Preparation of p2x3 antagonists

Abstract

Described herein are two methods for preparing P2X3 antagonist (5) -2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester in a stepwise manner as well as chemical intermediates used in the synthesis methods.

Description

[0001] Cross-references

[0002] This application claims the benefits of U.S. Provisional Application No. 62 / 977,004, filed February 14, 2020, and U.S. Provisional Application No. 63 / 144,902, filed February 2, 2021, the entire contents of which are incorporated herein by reference. Background Technology

[0003] P2X purine receptors are a family of ion channels activated by extracellular adenosine triphosphate (ATP). Purine receptors are involved in a variety of biological functions. The P2X3 receptor subunit is a member of this family. It was originally cloned from the rat dorsal root ganglion. Chen et al., Nature, Vol. 377, pp. 428-431 (1995). The nucleotide and amino acid sequences of rat and human P2X3 are currently known. Lewis et al., Nature, Vol. 377, pp. 432-435 (1995); and Garcia-Guzman et al., Brain Res. Mol. Brain Res., Vol. 47, pp. 59-66 (1997). Summary of the Invention

[0004] This article describes a method for synthesizing a P2X3 antagonist, wherein the P2X3 antagonist is (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1) or a pharmaceutically acceptable salt thereof.

[0005] One aspect is a method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1):

[0006]

[0007] Includes having a structure:

[0008] The compound is contacted with an amide coupling agent and methylamine or a salt thereof. In some embodiments, the amide coupling agent is carbonyl diimidazole. In some embodiments, the amide coupling agent is propanephosphonic anhydride (T3P).

[0009] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0010] The compound is formed by including the following structure:

[0011] The compound was prepared by contacting it with 2-amino-4-methylpyridine and optionally sodium borohydride in the presence of a solvent.

[0012] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0013] The compound is formed by including the following structure:

[0014] The compound is prepared by contacting it with a brominating agent. In some embodiments, the brominating agent is N-bromosuccinimide in the presence of an acid.

[0015] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0016] The compound is formed by including the following structure:

[0017] The compound is prepared by contacting it with methyl chloroformate and a base. In some embodiments, the base is sodium bicarbonate.

[0018] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0019] The compound is formed by including the following structure:

[0020] The compound is prepared by contacting it with hydrogen chloride in the presence of a solvent. In some embodiments, the solvent is ethyl acetate.

[0021] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0022] The compound is formed by including the following structure:

[0023] The compound is prepared by contacting it with a hydrogenation catalyst and hydrogen gas. In some embodiments, the hydrogenation catalyst is carbon-supported palladium, palladium hydroxide, carbon-supported rhodium, alumina-supported rhodium, platinum oxide, or carbon-supported platinum. In some embodiments, the hydrogenation catalyst is carbon-supported palladium.

[0024] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0025] The compound is formed by including the following structure:

[0026] Compounds with the following structures:

[0027] The compound is prepared by contacting a base with the compound. In some embodiments, the base is a mixture of potassium bicarbonate and potassium carbonate.

[0028] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0029] The compound is formed by including the following structure:

[0030] The compound is prepared by contacting it with 2,2,6,6-tetramethylpiperidine-1-oxy or propanephosphonic anhydride (T3P). Another aspect is the method for preparing (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1):

[0031]

[0032] Includes having a structure:

[0033] The compound is contacted with an amide coupling agent and methylamine or a salt thereof. In some embodiments, the amide coupling agent is carbonyl diimidazole. In some embodiments, the amide coupling agent is propanephosphonic anhydride (T3P).

[0034] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0035] The compound is formed by including the following structure:

[0036] The compound is prepared by contacting a base and optionally sodium borohydride in the presence of a solvent. In some embodiments, the solvent is an aqueous solution of tetrahydrofuran, dioxane, 2-methyltetrahydrofuran, an aqueous solution of methanol, an aqueous solution of ethanol, or an aqueous solution of acetonitrile. In some embodiments, the solvent is an aqueous solution of tetrahydrofuran. In some embodiments, the base is lithium hydroxide.

