 1H), 7.99-8.01 (m, 2H), 6.05-6.08 (m, 1H), 4.97-5.12 (m, 4H). LCMS: 311.0[MH].

[0404] Step 2: Synthesis of 3-(4-aminophenyl)-1-(oxetane-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0405]

[0406] The title compound was prepared via a similar procedure as described in step 2 of intermediate B2, starting from 3-(4-nitrophenyl)-1-(oxetane-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.596 g, 1.91 mmol) and Fe / NH4Cl, and was obtained as a pale yellow solid (0.42 g, quantitative yield), which was used without further purification. LCMS: 283.0 [M+H].

[0407] Intermediate B5

[0408] 3-(4-amino-3-fluorophenyl)-1-(oxetane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0409]

[0410] The title compound was prepared via a procedure similar to that described for intermediate B1, starting with 3-iodo-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A3, 0.110 g, 0.346 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaboranecyclopentane-2-yl)aniline (0.099 g, 0.420 mmol), and was obtained as a pale yellow gel (0.077 g, 74% yield). LCMS: 317.1 [M+H].

[0411] Intermediate B6

[0412] 1-Allyl-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0413]

[0414] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-allyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A4, 0.15 g, 0.49 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.13 g, 0.54 mmol), and was obtained as a yellow gel (0.13 g, 92% yield). LCMS: 285.0 [M+H].

[0415] Intermediate B7

[0416] 3-(4-aminophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0417] Step 1: Synthesis of 1-cyclopropyl-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0418]

[0419] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A1, 0.110 g, 0.36 mmol) and (4-nitrophenyl)boronic acid (0.067 g, 0.40 mmol), and was obtained as a pale yellow solid (0.060 g, 56% yield). 1H NMR (400MHz, DMSO-d6) δ=8.37 (d, J=8.8Hz, 2H), 8.30 (s, 1H), 7.91 (d, J=8.8Hz, 2H), 3.91-3.94 (m, 1H), 1.23-1.24 (m, 2H), 1.11-1.14 (m, 2H). LCMS: 297.0[M+H].

[0420] Step 2: Synthesis of 3-(4-aminophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0421]

[0422] The title compound was prepared from 1-cyclopropyl-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.060 g, 0.2 mmol) and Fe / NH4Cl via a procedure similar to that described in step 2 of intermediate B2, and was obtained as a pale yellow solid (0.047 g, 89% yield) used without further purification. LCMS: 266.9 [M+H].

[0423] Intermediate B8

[0424] 3-(4-aminophenyl)-1-cyclobutyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0425] Step 1: Synthesis of 1-cyclobutyl-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0426]

[0427] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-cyclobutyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A5, 0.150 g, 0.47 mmol) and (4-nitrophenyl)boronic acid (0.087 g, 0.52 mmol), and was obtained as a pale yellow solid (0.160 g, quantitative yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.34-8.41 (m, 2H), 8.28 (s, 1H), 7.94-7.97 (m, 2H), 5.32-5.43 (m, 1H), 2.65-2.75 (m, 2H), 1.86-1.94 (m, 4H). LCMS: 311.2[M+H].

[0428] Step 2: Synthesis of 3-(4-aminophenyl)-1-cyclobutyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0429]

[0430] The title compound was prepared from 1-cyclobutyl-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.16 g, 0.51 mmol) and Fe / NH4Cl via a procedure similar to that described in step 2 of intermediate B2, and was obtained as a pale yellow solid (0.140 g, quantitative yield) used without further purification. LCMS: 281.0 [M+H].

[0431] Intermediate B9

[0432] 3-(4-amino-3-fluorophenyl)-1-cyclobutyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0433]

[0434] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-cyclobutyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A5, 0.250 g, 0.793 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.225 g, 0.952 mmol), and was obtained as a pale yellow solid (0.100 g, 32% yield). LCMS: 299.1 [M+H].

[0435] Intermediate B10

[0436] 3-(4-aminophenyl)-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0437] Step 1: Synthesis of 3-(4-nitrophenyl)-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0438]

[0439] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A6, 0.165 g, 0.49 mmol) and (4-nitrophenyl)boronic acid (0.091 g, 0.54 mmol), and was obtained as a pale yellow solid (0.086 g, 53% yield). 1H NMR (400MHz, DMSO-d6) δ = 8.38-8.40 (m, 2H), 8.30 (s, 1H), 7.93-7.95 (m, 2H), 5.52-5.58 (m, 1H), 4.06-4.15 (m, 2H), 3.88-3.99 (m, 2H), 2.50-2.51 (m, 2H). LCMS: 327.2[M+H].

[0440] Step 2: Synthesis of 3-(4-aminophenyl)-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0441]

[0442] The title compound was prepared by a procedure similar to that described in step 2 of intermediate B2, starting with 3-(4-nitrophenyl)-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (0.115 g, 0.35 mmol) and Fe / NH4Cl, and was obtained as a pale yellow solid (0.083 g, 80% yield), which was used without further purification. 1 H NMR (400MHz, DMSO-d6) δ = 8.22 (s, 1H), 7.33 (dd, J = 2.0, 6.6Hz, 2H), 6.71 (dd, J = 2.0, 6.4Hz, 2H ), 5.48(bs, 2H), 5.44-5.47(m, 1H), 4.04-4.12(m, 2H), 3.86-3.94(m, 2H), 2.34-2.41(m, 2H). LCMS: 296.9[M+H].

[0443] Intermediate B11

[0444] 3-(4-amino-3-fluorophenyl)-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0445]

[0446] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A6, 0.351 g, 1.00 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.301 g, 1.27 mmol), and was obtained as a pale yellow solid (0.200 g, 60% yield). 1H NMR (400MHz, DMSO-d6) δ = 8.23 ​​(s, 1H), 7.24-7.28 (m, 1H), 7.18-7.21 (m, 1H), 6.88-6.92 (m, 1H), 5.46-5.50 (m, 3H), 4.04-4.12 (m, 2H), 3.87-3.94 (m, 2H), 2.33-2.41 (m, 2H). LCMS: 315.1 [M+H].

[0447] Intermediate B12

[0448] 3-(4-aminophenyl)-1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0449] Step 1: Synthesis of 3-(4-nitrophenyl)-1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0450]

[0451] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A7, 0.094 g, 0.273 mmol) and (4-nitrophenyl)boronic acid (0.055 g, 0.328 mmol), and was obtained as a pale yellow solid (0.078 g, 84% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.37-8.40 (m, 2H), 8.29 (s, 1H), 7.93-7.96 (m, 2H), 4.96-5.0 2(m, 1H), 4.00-4.04(m, 2H), 3.54-3.59(m, 2H), 2.16-2.24(m, 2H), 1.89-1.94(m, 2H). LCMS: 340.9[M+H].

[0452] Step 2: Synthesis of 3-(4-aminophenyl)-1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0453]

[0454] The title compound was prepared by a procedure similar to that described in step 2 of intermediate B2, starting with 3-(4-nitrophenyl)-1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.078 g, 0.23 mmol) and Fe / NH4Cl, and was obtained as a pale yellow solid (0.052 g, 73% yield), which was used without further purification. LCMS: 311.1 [M+H].

[0455] Intermediate B13

[0456] 3-(4-amino-3-fluorophenyl)-1-(tetrahydro-2H-pyran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0457]

[0458] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-(tetrahydro-2H-pyran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A8, 0.050 g, 0.14 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.040 g, 0.17 mmol), and was obtained as a pale yellow solid (0.015 g, 32% yield). LCMS: 329.2 [M+H].

[0459] Intermediate B14

[0460] 3-(4-amino-3-fluorophenyl)-1-(3,3-difluorocyclobutyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0461]

[0462] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-(3,3-difluorocyclobutyl)-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A9, 0.100 g, 0.285 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.810 g, 0.341 mmol), and was obtained as a pale yellow solid (0.088 g, 93% yield). LCMS: 334.9 [M+H].

[0463] Intermediate B15

[0464] 4-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)cyclohexane-1-ol

[0465] Step 1: Synthesis of 4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)cyclohexane-1-ol

[0466]

[0467] An aqueous solution of HCl (1.5 N, 5 mL) was added to a solution of 3-iodo-1-(1,4-dioxaspiro[4.5]dec-8-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine (A10, 0.313 g, 0.780 mmol), and the resulting mixture was stirred at room temperature for 12 hours. After the reaction was complete (as shown in UPLC), the solvent was removed under reduced pressure to obtain a residue (0.232 g, LCMS: 358.0 [M+H]), which was dissolved in THF (5 mL). The resulting solution was cooled to 0 °C, NaBH4 (0.050 g, 1.322 mmol) was added, and the mixture was stirred at room temperature for 1 hour. After the reaction was complete (as shown in UPLC), an aqueous solution of HCl (1.5 N, 5 mL) was added, and the mixture was extracted with EtOAc (2 × 10 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the title product (0.196 g) as a light brown gel, which was used without further purification. LCMS: 359.8 [M+H].

[0468] Step 2: Synthesis of 4-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)cyclohexane-1-ol

[0469]

[0470] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)cyclohexane-1-ol (0.196 g, 0.550 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.160 g, 0.655 mmol), and was obtained as a yellow gel (0.120 g, 64% yield). LCMS: 343.0 [M+H].

[0471] Intermediate B16

[0472] 3-(4-amino-3-fluorophenyl)-1-(1-(oxecyclobutane-3-yl)piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0473] Step 1: Synthesis of tert-butyl 4-(4-amino-3-(3-fluoro-4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidine-1-carboxylate

[0474]

[0475] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with tert-butyl 4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-carboxylate (A11, 0.10 g, 0.23 mmol) and (3-fluoro-4-nitrophenyl)boronic acid (0.05 g, 0.27 mmol), and was obtained as a yellow solid (0.04 g, 40% yield). LCMS: 458.1 [MH].

[0476] Step 2: Synthesis of 3-(3-fluoro-4-nitrophenyl)-1-(piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0477]

[0478] TFA (0.5 mL) was added to a solution of tert-butyl 4-(4-amino-3-(3-fluoro-4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-carboxylate (100 mg, 0.218 mmol) in anhydrous DCM (5 mL), and the resulting mixture was stirred at room temperature for 2 hours. After the reaction was complete (as shown by UPLC), the solvent was evaporated under reduced pressure to give the title product (110 mg) as a light brown gel, which was used without further purification. LCMS: 358.1 [MH].

[0479] Step 3: Synthesis of 3-(3-fluoro-4-nitrophenyl)-1-(1-(oxetane-3-yl)piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0480]

[0481] Oxetane-3-one (0.020 g, 0.277 mmol) and glacial acetic acid (catalytic amount) were added to a solution of 3-(3-fluoro-4-nitrophenyl)-1-(piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.100 g, 0.280 mmol) in DCM (5 mL), and the resulting mixture was stirred at room temperature for 4 hours. Then, sodium triacetoxyborohydride (0.178 g, 0.840 mmol) was added, and the resulting mixture was stirred at room temperature for 12 hours. After the reaction was complete (as shown in UPLC), the solution was diluted with DCM (5 mL) and washed with 10% NaHCO3 aqueous solution (5 mL) and brine (5 mL). The organic layer was dried over Na2SO4, filtered, and the solvent was evaporated under reduced pressure to give the title product (110 mg), which was used without further purification. LCMS: 414.2 [MH].

[0482] Step 4: Synthesis of 3-(4-amino-3-fluorophenyl)-1-(1-(oxetane-3-yl)piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0483]

[0484] Iron powder (0.135 g, 2.417 mmol) and ammonium chloride (0.142 g, 2.655 mmol) were added to a solution of 3-(3-fluoro-4-nitrophenyl)-1-(1-(oxetane-3-yl)piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.110 g, 0.266 mmol) in ethanol (5 mL) and water (2 mL), and the resulting mixture was stirred at 80 °C for 2 hours. After the reaction was complete (as shown by TLC), the reaction mixture was filtered through a diatomaceous earth mat, and the mat was then washed with EtOAc (5 mL × 2). The combined filtrates were concentrated under reduced pressure to obtain a residue, which was dissolved in EtOAC (10 mL) and then washed with brine (5 mL). The organic layer was separated, dried over Na2SO4, filtered, and concentrated under reduced pressure to give the title compound (0.05 g, 50% yield) as a grayish-white solid, which was used without further purification. LCMS: 383.9 [M+H].

[0485] Intermediate B17

[0486] 3-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydrothiophene 1,1-dioxide

[0487]

[0488] The title compound was prepared by a procedure similar to that described for intermediate B1, starting from 3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydrothiophene 1,1-dioxide (A12, 0.08 g, 0.21 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.06 g, 0.25 mmol), and was obtained as a yellow gel (0.03 g, 40% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.26 (s, 1H), 7.27-7.31 (m, 1H), 7.20-7.23 (m, 1H), 6.88-6.93 (m, 1H), 5.69(bs, 2H), 5.53(bs, 2H), 3.72-3.78(m, 2H), 3.51-3.56(m, 2H), 2.61-2.70(m, 2H). LCMS: 362.8[M+H].

[0489] Intermediate B18

[0490] 1-(4-amino-3-(4-aminophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol

[0491] Step 1: Synthesis of 1-(4-amino-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol

[0492]

[0493] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol (A13, 0.067 g, 0.201 mmol) and (4-nitrophenyl)boronic acid (0.054 g, 0.302 mmol), and was obtained as a yellow solid (0.059 g, 90% yield). 1 H NMR (400MHz, DMSO-d6) δ=8.40 (d, J=7.6Hz, 2H), 8.30 (s, 1H), 7.94 (d, J=6.8Hz, 2H), 4.32 (bs, 2H), 1.16 (bs, 6H). LCMS: 328.9[MH].

[0494] Step 2: Synthesis of 1-(4-amino-3-(4-aminophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol

[0495]

[0496] The title compound was prepared from 1-(4-amino-3-(4-nitrophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol (0.072 g, 0.219 mmol) and Fe / NH4Cl via a procedure similar to that described in step 2 of intermediate B2, and was obtained as a pale yellow solid (0.07 g, quantitative yield), which was used without further purification. LCMS: 299.0 [M+H].

[0497] Intermediate B19

[0498] 1-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol

[0499]

[0500] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol (A13, 0.110 g, 0.330 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.094 g, 0.396 mmol), and was obtained as a pale yellow solid (0.077 g, 66% yield). LCMS: 317.1 [M+H].

[0501] Intermediate B20

[0502] 3-(4-amino-3-fluorophenyl)-1-(pyridin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0503]

[0504] The title compound was prepared via a procedure similar to that described for intermediate B1, starting with 3-iodo-1-(pyridin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A14, 0.250 g, 0.73 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.193 g, 0.81 mmol), and was obtained as a yellow gel (0.140 g, 59% yield). LCMS: 321.9 [M+H].

[0505] Intermediate B21

[0506] 3-(4-amino-3-chlorophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0507] Step 1: Synthesis of 3-(3-chloro-4-nitrophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0508]

[0509] The title compound was prepared by a similar procedure to that described for intermediate B1, starting with 3-iodo-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A2, 0.115 g, 0.379 mmol) and 2-(3-chloro-4-nitrophenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentane (0.129 g, 0.455 mmol), and was obtained as a grayish-white solid (0.083 g, 66% yield). 1 HNMR (400MHz, DMSO-d6) δ = 8.28 (s, 1H), 8.21-8.23 (m, 1H), 7.95-7.95 (m, 1H), 7.82-7.85 (m, 1H), 5.07-5.13 (m, 1H), 1.51 (d, J = 6.4Hz, 6H). LCMS: 332.9[M+H].

[0510] Step 2: Synthesis of 3-(4-amino-3-chlorophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0511]

[0512] The title compound was prepared by a procedure similar to that described in step 2 of intermediate B2, starting with 3-(3-chloro-4-nitrophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.083 g, 0.25 mmol) and Fe / NH4Cl, and was obtained as a pale yellow solid (0.900 g, quantitative yield), which was used without further purification. LCMS: 302.9 [M+H].

[0513] Intermediate B22

[0514] 3-(4-amino-3-chlorophenyl)-1-methyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0515]

[0516] The title compound was prepared from 3-iodo-1-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A15, 0.100 g, 0.364 mmol) and 2-chloro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.092 g, 0.364 mmol) via a procedure similar to that described for intermediate B1, and was obtained as a yellow solid (0.094 g, 94% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.23 ​​(s, 1H), 7.45-7.46 (m, 1H), 7.31-7.33 (m, 1H), 6.91-6.93 (m, 1H), 5.68 (bs, 2H), 3.91 (s, 3H). LCMS: 275.0[M+H].

[0517] Intermediate B23

[0518] 3-(4-amino-3-fluorophenyl)-1-methyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0519]

[0520] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 3-iodo-1-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B15, 0.110 g, 0.400 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.104 g, 0.440 mmol), and was obtained as a brown gel (0.130 g, 97% yield). LCMS: 259.1 [M+H].

[0521] Intermediate B24

[0522] 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridine-4-amine

[0523]

[0524] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[4,3-c]pyridine-4-amine (A16, 0.190 g, 0.63 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.180 g, 0.75 mmol), and was obtained as a yellow gel (0.070 g, 39% yield). 1H NMR (400MHz, DMSO-d6) δ = 7.75-7.76 (m, 1H), 7.21-7.24 (m, 1H), 7.14-7.16 (m, 2H), 6 .85-6.87(m, 1H), 5.78(bs, 2H), 5.46(bs, 2H), 3.67-3.69(m, 1H), 1.09-1.10(m, 4H). LCMS: 317.1[M+H].

[0525] Intermediate B25

[0526] 3-(4-aminophenyl)-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridine-4-amine

[0527] Step 1: Synthesis of 1-cyclopropyl-3-(4-nitrophenyl)-1H-pyrazolo[4,3-c]pyridine-4-amine

[0528]

[0529] The title compound was prepared by a procedure similar to that described for intermediate B1, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[4,3-c]pyridine-4-amine (A16, 0.190 g, 0.63 mmol) and (4-nitrophenyl)boronic acid (0.126 g, 0.75 mmol), and was obtained as a pale yellow solid (0.090 g, 50% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.36-8.39 (m, 2H), 7.93-7.96 (m, 2H), 7.84 (d, J = 6.0Hz , 1H), 6.95 (d, J=6.0Hz, 1H), 5.98 (bs, 2H), 3.78-3.81 (m, 1H), 1.13-1.16 (m, 4H). LCMS: 296.1[M+H].

[0530] Step 2: Synthesis of 3-(4-aminophenyl)-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridine-4-amine

[0531]

[0532] The title compound was prepared from 1-cyclopropyl-3-(4-nitrophenyl)-1H-pyrazolo[4,3-d]pyrimidine-4-amine (0.093 g, 0.31 mmol) and Fe / NH4Cl via a procedure similar to that described in step 2 of intermediate B2, and was obtained as a pale yellow solid (0.052 g, 63% yield) used without further purification. LCMS: 266.0 [M+H].

[0533] Intermediate B26

[0534] 3-(4-amino-3-fluorophenyl)-1-(3-(benzyloxy)cyclobutyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0535]

[0536] A mixture of 1-(3-(benzyloxy)cyclobutyl)-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A17, 0.400 g, 0.712 mmol), 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhexacyclopentan-2-yl)aniline (0.186 g, 0.783 mmol) and K2CO3 (0.197 g, 1.424 mmol) in dioxane (7 mL), ethanol (3 mL), and water (3 mL) was purged with N2 for 10 min. Then PdCl2 (dppf) (0.026 g, 0.036 mmol) was added, and the resulting mixture was again purged with N2 for 10 min, and then stirred in a sealed tube at 80 °C for 16 h. After the reaction was complete (as shown by TLC and LCMS), the reaction mixture was filtered through a diatomaceous earth pad, which was then washed with EtOAc (3 × 10 mL × 3). The combined filtrates were concentrated under reduced pressure to give a crude material, which was purified by rapid chromatography (silica gel 230-400 mesh, eluted with 4% MeOH / DCM) to give the title compound (0.179 g, 62% yield) as a pale brown solid. 1 H NMR (400MHz, DMSO-d6) δ = 8.21 (s, 1H), 7.20-7.40 (m, 7H), 6.89-6.93 (m, 1H), 5.42-5.48 (m, 3H), 4.47-4.48 (m, 3H), 2.74-2.83 (m, 2H), 2.60 (s, 2H). LCMS: 405.1[M+H].

[0537] Intermediate B27

[0538] 3-(4-amino-3-methylphenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0539]

[0540] The title compound was prepared via a procedure similar to that described for intermediate B26, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A1, 0.250 g, 0.830 mmol) and 2-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.213 g, 0.913 mmol), and was obtained as a brown gel (0.22 g), which was used without further purification. LCMS: 281.3 [M+H].

[0541] Intermediate B28

[0542] 3-(4-amino-3-fluorophenyl)-1-(1-methylazacyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0543]

[0544] The title compound was obtained by following a similar procedure to that described for intermediate B26, starting with 3-iodo-1-(1-methylazacyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A18, 0.380 g, 1.151 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecylpentane-2-yl)aniline (0.300 g, 1.266 mmol), and was obtained as a brown solid (0.26 g, 48% yield). LCMS: 314.4 [M+H].

[0545] Intermediate B29

[0546] 3-(4-amino-2,5-difluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0547]

[0548] The title compound was obtained by following a procedure similar to that described for intermediate B26, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A1, 0.165 g, 0.548 mmol) and 2,5-difluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (prepared as described in PCT Publication No. WO 2020 / 172093, 0.300 g, 1.266 mmol), and was obtained as a pale yellow solid (0.051 g, 27% yield). LCMS: 303.2 [M+H].

[0549] Intermediate B30

[0550] (2-amino-5-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)methanol

[0551]

[0552] The title compound was obtained by following a similar procedure to that described for intermediate B26, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A1, 0.250 g, 0.830 mmol) and (2-amino-5-(4,4,5,5-tetramethyl-1,3-dioxacyclopentan-2-yl)phenyl)methanol (prepared as described in PCT Publication No. WO 2011 / 8130628, 0.207 g, 0.830 mmol), and was obtained as a light pink solid (0.122 g, 47% yield). LCMS: 297.1 [M+H].

[0553] Intermediate B31

[0554] 3-(6-aminopyridin-3-yl)-1-cyclopropyl-N-(2,4-dimethoxybenzyl)-1H-pyrazolo[4,3-c]pyridin-4-amine

[0555]

[0556] The title compound was obtained by following a similar procedure to that described for intermediate B26, starting with 1-cyclopropyl-N-(2,4-dimethoxybenzyl)-3-iodo-1H-pyrazolo[4,3-c]pyridine-4-amine (A19, 0.300 g, 0.666 mmol) and 5-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)pyridine-2-amine (0.147 g, 0.666 mmol), and was obtained as a brown gel (0.11 g, 30% yield). LCMS: 416.9 [M+H].

[0557] Intermediate B32

[0558] 2-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethane-1-ol

[0559]

[0560] The title compound was obtained by following a similar procedure to that described for intermediate B26, starting with 2-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethane-1-ol (A20, 0.420 g, 1.377 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.327 g, 1.379 mmol), and was obtained as a brown solid (0.390 g, 92% yield). LCMS: 289.1 [M+H].

[0561] Intermediate B33

[0562] 3-(4-amino-3-fluorophenyl)-1-(2-methoxyethyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0563]

[0564] The title compound was obtained by following a procedure similar to that described for intermediate B26, starting with 3-iodo-1-(2-methoxyethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A21, 0.620 g, 1.943 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.461 g, 1.943 mmol), and was obtained as a brown solid (0.42 g), which is suitable without further purification. LCMS: 302.9 [M+H].