[0037] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0038] The compound is formed by including the following structure:

[0039] The compound was prepared by contacting it with 2-amino-4-methylpyridine.

[0040] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0041] The compound is formed by including the following structure:

[0042] The compound is prepared by contacting it with a brominating agent. In some embodiments, the brominating agent is copper(II) bromide. In some embodiments, the brominating agent is liquid bromine.

[0043] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0044] The compound is formed by including the following structure:

[0045] The compound is prepared by contacting it with methyl chloroformate and a base. In some embodiments, the base is sodium bicarbonate.

[0046] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0047] The compound is formed by including the following structure:

[0048] The compound was prepared by contacting it with hydrogen chloride and methanol.

[0049] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0050] The compound is formed by including the following structure:

[0051] The compound is prepared by contacting it with hydrogen chloride in the presence of a solvent. In some embodiments, the solvent is ethyl acetate.

[0052] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0053] The compound is formed by including the following structure: The compound is prepared by contacting it with tert-butyl 3,5-difluorobenzoate and a base. In some embodiments, the base is lithium diisopropylamino.

[0054] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0055] The compound is formed by including the following structure:

[0056] The compound is prepared by contacting it with N,O-dimethylhydroxylamine, an amide coupling agent, and 1-hydroxybenzotriazole. In some embodiments, the amide coupling agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide.

[0057] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0058] The compound is formed by including the following structure:

[0059] The compound is prepared by contacting it with a hydrogenation catalyst and hydrogen gas. In some embodiments, the hydrogenation catalyst is carbon-supported palladium, palladium hydroxide, carbon-supported rhodium, alumina-supported rhodium, platinum oxide, or carbon-supported platinum. In some embodiments, the hydrogenation catalyst is carbon-supported palladium.

[0060] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0061] The compound is formed by including the following structure:

[0062] The compound is prepared by contacting a base with an aqueous solution of tetrahydrofuran. In some embodiments, the base is sodium hydroxide.

[0063] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0064] The compound is formed by including the following structure:

[0065] The compound was prepared by contacting it with (ethoxycarbonylmethylene)triphenylphosphine.

[0066] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0067] The compound is formed by including the following structure:

[0068] The compound was prepared by contacting it with 2,2,6,6-tetramethylpiperidine 1-oxy group or with T3P.

[0069] This article further discloses a method for preparing (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1), comprising:

[0070] A) Has a structure:

[0071] The compound reacts with 2,2,6,6-tetramethylpiperidine 1-oxy or T3P to produce a compound with the structure:

[0072] Compounds;

[0073] B) Then it has a structure:

[0074] Compounds and having the following structures: The compound reacts with a mixture of potassium bicarbonate and potassium carbonate to produce a compound with the structure:

[0075] Compounds;

[0076] C) Then it has a structure:

[0077] The compound reacts with carbon-supported palladium and hydrogen to produce a compound with the structure: Compounds;

[0078] D) Then it has a structure:

[0079] The compound reacts with hydrogen chloride in ethyl acetate to produce a compound with the structure: Compounds;

[0080] E) Then it has a structure:

[0081] The compound reacts with methyl chloroformate and sodium bicarbonate to produce a compound with the structure:

[0082] Compounds;

[0083] F) Then it has a structure:

[0084] The compound reacts with N-bromosuccinimide in the presence of acid to produce a compound with the structure:

[0085] Compounds;

[0086] G) Then it has a structure:

[0087] The compound reacts with 2-amino-4-methylpyridine and optionally sodium borohydride to produce a compound with the structure:

[0088] Compounds;

[0089] H) Then it has a structure:

[0090] The compound reacts with carbonyl diimidazole and methylamine to produce a compound with the structure: Compounds.