[0565] Intermediate B34

[0566] 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-N-(2,4-dimethoxybenzyl)-1H-pyrazolo[4,3-c]pyridine-4-amine

[0567]

[0568] The title compound was obtained by following a procedure similar to that described for intermediate B26, starting with 1-cyclopropyl-N-(2,4-dimethoxybenzyl)-3-iodo-1H-pyrazolo[4,3-c]pyridine-4-amine (A19, 0.600 g, 1.333 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.316 g, 1.333 mmol), and was obtained as a brown solid (0.43 g, 72% yield). 1H NMR (400MHz, DMSO-d6) δ=7.82 (d, J=6.0Hz, 1H), 7.07-7.19 (m, 3H), 6.83-6.89 (m, 2H), 6.51 (d, J=2.0Hz, 1H), 6.4 0-6.43 (m, 1H), 5.62 (t, J=5.6Hz, 1H), 5.49 (bs, 2H), 4.49 (d, J=5.6Hz, 2H), 3.65-3.72 (m, 7H), 1.08-1.09 (m, 4H). LCMS: 434.2[M+H].

[0569] Intermediate B35

[0570] 3-(6-aminopyridin-3-yl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0571]

[0572] The title compound was obtained by following a similar procedure to that described for intermediate B26, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A1, 0.250 g, 0.830 mmol) and 5-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)pyridine-2-amine (0.183 g, 0.830 mmol), and was obtained as a brown solid (0.085 g, 38% yield). LCMS: 268.6 [M+H].

[0573] Intermediate B36

[0574] 3-(4-amino-3-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidine-4-amine

[0575]

[0576] The title compound was obtained by following a procedure similar to that described for intermediate B26, starting with 3-iodo-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A22, 0.500 g, 1.278 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)aniline (0.454 g, 1.917 mmol), and was obtained as a brown solid (0.16 g, 30% yield). 1H NMR (400MHz, DMSO-d6) δ = 8.26 (s, 1H), 7.20-7.27 (m, 2H), 6.88-6.93 (m, 1H), 5.62 (s, 2H), 5.52 (bs, 2H), 3.62 (t, J=8.0Hz, 2H), 0.84 (t, J=8.0Hz, 2H), -0.12 (s, 9H). LCMS: 375.4[M+H].

[0577] Intermediate B37

[0578] 3-(4-amino-3-fluorophenyl)-1-(1-methylpiperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

[0579]

[0580] The title compound was obtained by following a procedure similar to that described for intermediate B26, starting with 3-iodo-1-(1-methylpiperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A23, 0.180 g, 0.503 mmol) and 2-fluoro-4-(4,4,5,5-tetramethyl-1,3,3-dioxaborhecyclopentan-2-yl)aniline (0.131 g, 0.553 mmol), and was obtained as a brown solid (0.22 g, 72% yield). LCMS: 342.2 [M+H].

[0581] General procedure for synthesizing carbamate intermediate C

[0582] Pyridine (1.2 equivalents) and phenyl chloroformate (1.5 equivalents) were added to a solution of amine (1.0 equivalent) in THF (10 volumes) at 0 °C. The reaction mixture was heated to 25 °C and stirred for 12 hours. After the reaction was complete (as shown by TLC), the mixture was diluted with EtOAc (10 mL) and washed with brine (5 mL). The organic layer was dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain a crude material, which was purified by rapid chromatography (silica gel 230-400 mesh, with 10% to 20% EtOAc / petroleum ether) to obtain the desired carbamate.

[0583] The following carbamates were prepared using the general procedure described above.

[0584]

[0585]

[0586]

[0587]

[0588] Note: The amines used to synthesize carbamates are commercially available or synthesized using the following literature procedures:

[0589] The synthesis of 3-(1-(trifluoromethyl)cyclopropyl)isoxazole-5-amine (C6 precursor) and 5-(1-(trifluoromethyl)cyclopropyl)isoxazole-3-amine (C7 precursor) from methyl 1-(trifluoromethyl)cyclopropane-1-carboxylate is reported in Synthesis 2013, 45, 171-173.

[0590] 3-(1,1,1-trifluoro-2-methylpropane-2-yl)isoxazole-5-amine (C8 precursor) and 3-(2-fluoropropane-2-yl)isoxazole-5-amine (C9 precursor) were synthesized from methyl 3,3,3-trifluoro-2,2-dimethylpropionate and methyl 2-fluoro-2-methylpropionate, respectively, followed by the procedure reported in Synthesis 2013, 45, 171-173.

[0591] The synthesis of 2-(5-aminoisoxazol-3-yl)-2-methylpropionitrile (a C20 precursor) from 2,2-dimethyl-3-oxopyranonitrile is reported in J. Med. Chem. 2012, 55, 1082-1105.

[0592] The synthesis of 3-(((tert-butyldiphenylsilyl)oxy)methyl)isoxazole-5-amine (a precursor of C21) from 4-((tert-butyldiphenylsilyl)oxy)-3-oxobutyronitrile is reported in PCT Publication No. WO2013 / 104561.

[0593] The synthesis of 3-(tert-butyl)-4-methylisoxazol-5-amine (a precursor of C22) from 2,4,4-trimethyl-3-oxopentanilide is reported in PCT Publication No. WO 2012 / 019015.

[0594] Synthesis of 3-(1-methylcyclobutyl)isoxazole-5-amine (C19 precursor):

[0595]

[0596] NH₂OH·H₂SO₄ (0.699 g, 4.25 mmol) was added to a stirred solution of 3-(1-methylcyclobutyl)-3-oxopropionitrile (prepared as reported in PCT Publication No. WO 2017 / 060874, 0.500 g, 3.86 mmol) and sodium hydroxide (0.170 g, 4.25 mmol) in EtOH (10 mL) and water (10 mL). The pH of the resulting mixture was adjusted to 7.5 using an aqueous solution of NaOH (1 M), and the reaction mixture was stirred at 80 °C for 15 h. After the reaction was complete (as shown by TLC), the reaction mixture was concentrated under reduced pressure to obtain a residue, which was dissolved in DCM (25 mL), washed with water (10 mL), dried over Na₂SO₄, filtered, and concentrated under reduced pressure. The crude material was purified by rapid chromatography (silica gel 230-400 mesh, eluted with 40% EtOAc / petroleum ether) to give the title product (0.110 g, 19% yield) as a grayish-white solid. 1 H NMR (400MHz, CDCl3) δ=5.04 (s, 1H), 2.43-2.49 (m, 2H), 1.96-2.02 (m, 4H), 1.50 (s, 3H). LCMS: 153.2[M+H].

[0597] Preparation of Examples

[0598] General urea formation procedures used to synthesize Examples 1 to 72

[0599] Method A – Triethylamine (2.0 equivalents) was added to a mixture of amine intermediate D (1.0 equivalents) and carbamate intermediate E (1.0 equivalents) in THF (10 volumes), and the resulting mixture was stirred in a sealed tube at 60°C for 12 hours. After the reaction was complete (as shown in LCMS), the reaction mixture was concentrated under reduced pressure to obtain a crude material, which was purified by reversed-phase preparative HPLC to obtain the desired product.

[0600] Method B – DMAP (0.05 equivalents) and DIPEA (1.5 equivalents) were added to a solution of amine intermediate D (1.0 equivalents) and carbamate intermediate E (1.0 equivalents) in THF (10 volumes), and the resulting mixture was stirred in a sealed tube at 60°C for 12 hours. After the reaction was complete (as shown in LCMS), the reaction mixture was concentrated under reduced pressure to obtain a crude material, which was purified by reversed-phase preparative HPLC to obtain the desired product.

[0601] The following compounds were prepared using the general procedure described above.

[0602] Example 1

[0603] 1-(4-(4-amino-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0604]

[0605] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-aminophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B2, 0.250 g, 0.93 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.242 g, 0.93 mmol), and was obtained as a grayish-white solid (0.150 g, 37% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.15 (bs, 1H), 9.09 (bs, 1H), 8.24 (bs, 1H), 7.55-7. 66 (m, 4H), 6.10 (s, 1H), 5.03-5.10 (m, 1H), 1.49 (d, J=6.4Hz, 6H), 1.27 (s, 9H). LCMS: 435.3[M+H].

[0606] Example 2

[0607] 1-(4-(4-amino-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0608]

[0609] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-aminophenyl)-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B4, 0.210 g, 0.744 mmol) and (3-(tert-butyl)isoxazol-3-yl)carbamate (C4, 0.193 g, 0.744 mmol), and was obtained as a grayish-white solid (0.053 g, 16% yield). 1 HNMR (400MHz, DMSO-d6) δ = 10.23 (bs, 1H), 9.15 (bs, 1H), 8.31 (s, 1H), 7.74 (s, 4H), 6.1 6 (s, 1H), 6.06-6.10 (m, 1H), 5.16 (t, J=6.4Hz, 2H), 5.05 (t, J=6.8Hz, 2H), 1.33 (s, 9H). LCMS: 449.2[M+H].

[0610] Example 3

[0611] 1-(4-(4-amino-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(5-(tert-butyl)isoxazo-3-yl)urea

[0612]

[0613] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-aminophenyl)-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B4, 0.070 g, 0.248 mmol) and (5-(tert-butyl)isoxazol-3-yl)carbamate (C5, 0.065 g, 0.248 mmol), and was obtained as a grayish-white solid (0.035 g, 32% yield). 1 HNMR (400MHz, DMSO-d6) δ = 9.59 (bs, 1H), 9.07 (bs, 1H), 8.26 (s, 1H), 7.66 (bs, 4H), 6.5 3 (s, 1H), 5.98-6.04 (m, 1H), 5.09 (t, J=6.4Hz, 2H), 5.00 (t, J=6.8Hz, 2H), 1.31 (s, 9H). LCMS: 449.2[M+H].

[0614] Example 4

[0615] 1-(4-(4-amino-1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0616]

[0617] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-aminophenyl)-1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B12, 0.05 g, 0.16 mmol) and (3-(tert-butyl)isoxazol-3-yl)carbamate (C4, 0.042 g, 0.16 mmol), and was obtained as a grayish-white solid (0.013 g, 20% yield). 1HNMR (400MHz, DMSO-d6) δ = 10.13 (bs, 1H), 9.08 (bs, 1H), 8.25 (s, 1H), 7.61-7.67 (m, 4H), 6.10 (s, 1H), 4.9 0-4.98 (m, 1H), 4.00-4.03 (m, 2H), 3.53-3.58 (m, 2H), 2.19-2.26 (m, 2H), 1.87-1.91 (m, 2H), 1.27 (s, 9H). LCMS: 477.1[M+H].

[0618] Example 5

[0619] 1-(4-(4-amino-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0620]

[0621] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-aminophenyl)-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B10, 0.083 g, 0.28 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.072 g, 0.28 mmol), and was obtained as a white solid (0.048 g, 36% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.17 (bs, 1H), 9.08 (bs, 1H), 8.26 (s, 1H), 7.60-7.67 (m, 4H), 6.10 (s , 1H), 5.49-5.52(m, 1H), 4.07-4.14(m, 2H), 3.89-3.97(m, 2H), 2.38-2.43(m, 2H), 1.27(s, 9H). LCMS: 463.0[M+H].

[0622] Example 6

[0623] 1-(4-(4-amino-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-chlorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0624]

[0625] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-chlorophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B21, 0.080 g, 0.264 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.067 g, 0.264 mmol), and was obtained as a grayish-white solid (0.003 g, 3% yield). 1 H NMR (400MHz, DMSO-6) δ = 10.85 (bs, 1H), 8.66 (bs, 1H), 8.35 (d, J = 8.4Hz, 1H), 8.25 (s, 1H), 7.72 (d, J =2.0Hz, 1H), 7.61-7.64 (m, 1H), 6.12 (s, 1H), 5.04-5.10 (m, 1H), 1.50 (d, J = 6.8Hz, 6H), 1.28 (s, 9H). LCMS: 469.1[M+H].

[0626] Example 7

[0627] 1-(4-(4-amino-1-cyclobutyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0628]

[0629] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-aminophenyl)-1-cyclobutyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B8, 0.140 g, 0.49 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.130 g, 0.49 mmol), and was obtained as a grayish-white solid (0.067 g, 29% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.15 (bs, 1H), 9.09 (bs, 1H), 8.24 (s, 1H), 7.62-7.68 (m, 4H), 6.10 (s , 1H), 5.32-5.36(m, 1H), 2.69-2.74(m, 2H), 2.39-2.42(m, 2H), 1.87-1.89(m, 2H), 1.27(s, 9H). LCMS: 447.2[M+H].

[0630] Example 8

[0631] 1-(4-(4-amino-1-cyclobutyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0632]

[0633] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-aminophenyl)-1-cyclobutyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B9, 0.100 g, 0.33 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.087 g, 0.33 mmol), and was obtained as a pale yellow solid (0.005 g, 4% yield). 1 H NMR (400MHz, DMSO-d6) δ = 9.10 (bs, 1H), 8.23-8.28 (m, 2H), 7.47-7.53 (m, 2H), 6.90 (bs, 1H), 6.09 (s, 1H), 5.29-5.37 (m, 1H), 2.67-2.73 (m, 2H), 2.38-2.41 (m, 2H), 1.85-1.89 (m, 2H), 1.26 (s, 9H). LCMS: 465.1[M+H].

[0634] Example 9

[0635] 1-(4-(4-amino-1-(2-hydroxy-2-methylpropyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0636]

[0637] The title compound was prepared following the general procedure for urea formation (Method A) starting from 1-(4-amino-3-(4-aminophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol (B18, 0.119 g, 0.4 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.103 g, 0.4 mmol), and was obtained as a grayish-white solid (0.017 g, 9% yield). 1 H NMR (400MHz, DMSO-d6) δ = 9.44 (bs, 1H), 8.24 (s, 1H), 7.59-7.68 (m, 4H), 6.07 (s, 1H), 4.81 (bs, 1H), 4.27 (bs, 2H), 1.26 (s, 9H), 1.15 (s, 6H). LCMS: 465.1[M+H].

[0638] Example 10

[0639] 1-(4-(4-amino-1-(2-hydroxy-2-methylpropyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0640]

[0641] The title compound was prepared following the general procedure for urea formation (Method A) starting from 1-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol (B19, 0.077 g, 0.243 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.063 g, 0.243 mmol), and was obtained as a grayish-white solid (0.013 g, 11% yield). 1 H NMR (400MHz, CD3OD) δ = 8.33-8.37 (m, 1H), 8.29 (s, 1H), 7.53-7.59 (m, 2H), 6.19 (s, 1H), 4.42 (s, 2H), 1.35 (s, 9H), 1.29 (s, 6H). LCMS: 483.3[M+H].

[0642] Example 11

[0643] 1-(4-(4-amino-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazol-5-yl)urea

[0644]

[0645] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-isopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B3, 0.200 g, 0.69 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.182 g, 0.69 mmol), and was obtained as a grayish-white solid (0.011 g, 3% yield). 1H NMR (400MHz, DMSO-d6) δ = 10.40 (bs, 1H), 8.93 (bs, 1H), 8.24-8.30 (m, 2H), 7.47-7.52 (m , 2H), 6.95 (bs, 2H), 6.11 (s, 1H), 5.03-5.10 (m, 1H), 1.49 (d, J=6.4Hz, 6H), 1.27 (s, 9H). LCMS: 453.2[M+H].

[0646] Example 12

[0647] 1-(4-(4-amino-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0648]

[0649] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-(oxetane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B5, 0.200 g, 0.66 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.173 g, 0.66 mmol), and was obtained as a grayish-white solid (0.027 g, 10% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.97 (bs, 1H), 8.29-8.33 (m, 1H), 8.25 (s, 1H), 7.52-7.59 (m, 2 H), 6.11(s, 1H), 5.98-6.05(m, 1H), 5.01-5.11(m, 2H), 4.98-4.99(m, 2H), 1.27(s, 9H). LCMS: 467.1[M+H].

[0650] Example 13

[0651] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0652]

[0653] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.150 g, 0.52 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.137 g, 0.52 mmol), and was obtained as a white solid (0.035 g, 15% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.42 (bs, 1H), 8.98 (bs, 1H), 8.25-8.29 (m, 2H), 7.44-7.50 (m, 2H), 6. 95(bs, 2H), 6.10(s, 1H), 3.85-3.87(m, 1H), 1.27(s, 9H), 1.20-1.23(m, 2H), 1.08-1.10(m, 2H). LCMS: 451.2[M+H].

[0654] Example 14

[0655] 1-(4-(4-amino-1-(1-(oxecyclobutane-3-yl)piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0656]

[0657] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-(1-(oxecyclobutan-3-yl)piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B16, 0.050 g, 0.13 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.034 g, 0.13 mmol), and was obtained as a grayish-white solid (0.006 g, 8% yield). 1 H NMR (400MHz, CD3OD) δ = 8.32-8.36 (m, 1H), 8.27 (s, 1H), 7.51-7.57 (m, 2H), 6.19 (s, 1H), 4.65-4.82 (m, 5H), 3 .61-3.64(m, 1H), 2.98-3.01(m, 2H), 2.37-2.44(m, 2H), 2.13-2.19(m, 2H), 2.04-2.07(m, 2H), 1.35(s, 9H). LCMS: 550.0[M+H].

[0658] Example 15

[0659] 1-(4-(4-amino-1-(4-hydroxycyclohexyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0660]

[0661] The title compound was prepared following the general procedure for urea formation (Method A) starting with 4-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)cyclohexane-1-ol (B15, 0.075 g, 0.22 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.057 g, 0.22 mmol), and was obtained as a grayish-white solid (0.007 g, 6% yield). 1 H NMR (400MHz, CD3OD) δ = 8.32-8.36 (m, 1H), 8.27 (s, 1H), 7.50-7.55 (m, 2H), 6.19 (s, 1H), 4.74-4.80 (m, 1H), 3.72-3.73 (m, 1H), 2.14-2.21 (m, 4H), 2.03-2.06 (m, 2H), 1.55-1.59 (m, 2H), 1.35 (s, 9H). LCMS: 509.2[M+H].

[0662] Example 16

[0663] 1-(4-(4-amino-1-(3,3-difluorocyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0664]

[0665] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-(3,3-difluorocyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B14, 0.110 g, 0.33 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.086 g, 0.33 mmol), and was obtained as a grayish-white solid (0.020 g, 13% yield). 1H NMR (400MHz, DMSO-d6) δ = 9.28 (bs, 1H), 8.26-8.30 (m, 2H), 7.48-7.55 (m, 2H), 7.08(bs, 1H), 6.07(s, 1H), 5.28-5.33(m, 1H), 3.25-3.34(m, 4H), 1.27(s, 9H). LCMS: 501.1[M+H].

[0666] Example 17

[0667] 1-(4-(4-amino-1-(pyridin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0668]

[0669] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-(pyridin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B20, 0.140 g, 0.43 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.113 g, 0.43 mmol), and was obtained as a grayish-white solid (0.006 g, 3% yield). 1 HNMR (400MHz, DMSO-d6) δ = 10.44 (bs, 1H), 9.00 (bs, 1H), 8.73-8.74 (m, 2H), 8.34-8.47 (m, 4H), 7.59-7.68 (m, 2H), 6.13 (s, 1H), 1.28 (s, 9H). LCMS: 488.2[M+H].

[0670] Example 18

[0671] 1-(4-(4-amino-1-(1,1-tetrahydrothiophene-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazol-5-yl)urea

[0672]

[0673] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)tetrahydrothiophene 1,1-dioxide (B17, 0.030 g, 0.082 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.021 g, 0.082 mmol), and was obtained as a white solid (0.003 g, 7% yield). 1 H NMR (400MHz, CD3OD) δ = 8.31-8.36 (m, 2H), 7.54-7.60 (m, 2H), 6.19 (s, 1H), 5.78-5.80 (m, 1H), 3.58-3.63 (m, 3H), 2.78-2.89 (m, 2H), 1.32 (s, 9H). LCMS: 529.2[M+H].

[0674] Example 19

[0675] 1-(4-(4-amino-1-(tetrahydro-2H-pyran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0676]

[0677] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-(tetrahydro-2H-pyran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B13, 0.015 g, 0.045 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.011 g, 0.045 mmol), and was obtained as a grayish-white solid (0.002 g, 9% yield). 1 H NMR (400MHz, CD3OD) δ = 8.54 (s, 1H), 8.35 (d, J = 7.2Hz, 1H), 8.28 (s, 1H), 7.50-7.56 (m, 2H), 6.19 (s, 1H), 4.60 (s, 1H), 3.87-4.05(m, 3H), 3.50-3.58(m, 1H), 2.38-2.42(m, 1H), 2.21-2.23(m, 1H), 1.92-2.03(m, 2H), 1.35(s, 9H). LCMS: 495.1[M+H].

[0678] Example 20

[0679] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0680]

[0681] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-aminophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B7, 0.100 g, 0.38 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.100 g, 0.38 mmol), and was obtained as a white solid (0.045 g, 27% yield). 1 HNMR (400MHz, DMSO-d6) δ = 10.11 (bs, 1H), 9.10 (bs, 1H), 8.26 (s, 1H), 7.58-7.65 (m, 4H ), 6.09 (s, 1H), 3.83-3.89 (m, 1H), 1.24 (s, 9H), 1.19-1.23 (m, 2H), 1.06-1.11 (m, 2H). LCMS: 433.2[M+H].

[0682] Example 21

[0683] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0684]

[0685] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.100 g, 0.31 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.091 g, 0.31 mmol), and was obtained as a grayish-white solid (0.020 g, 12% yield). 1H NMR (400MHz, DMSO-d6) δ = 10.63 (bs, 1H), 8.92 (bs, 1H), 8.25-8.29 (m, 2H), 7.44-7.51 (m, 2H), 6.87 (bs, 2H), 6.20 (s, 1H), 3.84-3.89 (m, 1H), 1.45-1.76 (m, 2H), 1.36-1.41 (m, 2H), 1.19-1.23 (m, 2H), 1.06-1.11 (m, 2H). LCMS: 503.1[M+H].

[0686] Example 22

[0687] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1,1,1-trifluoro-2-methylpropane-2-yl)isoxazo-5-yl)urea

[0688]

[0689] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.17 mmol) and (3-(1,1,1-trifluoro-2-methylpropane-2-yl)isoxazol-5-yl)carbamate (C8, 0.045 g, 0.17 mmol), and was obtained as a grayish-white solid (0.004 g, 4% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.62 (bs, 1H), 8.96 (bs, 1H), 8.25-8.28 (m, 2H), 7.45-7.52 (m, 2H), 6. 99(bs, 2H), 6.25(s, 1H), 3.84-3.90(m, 1H), 1.53(s, 6H), 1.19-1.23(m, 2H), 1.10-1.12(m, 2H). LCMS: 505.1[M+H].

[0690] Example 23

[0691] 1-(4-(4-amino-1-(oxecyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(tert-butyl)isoxazo-3-yl)urea

[0692]

[0693] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-(oxetane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B5, 0.150 g, 0.5 mmol) and (5-(tert-butyl)isoxazol-3-yl)carbamate (C5, 0.130 g, 0.5 mmol), and was obtained as a grayish-white solid (0.015 g, 6% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.00 (bs, 1H), 9.09 (bs, 1H), 8.32-8.37 (m, 1H), 8.26 (s, 1H), 7.51-7.59 (m, 2H) , 6.80 (bs, 2H), 6.53 (s, 1H), 5.99-6.04 (m, 1H), 5.10 (t, J=6.4Hz, 2H), 5.00 (t, J=6.4Hz, 2H), 1.31 (s, 9H). LCMS: 467.2[M+H].

[0694] Example 24

[0695] 1-(4-(4-amino-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0696]

[0697] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-(tetrahydrofuran-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B11, 0.200 g, 0.59 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.154 g, 0.59 mmol), and was obtained as a white solid (0.030 g, 10% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.40 (bs, 1H), 8.92 (bs, 1H), 8.26-8.31 (m, 2H), 7.47-7.54 (m, 2H), 6.80 (bs, 2H) ), 6.11(s, 1H), 5.47-5.53(m, 1H), 4.05-4.13(m, 2H), 3.87-3.96(m, 2H), 2.40-2.41(m, 2H), 1.27(s, 9H). LCMS: 481.2[M+H].

[0698] Example 25

[0699] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0700]

[0701] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.070 g, 0.24 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.077 g, 0.24 mmol), and was obtained as a grayish-white solid (0.013 g, 11% yield). 1 H NMR (400MHz, DMSO-d6) δ = 11.78 (bs, 1H), 8.76 (s, 1H), 7.43-7.45 (m, 2H), 6.84-6.88 (m, 2H) , 5.46 (s, 2H), 3.91-3.97 (m, 1H), 1.49-1.54 (m, 4H), 1.19-1.22 (m, 2H), 1.10-1.15 (m, 2H). LCMS: 503.1[M+H].

[0702] Example 26

[0703] 1-(4-(1-allyl-4-amino-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(tert-butyl)isoxazo-3-yl)urea

[0704]

[0705] The title compound was prepared following the general procedure for urea formation (Method A) starting from 1-allyl-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B6, 0.130 g, 0.45 mmol) and (5-(tert-butyl)isoxazol-3-yl)carbamate (C5, 0.119 g, 0.45 mmol), and was obtained as a grayish-white solid (0.012 g, 6% yield). 1H NMR (400MHz, DMSO-d6) δ = 10.46 (bs, 1H), 9.52 (bs, 1H), 8.24-8.28 (m, 2H), 7.45-7.50 (m, 2H), 6.9 5 (bs, 2H), 6.52 (s, 1H), 6.00-6.10 (m, 1H), 5.18-5.21 (m, 2H), 5.13 (d, J=1.6Hz, 2H), 1.31 (s, 9H). LCMS: 451.2[M+H].

[0706] Example 27

[0707] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(2-fluoropropane-2-yl)isoxazo-5-yl)urea

[0708]

[0709] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (3-(2-fluoropropane-2-yl)isoxazol-5-yl)carbamate (C9, 0.047 g, 0.176 mmol), and was obtained as a grayish-white solid (0.007 mg, 9% yield). 1 H NMR (400MHz, DMSO-d6) δ = 8.32-8.35 (m, 1H), 8.25 (s, 1H), 7.32-7.37 (m, 2H), 5.92 (s, 1 H), 3.83-3.89 (m, 1H), 1.66 (s, 3H), 1.61 (s, 3H), 1.20-1.24 (m, 2H), 1.07-1.09 (m, 2H). LCMS: 455.1[M+H].

[0710] Example 28

[0711] 1-(4-(4-amino-1-methyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-chlorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0712]

[0713] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-chlorophenyl)-1-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B22, 0.068 g, 0.248 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.064 g, 0.248 mmol), and was obtained as a grayish-white solid (0.005 g, 4% yield). 1 H NMR (400MHz, CD3OD) δ = 8.70-8.68 (m, 1H), 7.47-7.67 (m, 2H), 7.00-7.02 (m, 1H), 6.20 (s, 1H), 4.10 (s, 3H), 1.34 (s, 9H). LCMS: 441.1[M+H].

[0714] Example 29

[0715] 1-(4-(4-amino-1-methyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0716]

[0717] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B23, 0.052 g, 0.201 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.052 g, 0.201 mmol), and was obtained as a grayish-white solid (0.005 g, 6% yield). 1 H NMR (400MHz, CD3OD) δ=10.60 (bs, 1H), 9.05 (bs, 1H), 8.25-8.30 (m, 2H), 7.45-7.51 (m, 2H), 6.90 (s, 2H), 6.09 (s, 1H), 3.95 (s, 3H), 1.27 (s, 9H). LCMS: 423.1[MH].

[0718] Example 30

[0719] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0720]

[0721] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridine-4-amine (B24, 0.070 g, 0.24 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.064 g, 0.24 mmol), and was obtained as a grayish-white solid (0.004 g, 4% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.52 (bs, 1H), 9.09 (bs, 1H), 8.23-8.27 (m, 1H), 7.80 (s, 1H), 7.44-7.53 (m, 2H ), 6.90 (d, J=6.0Hz, 1H), 6.10 (s, 1H), 5.86 (bs, 2H), 3.72-3.74 (m, 1H), 1.27 (s, 9H), 1.12-1.13 (m, 4H). LCMS: 450.2[M+H].

[0722] Example 31

[0723] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridin-3-yl)phenyl)-3-(3-(tert-butyl)isoxazo-5-yl)urea

[0724]

[0725] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-aminophenyl)-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridine-4-amine (B25, 0.052 g, 0.2 mmol) and (3-(tert-butyl)isoxazol-5-yl)carbamate (C4, 0.05 g, 0.2 mmol), and was obtained as a white solid (0.007 g, 8% yield). 1 HNMR (400MHz, DMSO-d6) δ = 10.85 (bs, 1H), 9.68 (bs, 1H), 7.78 (bs, 1H), 7.66 (d, J = 8.4Hz, 2H), 7.56 (d, J = 8.4H z, 2H), 6.90 (d, J=6.0Hz, 1H), 6.08 (s, 1H), 5.75 (bs, 2H), 3.68-3.73 (m, 1H), 1.26 (s, 9H), 1.11-1.12 (m, 4H). LCMS: 432.3[M+H].

[0726] Example 32

[0727] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-methylcyclopropyl)isoxazol-5-yl)

[0728]

[0729] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (3-(1-methylcyclopropyl)isoxazol-5-yl)carbamate (C10, 0.045 g, 0.176 mmol), and was obtained as a grayish-white solid (0.009 g, 11% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.29 (bs, 1H), 8.90 (bs, 1H), 8.24-8.28 (m, 2H), 7.44-7.51 (m, 2H), 6.94 (bs, 2H), 5.86 (s , 1H), 3.84-3.89(m, 1H), 1.38(s, 3H), 1.19-1.22(m, 2H), 1.08-1.11(m, 2H), 0.94-0.97(m, 2H), 0.82-0.85(m, 2H). LCMS: 449.1[M+H].

[0730] Example 33

[0731] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(tert-butyl)-1,3,4-thiadiazol-2-yl)

[0732]

[0733] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (5-(tert-butyl)-1,3,4-thiadiazol-2-yl)carbamate (C11, 0.049 g, 0.176 mmol), and was obtained as a grayish-white solid (0.012 g, 14% yield). 1H NMR (400MHz, DMSO-d6) δ = 8.67 (bs, 1H), 8.33-8.41 (m, 1H), 8.25 (s, 1H), 7.34-7.39 (m, 2 H), 6.87 (bs, 2H), 3.83-3.87 (m, 1H), 1.34 (s, 9H), 1.20-1.23 (m, 2H), 1.06-1.11 (m, 2H). LCMS: 468.2[M+H].

[0734] Example 34

[0735] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(4-(tert-butyl)thiazolyl-2-yl)urea

[0736]

[0737] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (4-(tert-butyl)thiazolyl-2-yl)carbamate (C12, 0.049 g, 0.176 mmol), and was obtained as a grayish-white solid (0.004 g, 5% yield). 1 H NMR (400MHz, CD3OD) δ = 8.45-8.49 (m, 1H), 8.40 (s, 1H), 7.50-7.58 (m, 2H), 6.65 (s, 1H), 3.96-4.02 (m, 1H), 1.31-1.40 (m, 11H), 1.21-1.25 (m, 2H). LCMS: 467.2[M+H].

[0738] Example 35

[0739] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)isothiazolyl-5-yl)urea

[0740]

[0741] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (3-(tert-butyl)isothiazolyl-5-yl)carbamate (C13, 0.049 g, 0.176 mmol), and was obtained as a grayish-white solid (0.016 g, 19% yield). 1 H NMR (400MHz, DMSO-d6) δ = 11.42 (bs, 1H), 8.51 (bs, 1H), 8.26 (s, 1H), 8.07-8.11 (m, 1H), 7 .39-7.43 (m, 2H), 6.70 (s, 1H), 3.84-3.87 (m, 1H), 1.21-1.26 (m, 11H), 1.08-1.10 (m, 2H). LCMS: 465.1[MH].

[0742] Example 36

[0743] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(tert-butyl)-1,3,4-oxadiazol-2-yl)urea

[0744]

[0745] The title compound was prepared following the general procedure for urea formation (Method A) starting with (4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)carbamate (C3, 0.050 g, 0.124 mmol) and 5-(tert-butyl)-1,3,4-oxadiazol-2-amine (0.017 g, 0.124 mmol), and was obtained as a grayish-white solid (0.002 g, 4% yield). 1 H NMR (400MHz, CD3OD) δ = 8.37-8.43 (m, 2H), 7.46-7.62 (m, 2H), 3.93-3.95 (m, 1H), 1.45 (s, 9H), 1.31-1.38 (m, 2H), 1.20-1.23 (m, 2H). LCMS: 451.9[M+H].

[0746] Example 37

[0747] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)-1,2,4-thiadiazol-5-yl)urea

[0748]

[0749] The title compound was prepared following the general procedure for urea formation (Method A) starting with (4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)carbamate (C3, 0.050 g, 0.124 mmol) and 3-(tert-butyl)-1,2,4-thiadiazole-5-amine (0.019 g, 0.124 mmol), and was obtained as a grayish-white solid (0.001 g, 1% yield). 1 H NMR (400MHz, CD3OD) δ = 8.27-8.39 (m, 2H), 7.51-7.56 (m, 2H), 3.80-3.86 (m, 1H), 1.40 (s, 9H), 1.31-1.33 (m, 2H), 1.17-1.21 (m, 2H). LCMS: 468.0[M+H].

[0750] Example 38

[0751] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(1-(tert-butyl)-1H-1,2,4-triazol-3-yl)urea

[0752]

[0753] The title compound was prepared following the general procedure for urea formation (Method A) starting from (4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)carbamate (C3, 0.050 g, 0.124 mmol) and 1-(tert-butyl)-1H-1,2,4-triazol-3-amine (0.017 g, 0.124 mmol), and was obtained as a grayish-white solid (0.009 g, 15% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.80 (bs, 1H), 10.21 (bs, 1H), 8.55 (s, 1H), 8.42-8.46 (m, 1H), 8.26 (s, 1H), 7 .44-7.52 (m, 2H), 6.94 (bs, 2H), 3.83-3.88 (m, 1H), 1.57 (s, 9H), 1.19-1.23 (m, 2H), 1.06-1.11 (m, 2H). LCMS: 451.0[M+H].

[0754] Example 39

[0755] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(trifluoromethyl)isoxazo-5-yl)urea

[0756]

[0757] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (3-(trifluoromethyl)isoxazol-5-yl)carbamate (C14, 0.048 g, 0.176 mmol), and was obtained as a white solid (0.004 g, 5% yield). 1 H NMR (400MHz, DMSO-d6) δ = 11.11 (bs, 1H), 8.97 (bs, 1H), 8.23-8.27 (m, 2H), 7.46-7.52 (m, 2H), 6.95(bs, 2H), 6.53(s, 1H), 3.84-3.88(m, 1H), 1.19-1.24(m, 2H), 1.07-1.12(m, 2H). LCMS: 462.9[M+H].

[0758] Example 40

[0759] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(pentan-3-yl)isoxazol-5-yl)urea

[0760]

[0761] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.05 g, 0.176 mmol) and (3-(pentan-3-yl)isoxazol-5-yl)carbamate (C15, 0.048 g, 0.176 mmol), and was obtained as a pale brown solid (0.006 g, 7% yield). 1H NMR (400MHz, DMSO-d6) δ = 10.44 (bs, 1H), 8.97 (bs, 1H), 8.26-8.29 (m, 2H), 7.44-7.50 (m, 2H), 6.81 (bs, 2H), 6.00 (s, 1H), 3.84-3.86 (m, 1H), 1.51-1.66 (m, 4H), 1.08-1.21 (m, 4H), 0.79-0.82 (m, 6H). LCMS: 465.0[M+H].

[0762] Example 41

[0763] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-isopropylisoxazol-5-yl)urea

[0764]

[0765] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (3-isopropylisoxazol-5-yl)carbamate (C16, 0.043 g, 0.176 mmol), and was obtained as a grayish-white solid (0.007 g, 9% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.49 (bs, 1H), 9.01 (bs, 1H), 8.27-8.32 (m, 2H), 7.44-7.52 (m, 2H ), 6.07 (s, 1H), 3.87-3.92 (m, 1H), 2.90-2.97 (m, 1H), 1.18-1.24 (m, 8H), 1.10-1.12 (m, 2H). LCMS: 437.0[M+H].

[0766] Example 42

[0767] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-ethylisoxazol-5-yl)urea

[0768]

[0769] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.050 g, 0.176 mmol) and (3-ethylisoxazol-5-yl)carbamate (C17, 0.041 g, 0.176 mmol), and was obtained as a grayish-white solid (0.006 g, 8% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.44 (bs, 1H), 8.94 (bs, 1H), 8.26-8.30 (m, 2H), 7.44-7.51 (m, 2H), 6.99 (bs, 2H), 6.05 (s, 1H), 3.84-3.89 (m, 1H), 2.53-2.59 (m, 2H), 1.18-1.21 (m, 5H), 1.08-1.10 (m, 2H). LCMS: 423.0[M+H].

[0770] Example 43

[0771] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(sec-butyl)isoxazo-5-yl)urea

[0772]

[0773] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.080 g, 0.281 mmol) and (3-(sec-butyl)isoxazol-5-yl)carbamate (C18, 0.073 g, 0.281 mmol), and was obtained as a white solid (0.004 g, 4% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.42 (bs, 1H), 8.96 (bs, 1H), 8.26-8.33 (m, 2H), 7.44-7.52 (m, 2H), 6.04 (s , 1H), 3.87-3.92(m, 1H), 2.68-2.73(m, 1H), 1.54-1.60(m, 2H), 1.08-1.24(m, 7H), 0.82-0.84(m, 3H). LCMS: 451.1[M+H].

[0774] Example 44

[0775] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-methylcyclobutyl)isoxazo-5-yl)urea

[0776]

[0777] The title compound was prepared following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.100 g, 0.352 mmol) and (3-(1-methylcyclobutyl)isoxazol-5-yl)carbamate (C19, 0.096 g, 0.352 mmol), and was obtained as a white solid (0.002 g, 1% yield). 1 HNMR (400MHz, CD3OD) δ = 8.35-8.41 (m, 2H), 7.49-7.56 (m, 2H), 6.16 (s, 1H), 3.98-4 .02(m, 1H), 2.46-2.51(m, 2H), 1.96-2.13(m, 4H), 1.54(s, 3H), 1.23-1.38(m, 4H). LCMS: 463.1[M+H].

[0778] Example 45

[0779] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0780]

[0781] The title compound was prepared following the general procedure for urea formation (Method A) starting from 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridine-4-amine (B24, 0.160 g, 0.565 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.176 g, 0.565 mmol), and was obtained as a grayish-white solid (0.012 g, 4% yield). 1H NMR (400MHz, DMSO-d6) δ = 10.69 (bs, 1H), 8.97 (bs, 1H), 8.23-8.27 (m, 1H), 7.80 (d, J = 6.0Hz, 1H), 7.46-7.54 (m, 2H), 6.91 (d, J=6.0Hz, 1H), 6.22 (s, 1H), 5.85 (bs, 2H), 3.72-3.74 (m, 1H), 1.39-1.49 (m, 4H), 1.12-1.13 (m, 4H). LCMS: 502.2[M+H].

[0782] Example 46

[0783] 1-(4-(4-amino-1-(3-hydroxycyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0784] Step 1: Synthesis of 1-(4-(4-amino-1-(3-(benzyloxy)cyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0785]

[0786] The title compound was obtained following the general procedure for urea formation (Method B), starting with 3-(4-amino-3-fluorophenyl)-1-(3-(benzyloxy)cyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B26, 0.090 g, 0.223 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.076 g, 0.245 mmol), and was obtained as a brown gel (70 mg), which was used without further purification. LCMS: 623.4 [M+H].

[0787] Step 2: Synthesis of 1-(4-(4-amino-1-(3-hydroxycyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0788]

[0789] Boron trichloride (1 M in DCM, 0.899 mL, 0.899 mmol) was added dropwise to a solution of 1-(4-(4-amino-1-(3-(benzyloxy)cyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)urea (0.070 g, 0.112 mmol) in anhydrous DCM (5 mL) at -60 °C, and the resulting mixture was stirred at 0 °C for 3 h. After the reaction was complete (as shown by TLC and LCMS), the reaction mixture was cooled to -70 °C, neutralized with ammonia (25% in water), and extracted with DCM (10 mL × 2). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain a crude material, which was purified by preparative HPLC (eluting with 1% TFA / (water and CAN)) to give the title product as a white solid (0.009 g, 15% yield). 1 H NMR (400MHz, CD3OD) δ = 8.34-8.38 (m, 2H), 7.53-7.61 (m, 2H), 6.34 (s, 1H), 5.61-5.64 (m, 1H), 4.73-4.79 (m, 1H), 2.94-3.01 (m, 2H), 2.57-2.63 (m, 2H), 1.39-1.48 (m, 4H). LCMS: 533.7[M+H].

[0790] Example 47

[0791] 1-(5-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)

[0792] Step 1: Synthesis of 1-(5-(4,4,5,5-tetramethyl-1,3,2-dioxaborphane-2-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0793]

[0794] The title compound was obtained by following the general procedure for urea formation (Method A) starting with 5-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)pyridine-2-amine (0.100 g, 0.454 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.142 g, 0.454 mmol), and was obtained as a brown gel (150 mg) used without further purification. LCMS: 439.5 [M+H].

[0795] Step 2: Synthesis of 1-(5-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0796]

[0797] The title compound was obtained by following a procedure similar to that described for intermediate B26, starting with 1-cyclopropyl-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A1, 0.100 g, 0.332 mmol) and 1-(5-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)urea (0.146 g, 0.332 mmol), and was obtained as a grayish-white solid (0.013 g, 8% yield). 1 H NMR (400MHz, DMSO-d6) δ = 11.68 (bs, 1H), 9.93 (bs, 1H), 8.53 (d, J = 2.0Hz, 1H), 8.28 (s, 1H), 8 .02-8.05 (m, 1H), 7.72 (d, J=8.4Hz, 1H), 6.27 (s, 1H), 3.86-3.90 (m, 1H), 1.10-1.49 (m, 8H). LCMS: 486.2[M+H].

[0798] Example 48

[0799] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methylphenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0800]

[0801] The title compound was obtained by starting with 3-(4-amino-3-methylphenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B27, 0.110 g, 0.392 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.135 g, 0.432 mmol) according to the general procedure for urea formation (Method B), and was obtained as a light brown solid (0.051 g, 26% yield). 1H NMR (400MHz, CD3OD) δ=8.41 (s, 1H), 7.95-7.97 (m, 1H), 7.52-7.57 (m, 2H), 6.31 (s, 1H), 3.98-4.01 (m, 1H), 2.41 (s, 3H), 1.21-1.48 (m, 8H). LCMS: 499.1[M+H].

[0802] Example 49

[0803] 1-(4-(4-amino-1-(1-methylazacyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0804]

[0805] The title compound was obtained by starting with 3-(4-amino-3-fluorophenyl)-1-(1-methylazacyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B28, 0.130 g, 0.278 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.095 g, 0.306 mmol) according to the general procedure for urea formation (Method B), and was obtained as a white solid (0.010 g, 7% yield). 1 H NMR (400MHz, CD3OD) δ = 8.35-8.39 (m, 1H), 8.30 (s, 1H), 7.58-7.65 (m, 2H), 6.35 (s, 1H), 5.74-5.78 (m, 1H), 4.38-4.48 (m, 4H), 2.92 (s, 3H), 1.40-1.49 (m, 4H). LCMS: 532.2[M+H].

[0806] Example 50

[0807] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2,5-difluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0808]

[0809] The title compound was obtained by starting with 3-(4-amino-2,5-difluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B29, 0.050 g, 0.165 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.0516 g, 0.165 mmol) according to the general procedure for urea formation (Method B), and was obtained as a grayish-white solid (0.002 g, 3% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.60 (bs, 1H), 9.08 (bs, 1H), 8.17 (s, 1H), 8.03-8.08 (m, 1H), 7 .37-7.42(m, 1H), 6.16(s, 1H), 3.78-3.82(m, 1H), 1.32-1.42(m, 4H), 1.02-1.20(m, 4H). LCMS: 521.3[M+H].

[0810] Example 51

[0811] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-(hydroxymethyl)phenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0812]

[0813] The title compound was obtained from (2-amino-5-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)methanol (B30, 0.120 g, 0.405 mmol)) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)phenyl carbamate (C6, 0.126 g, 0.405 mmol) as an off-white solid (0.025 g, 12% yield). 1 H NMR (400MHz, DMSO-d6) δ = 11.14 (bs, 1H), 8.65 (bs, 1H), 8.38 (s, 1H), 8.06-8.08 (m, 1H ), 7.54-7.62(m, 2H), 6.19(s, 1H), 4.60(s, 2H), 3.90-3.94(m, 1H), 1.24-1.48(m, 8H). LCMS: 515.2[M+H].

[0814] Example 52

[0815] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(2-cyanopropane-2-yl)isoxazo-5-yl)urea

[0816]

[0817] The title compound was prepared following the general procedure for urea formation (Method B) starting from 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.120 g, 0.422 mmol) and (3-(2-cyanopropane-2-yl)isoxazol-5-yl)carbamate (C20, 0.126 g, 0.464 mmol), and was obtained as a grayish-white solid (0.029 mg, 14% yield). 1 H NMR (400MHz, CD3OD) δ = 8.30-8.37 (m, 2H), 7.50-7.55 (m, 2H), 6.35 (s, 1H), 3.80-3.86 (m, 1H), 1.78 (s, 6H), 1.17-1.37 (m, 4H). LCMS: 462.2[M+H].

[0818] Example 53

[0819] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(hydroxymethyl)isoxazo-5-yl)urea

[0820] Step 1: Synthesis of 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(((tert-butyldimethylsilyl)oxy)methyl)isoxazo-5-yl)urea

[0821]

[0822] The title compound was prepared following the general procedure for urea formation (Method B) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.100 g, 0.352 mmol) and (3-(((tert-butyldimethylsilyl)oxy)methyl)isoxazol-5-yl)carbamate (C21, 0.135 g, 0.387 mmol), and was obtained as a white solid (0.020 g, 8% yield). 1H NMR (400MHz, CD3OD) δ = 8.30-8.36 (m, 2H), 7.49-7.54 (m, 2H), 6.24 (s, 1H), 4.73 (s, 2H) ), 3.80-3.85(m, 1H), 1.29-1.33(m, 2H), 1.16-1.21(m, 2H), 0.99(s, 9H), 0.12(s, 6H). LCMS: 539.3[M+H].

[0823] Step 2: Synthesis of 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(hydroxymethyl)isoxazol-5-yl)urea

[0824]

[0825] At 0 °C, TBAF (1 M in THF, 0.037 mL, 0.037 mmol) was added to a solution of 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(((tert-butyldimethylsilyl)oxy)methyl)isoxazo-5-yl)urea (0.020 g, 0.037 mmol) in THF (3 mL), and the resulting mixture was stirred at 25 °C for 1 h. After the reaction was complete (as shown by TLC), the reaction mixture was concentrated under reduced pressure to give a crude material, which was purified by preparative HPLC (ELSD method, eluted with 0.1% TFA / (water and CAN)) to give the title compound (0.005 g, 31% yield, TFA salt) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ = δ 10.46 (bs, 1H), 8.98 (bs, 1H), 8.27-8.35 (m, 2H), 7.4 5-7.53 (m, 2H), 6.14 (s, 1H), 4.44 (s, 2H), 3.87-3.92 (m, 1H), 1.10-1.23 (m, 4H). LCMS: 425.2[M+H].

[0826] Example 54

[0827] 1-(5-(4-amino-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridin-3-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0828] Step 1: Synthesis of 1-(5-(1-cyclopropyl-4-((2,4-dimethoxybenzyl)amino)-1H-pyrazolo[4,3-c]pyridin-3-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0829]

[0830] The title compound was obtained according to the general procedure for urea formation (Method B), starting with 3-(6-aminopyridin-3-yl)-1-cyclopropyl-N-(2,4-dimethoxybenzyl)-1H-pyrazolo[4,3-c]pyridin-4-amine (B31, 0.050 g, 0.120 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.037 g, 0.120 mmol), and was given as a grayish-white gel (0.012 g, 15% yield). LCMS: 635.2 [M+H].

[0831] Step 2: Synthesis of 1-(5-(4-amino-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridin-3-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0832]

[0833] Triethylsilane (0.1 mL, 0.626 mmol) and TFA (0.1 mL, 1.298 mmol) were added to a solution of 1-(5-(1-cyclopropyl-4-((2,4-dimethoxybenzyl)amino)-1H-pyrazolo[4,3-c]pyridin-3-yl)pyridin-2-yl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)urea (0.012 g, 0.019 mmol) in DCM (2 mL), and the resulting mixture was stirred at 25 °C for 12 h. After the reaction was complete (as shown in LCMS), the reaction mixture was concentrated under reduced pressure to give a crude material, which was purified by preparative HPLC (eluting with 10 mM NH4OAc in ACN) to give the title product (0.001 g, 10% yield) as a grayish-white solid. 1 H NMR (400MHz, CD3OD) δ=8.64 (d, J=1.6Hz, 1H), 8.09-8.11 (m, 1H), 7.80 (d, J=6.0Hz, 1H), 7. 47-7.49 (m, 1H), 7.04 (d, J=6.4Hz, 1H), 6.39 (s, 1H), 3.80-3.86 (m, 1H), 0.90-1.50 (m, 8H). LCMS: 485.4[M+H].

[0834] Example 55

[0835] 1-(4-(4-amino-1-(2-hydroxyethyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0836]

[0837] The title compound was obtained from 2-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethane-1-ol (B32, 0.11 g, 0.382 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.10 g, 0.320 mmol) as an off-white solid (0.002 g, 1% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.66 (bs, 1H), 8.99 (bs, 1H), 8.26-8.30 (m, 2H), 7.48-7.54 (m, 2H), 6.94 ( bs, 2H), 6.22 (s, 1H), 4.90 (bs, 1H), 4.39 (t, J=6.0Hz, 2H), 3.85 (t, J=6.0Hz, 2H), 1.39-1.49 (m, 4H). LCMS: 507.2[M+H].

[0838] Example 56

[0839] 1-(4-(4-amino-1-(2-hydroxyethyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0840]

[0841] The title compound was obtained by starting with 2-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethane-1-ol (B32, 0.100 g, 0.347 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.108 g, 0.347 mmol) according to the general procedure for urea formation (Method B), and was obtained as a grayish-white solid (0.004 g, 3% yield). 1H NMR (400MHz, CD3OD) δ=8.36-8.41 (m, 2H), 7.52-7.59 (m, 2H), 6.83 (s, 1H), 4.58 (t, J=5.6Hz, 2H), 4.07 (t, J=5.2Hz, 2H), 1.49-1.59 (m, 4H). LCMS: 507.2[M+H].

[0842] Example 57

[0843] 1-(4-(4-amino-1-(2-methoxyethyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0844]

[0845] The title compound was obtained by starting with 3-(4-amino-3-fluorophenyl)-1-(2-methoxyethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B33, 0.125 g, 0.413 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.129 g, 0.413 mmol) according to the general procedure for urea formation (Method B), and was obtained as a grayish-white solid (0.020 g, 9% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.61 (bs, 1H), 8.98 (bs, 1H), 8.25-8.29 (m, 2H), 7.48-7.54 (m, 2H), 6.96 ( bs, 2H), 6.22 (s, 1H), 4.50 (t, J=5.2Hz, 2H), 3.82 (t, J=5.2Hz, 2H), 3.23 (s, 3H), 1.38-1.47 (m, 4H). LCMS: 521.2[M+H].

[0846] Example 58

[0847] 1-(4-(4-amino-1-(2-methoxyethyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0848]

[0849] The title compound was obtained by starting with 3-(4-amino-3-fluorophenyl)-1-(2-methoxyethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B33, 0.125 g, 0.413 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.129 g, 0.413 mmol) according to the general procedure for urea formation (Method B), and was obtained as a grayish-white solid (0.015 g, 7% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.07 (bs, 1H), 9.00 (bs, 1H), 8.31-8.35 (m, 2H), 7.47-7.55 (m , 2H), 6.92 (s, 1H), 4.53 (bs, 2H), 3.83 (t, J=5.2Hz, 2H), 3.23 (s, 3H), 1.51-1.52 (m, 4H). LCMS: 521.2[M+H].

[0850] Example 59

[0851] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-methylphenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0852]

[0853] The title compound was obtained by starting with 3-(4-amino-3-methylphenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B27, 0.100 g, 0.357 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.123 g, 0.392 mmol) according to the general procedure for urea formation (Method B), and was obtained as a white solid (0.026 g, 15% yield). 1 H NMR (400MHz, CD3OD) δ = 8.40 (s, 1H), 8.01-8.03 (m, 1H), 7.51-7.57 (m, 2H), 6.74 (s, 1H), 3.97-4.01 (m, 1H), 2.42 (s, 3H), 1.49-1.59 (m, 4H), 1.21-1.38 (m, 4H). LCMS: 499.5[M+H].

[0854] Example 60

[0855] 1-(4-(4-amino-1-(3-hydroxycyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0856] Step 1: 1-(4-(4-amino-1-(3-(benzyloxy)cyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0857]

[0858] The title compound was obtained following the general procedure for urea formation (Method B), starting with 3-(4-amino-3-fluorophenyl)-1-(3-(benzyloxy)cyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B26, 0.090 g, 0.223 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.076 g, 0.245 mmol), and was obtained as a brown gel (70 mg), which was used without further purification. LCMS: 623.4 [M+H].

[0859] Step 2: Synthesis of 1-(4-(4-amino-1-(3-hydroxycyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0860]

[0861] Boron trichloride (1 M in DCM, 0.899 mL, 0.899 mmol) was added dropwise to a solution of 1-(4-(4-amino-1-(3-(benzyloxy)cyclobutyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)urea (0.070 g, 0.112 mmol) in anhydrous DCM (5 mL) at -60 °C, and the resulting mixture was stirred at 0 °C for 3 h. After the reaction was complete (as shown by TLC and LCMS), the reaction mixture was cooled to -70 °C, neutralized with ammonia (25% in water), and extracted with DCM (2 × 10 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain a crude material, which was purified by preparative HPLC (eluting with 1% TFA in a mixture of water and CAN) to give the title product as a white solid (0.009 g, 15% yield). 1H NMR (400MHz, CD3OD) δ = 8.36-8.40 (m, 2H), 7.53-7.61 (m, 2H), 6.82 (s, 1H), 5.62-5.69 (m, 1H), 4.74-4.80 (m, 1H), 2.94-2.99 (m, 2H), 2.58-2.63 (m, 2H), 1.49-1.58 (m, 4H). LCMS: 533.7[M+H].

[0862] Example 61

[0863] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0864] Step 1: Synthesis of 1-(4-(1-cyclopropyl-4-((2,4-dimethoxybenzyl)amino)-1H-pyrazolo[4,3-c]pyridin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0865]

[0866] The title compound was obtained by following the general procedure for urea formation (Method B), starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-N-(2,4-dimethoxybenzyloxy)-1H-pyrazolo[4,3-c]pyridine-4-amine (B34, 0.220 g, 0.508 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.158 g, 0.508 mmol), and was obtained as a brown gel (0.093 g, 28% yield), which was used without further purification. LCMS: 652.3 [M+H].

[0867] Step 2: Synthesis of 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[4,3-c]pyridin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0868]

[0869] At 0 °C, triethylsilane (0.1 mL) and TFA (0.1 mL) were added to a solution of 1-(4-(1-cyclopropyl-4-((2,4-dimethoxybenzyl)amino)-1H-pyrazolo[4,3-c]pyridin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)urea (0.093 g, 0.143 mmol) in DCM (2 mL), and the resulting mixture was stirred at 25 °C for 12 h. After the reaction was complete (as shown in LCMS), the reaction mixture was concentrated under reduced pressure to give a crude material, which was purified by preparative HPLC (eluting with 0.1% TFA in a mixture of water and CAN) to give the title product (0.050 g, 68% yield) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ = 10.10 (bs, 1H), 9.06 (bs, 1H), 8.33-8.37 (m, 1H), 7.89 (bs, 2H), 7.83 (d, J = 7.2Hz, 1H), 7.54-7.5 7 (m, 1H), 7.44-7.47 (m, 1H), 7.32 (d, J=7.2Hz, 1H), 6.91 (s, 1H), 3.94-3.96 (m, 1H), 1.51-1.56 (m, 4H), 1.19-1.20 (m, 4H). LCMS: 502.5[M+H].

[0870] Example 62

[0871] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2,5-difluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0872]

[0873] The title compound was obtained by starting with 3-(4-amino-2,5-difluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B29, 0.100 g, 0.331 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.103 g, 0.331 mmol) according to the general procedure for urea formation (Method B), and was obtained as a grayish-white solid (0.002 g, 1% yield). 1H NMR (400MHz, CD3OD) δ = 8.24-8.29 (m, 2H), 7.38-7.43 (m, 1H), 6.85 (s, 1H), 3.80-3.85 (m, 1H), 1.47-1.59 (m, 4H), 1.20-1.33 (m, 4H). LCMS: 521.2[M+H].

[0874] Example 63

[0875] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-(hydroxymethyl)phenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0876]

[0877] The title compound was obtained by starting with (2-amino-5-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)phenyl)methanol (B30, 0.086 g, 0.290 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)phenyl carbamate (C7, 0.091 g, 0.290 mmol) as an off-white solid (0.004 g, 3% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.39 (bs, 1H), 8.58 (bs, 1H), 8.27 (s, 1H), 8.06-8.08 (m, 1H), 7.52-7.62 (m, 2H), 6.89 (s, 1H), 5.45 (t, J=5.6Hz, 1H), 4.59 (d, J=5.6Hz, 2H), 3.84-3.89 (m, 1H), 1.50-1.57 (m, 4H), 1.20-1.23 (m, 4H). LCMS: 515.2[M+H].

[0878] Example 64

[0879] 1-(5-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)pyridin-2-yl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0880]

[0881] The title compound was obtained by starting with 3-(6-aminopyridin-3-yl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B35, 0.085 g, 0.318 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.099 g, 0.318 mmol) according to the general procedure for urea formation (Method B), and was obtained as a grayish-white solid (0.003 g, 2% yield). 1 H NMR (400MHz, DMSO-d6) δ = 11.03 (bs, 1H), 9.83 (bs, 1H), 8.51 (bs, 1H), 8.27 (s, 1H), 8.01-8.04 (m , 1H), 7.72-7.74(m, 1H), 6.97(s, 1H), 3.86-3.88(m, 1H), 1.49-1.58(m, 4H), 1.12-1.24(m, 4H). LCMS: 486.1[M+H].

[0882] Example 65

[0883] 1-(4-(4-amino-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(tert-butyl)-4-methylisoxazol-5-yl)urea

[0884]

[0885] The title compound was prepared following the general procedure for urea formation (Method B) starting with 3-(4-amino-3-fluorophenyl)-1-cyclopropyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B1, 0.100 g, 0.352 mmol) and (3-(tert-butyl)-4-methylisoxazol-5-yl)carbamate (C22, 0.096 g, 0.350 mmol), and was obtained as a white solid (0.013 g, 8% yield). 1 HNMR (400MHz, CD3OD) δ = 8.39 (s, 1H), 8.22-8.26 (m, 1H), 7.46-7.55 (m, 2H), 3.94-3.97 (m, 1H), 2.09 (s, 3H), 1.35-1.39 (m, 11H), 1.21-1.23 (m, 2H). LCMS: 465.2[M+H].

[0886] Example 66

[0887] 1-(4-(4-amino-1-(2-hydroxy-2-methylpropyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0888]

[0889] The title compound was obtained by starting with 1-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol (B19, 0.100 g, 0.316 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.099 g, 0.316 mmol) according to the general procedure for urea formation (Method B), and was obtained as a white solid (0.033 g, 19% yield). 1 H NMR (400MHz, DMSO-d6) δ = 10.63 (bs, 1H), 8.97 (bs, 1H), 8.26-8.30 (m, 2H), 7.48-7.54 (m, 2 H), 6.96 (bs, 2H), 6.22 (s, 1H), 4.77 (bs, 1H), 4.28 (s, 2H), 1.39-1.49 (m, 4H), 1.15 (s, 6H). LCMS: 535.3[M+H].

[0890] Example 67

[0891] 1-(4-(4-amino-1-(2-hydroxy-2-methylpropyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0892]

[0893] The title compound was obtained by starting with 1-(4-amino-3-(4-amino-3-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-methylpropane-2-ol (B19, 0.100 g, 0.316 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.099 g, 0.316 mmol) according to the general procedure for urea formation (Method B), and was obtained as a white solid (0.022 g, 13% yield). 1H NMR (400MHz, DMSO-d6) δ = 10.08 (bs, 1H), 9.04 (bs, 1H), 8.42 (s, 1H), 8.31-8.35 (m, 1H), 7.47-7.55(m, 2H), 6.91(s, 1H), 4.31(s, 2H), 1.48-1.58(m, 4H), 1.17(s, 6H). LCMS: 535.3[M+H].

[0894] Example 68

[0895] 1-(4-(4-amino-1-(1-methylazacyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0896]

[0897] The title compound was obtained by starting with 3-(4-amino-3-fluorophenyl)-1-(1-methylazacyclobutane-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B28, 0.130 g, 0.278 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.095 g, 0.306 mmol) as a white solid (0.006 g, 4% yield). 1 H NMR (400MHz, CD3ODδ=8.26-8.41(m, 2H), 7.56-7.64(m, 2H), 6.84(s, 1H), 5.71-5. 74 (m, 1H), 4.29-4.90 (m, 4H), 2.85 (s, 3H), 1.52-1.59 (m, 4H). LCMS: 532.3 [M+H].

[0898] Example 69

[0899] 1-(4-(4-amino-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0900] Step 1: Synthesis of 1-(4-(4-amino-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0901]

[0902] The title compound was obtained following the general procedure for urea formation (Method A), starting with 3-(4-amino-3-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B36, 0.06 g, 0.160 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.05 g, 0.160 mmol), and was obtained as a pale brown solid (80 mg), which was used without further purification. LCMS: 593.3 [M+H].

[0903] Step 2: Synthesis of 1-(4-(4-amino-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0904]

[0905] At 0 °C, HCl (g) (4 M, 1 mL, 0.135 mmol) in dioxane was added to a solution of 1-(4-(4-amino-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)urea (0.08 g, 0.135 mmol) in DCM (5 mL), and the resulting mixture was stirred at 25 °C for 12 hours. After the reaction was complete (as shown by UPLC), the reaction mixture was concentrated under reduced pressure to give a crude material, which was purified by preparative HPLC (eluting with 10 mM NH4OAc in ACN) to give the title product (0.003 g, 5% yield) as a grayish-white solid. 1 H NMR (400MHz, CD3OD) δ = 8.30-8.34 (m, 1H), 8.24 (s, 1H), 7.49-7.54 (m, 2H), 6.31 (s, 1H), 1.37-1.46 (m, 4H). LCMS: 463.3[M+H].

[0906] Example 70

[0907] 1-(4-(4-amino-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0908] Step 1: Synthesis of 1-(4-(4-amino-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0909]

[0910] The title compound was obtained by following the general procedure for urea formation (Method A) starting with 3-(4-amino-3-fluorophenyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B36, 0.06 g, 0.160 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.05 g, 0.160 mmol), and was obtained as a pale brown solid (80 mg), which was used without further purification. LCMS: 593.3 [M+H].

[0911] Step 2: Synthesis of 1-(4-(4-amino-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0912]

[0913] At 0 °C, HCl (g) (4 M, 1 mL, 0.135 mmol) in dioxane was added to a solution of 1-(4-(4-amino-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)urea (0.08 g, 0.135 mmol) in DCM (5 mL), and the resulting mixture was stirred at 25 °C for 12 h. After the reaction was complete (as shown by UPLC), the reaction mixture was concentrated under reduced pressure to give a crude material, which was purified by preparative HPLC (eluting with 10 mM NH4OAc in CAN) to give the title product (0.007 g, 11% yield) as a grayish-white solid. 1 H NMR (400MHz, DMSO-d6) δ = 13.61 (bs, 1H), 10.04 (bs, 1H), 8.96 (bs, 1H), 8.27- 8.30 (m, 1H), 8.22 (s, 1H), 7.46-7.53 (m, 2H), 6.92 (s, 1H), 1.51-1.58 (m, 4H). LCMS: 463.3[M+H].

[0914] Example 71

[0915] 1-(4-(4-amino-1-(1-methylpiperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(3-(1-(trifluoromethyl)cyclopropyl)isoxazo-5-yl)urea

[0916]

[0917] The title compound was obtained by starting with 3-(4-amino-3-fluorophenyl)-1-(1-methylpiperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B37, 0.100 g, 0.179 mmol) and (3-(1-(trifluoromethyl)cyclopropyl)isoxazol-5-yl)carbamate (C6, 0.061 g, 0.197 mmol) according to the general procedure for urea formation (Method B), and was obtained as a white solid (0.008 g, 8% yield). 1 H NMR (400MHz, CD3OD) δ = 8.34-8.38 (m, 2H), 7.52-7.57 (m, 2H), 6.35 (s, 1H), 5.12-5.17 (m, 1H), 3.73-3 .76 (m, 2H), 3.33-3.39 (m, 2H), 3.00 (s, 3H), 2.59-2.62 (m, 2H), 2.35-2.39 (m, 2H), 1.45-1.49 (m, 4H). LCMS: 560.2[M+H].

[0918] Example 72

[0919] 1-(4-(4-amino-1-(1-methylpiperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-2-fluorophenyl)-3-(5-(1-(trifluoromethyl)cyclopropyl)isoxazo-3-yl)urea

[0920]

[0921] The title compound was obtained by starting with 3-(4-amino-3-fluorophenyl)-1-(1-methylpiperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (B37, 0.100 g, 0.179 mmol) and (5-(1-(trifluoromethyl)cyclopropyl)isoxazol-3-yl)carbamate (C7, 0.061 g, 0.197 mmol) as a brown solid (0.004 g, 4% yield). 1H NMR (400MHz, CD3OD) δ = 8.36-8.40 (m, 2H), 7.51-7.56 (m, 2H), 6.82 (s, 1H), 5.10-5.18 (m, 1H), 3.73-3 .76 (m, 2H), 3.37-3.39 (m, 2H), 3.00 (s, 3H), 2.59-2.62 (m, 2H), 2.35-2.38 (m, 2H), 1.39-1.59 (m, 4H). LCMS: 560.3[M+H].

[0922] Biological Example 1

[0923] Biochemical determination of compounds

[0924] The inhibitory activity of representative compounds on the release of NEK7 and IL-1β was tested according to the above procedure. The results are given in the table below.

[0925] Table 2. Activities of representative compounds

[0926]

[0927]

[0928]

[0929] For NEK7 IC in Table 2 50 active:

[0930] * IC 50 Greater than 501 nM

[0931] ** IC 50 301-500nM

[0932] *** IC 50 151-300nM

[0933] **** IC 50 Less than 150 nM

[0934] For IL-1βIC in Table 2 50 active:

[0935] + IC 50 Greater than 501 nM

[0936] ++ IC 50 201-500nM

[0937] +++ IC 50 51-200nM

[0938] ++++IC 50Less than 50 nM

[0939] - Indicates no value was detected.

[0940] The various embodiments described above may be combined to provide other embodiments. All U.S. patents, U.S. patent application publications, U.S. patent publications, foreign patents, foreign patent applications and non-patent publications referenced in this specification and / or listed in the application data sheets, including but not limited to U.S. Provisional Application No. 63 / 036,244, filed June 8, 2020; U.S. Provisional Application No. 63 / 170,776, filed April 5, 2021; and U.S. Provisional Application No. 63 / 185,257, filed May 6, 2021, are incorporated herein by reference in their entirety. Where necessary, aspects of the embodiments may be modified to incorporate the ideas of various patents, applications and publications, thereby providing other embodiments.

[0941] These and other changes can be made to the embodiments based on the detailed description above. Generally, the terminology used in the appended claims should not be construed as limiting the claims to the specific embodiments disclosed in this specification and claims, but rather should be interpreted to include all possible embodiments together with the full scope of equivalents defined in such claims. Therefore, the claims are not limited by this disclosure.

Claims

1. A compound having one of the following structures: ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ;or , Or a salt acceptable for its medication.

2. A compound having one of the following structures: ； ； ； ； ； ； ;or , Or a salt acceptable for its medication.

3. A pharmaceutical composition comprising the compound of claim 1 or 2 and a pharmaceutically acceptable carrier, diluent, or excipient.

4. Use of the compound of claim 1 or 2 or the pharmaceutical composition of claim 3 in the preparation of a medicament for treating NLRP3-mediated conditions in individuals in need.

5. The use as described in claim 4, wherein the condition is selected from autoimmune diseases, inflammatory diseases, cardiovascular diseases, neurodegenerative diseases, bacterial and viral infections, allergies, asthma, multiple organ failure, kidney disease, cancer, transplant rejection, and respiratory diseases.

6. The use as described in claim 4, wherein the condition is selected from type II diabetes, atherosclerosis, Alzheimer's disease, fatty liver, metabolic syndrome, asthma, psoriasis, obesity, acute and chronic tissue damage caused by infection, gout, arthritis, macular degeneration, enteritis, hepatitis, peritonitis, silicosis, UV-induced sunburn, contact hypersensitivity, sepsis, cancer, neurodegenerative diseases, multiple sclerosis, and Murphy-Weil syndrome.

7. The use as described in claim 4, wherein the condition is pancreatitis.

8. The use as described in claim 4, wherein the condition is aplasia.

9. The use as described in claim 4, wherein the condition is platelet aggregation.

10. The use as described in claim 4, wherein the condition is an ischemic condition.

11. The use as described in claim 4, wherein the condition is lung injury.

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