[0091] This article further discloses a method for preparing (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1), comprising:

[0092] A) Has a structure:

[0093] The compound reacts with 2,2,6,6-tetramethylpiperidine 1-oxy or T3P to produce a compound with the structure:

[0094] Compounds;

[0095] B) Then it has a structure:

[0096] The compound reacts with (ethoxycarbonylmethylene)triphenylphosphine to produce a compound with the structure:

[0097] Compounds;

[0098] C) Then it has a structure:

[0099] The compound reacts with sodium hydroxide in an aqueous solution of tetrahydrofuran to produce a compound with the structure:

[0100] Compounds;

[0101] D) Then it has a structure:

[0102] The compound reacts with carbon-supported palladium and hydrogen to produce a compound with the structure:

[0103] Compounds;

[0104] E) Then it has a structure:

[0105] The compound reacts with N,O-dimethylhydroxylamine, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and 1-hydroxybenzotriazole to produce a compound with the structure:

[0106] Compounds;

[0107] F) Then it has a structure:

[0108] The compound reacts with tert-butyl 3,5-difluorobenzoate and lithium diisopropylamino to produce a compound with the structure:

[0109] Compounds;

[0110] G) Then it has a structure:

[0111] The compound reacts with hydrogen chloride in ethyl acetate to produce a compound with the structure:

[0112] Compounds;

[0113] H) Then it has a structure:

[0114] The compound reacts with hydrogen chloride and methanol to produce a compound with the structure: Compounds;

[0115] I) Then it has a structure:

[0116] The compound reacts with methyl chloroformate and sodium bicarbonate to produce a compound with the structure:

[0117] Compounds;

[0118] J) Then it has a structure:

[0119] The compound reacts with copper(II) bromide to produce a compound with the structure: Compounds;

[0120] K) then has a structure:

[0121] The compound reacts with 2-amino-4-methylpyridine to produce a compound with the structure:

[0122] Compounds;

[0123] L) then has a structure:

[0124] The compound reacts with lithium hydroxide in an aqueous tetrahydrofuran solution, and then with sodium borohydride to produce a compound with the structure: Compounds;

[0125] M) then has a structure:

[0126] The compound reacts with carbonyl diimidazole and methylamine to produce a compound with the structure: Compounds.

[0127] This article further discloses a compound having the following structure:

[0128] Or its pharmaceutically acceptable salt.

[0129] This article further discloses a compound having the following structure:

[0130] Or its pharmaceutically acceptable salt.

[0131] This article further discloses a compound having the following structure:

[0132] Or its pharmaceutically acceptable salt.

[0133] This article further discloses a compound having the following structure:

[0134] Or its pharmaceutically acceptable salt.

[0135] This article further discloses a compound having the following structure:

[0136] Or its pharmaceutically acceptable salt.

[0137] This article further discloses a compound having the following structure:

[0138] Or its pharmaceutically acceptable salt.

[0139] This article further discloses a compound having the following structure:

[0140] Or its pharmaceutically acceptable salt.

[0141] This article further discloses a compound having the following structure:

[0142] Or its pharmaceutically acceptable salt.

[0143] This article further discloses a compound having the following structure:

[0144] Or its pharmaceutically acceptable salt.

[0145] This article further discloses a compound having the following structure:

[0146] Or its pharmaceutically acceptable salt.

[0147] Incorporation

[0148] All publications, patents and patent applications mentioned in this specification are incorporated herein by reference, as if each individual publication, patent or patent application were expressly and individually indicated to be incorporated by reference. Detailed Implementation

[0149] Large-scale preparation of clinically useful drug candidates typically requires good manufacturing practices. This article provides certain processes and methods for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1) or a pharmaceutically acceptable salt thereof.

[0150] definition

[0151] As used in the specification and appended claims, unless otherwise stated, the following terms have the meanings as follows.

[0152] As used herein and in the appended claims, the singular forms “a,” “and,” and “the” include plural indicators unless the context clearly specifies otherwise. Thus, for example, reference to “reagent” includes a variety of such reagents, and reference to “the cell” includes reference to one or more cells and their equivalents, etc.

[0153] When the scope is used herein for physical properties, such as molecular weight, or chemical properties, such as chemical formula, it is intended to include all combinations and sub-combinations of the scope and specific embodiments thereof.

[0154] When referring to a number or range of values, the term “approximately” means that the number or range of values ​​referred to is an approximation within experimental variability (or statistical experimental error), and therefore the number or range of values ​​varies between 1% and 15% of the specified number or range of values.

[0155] The term “comprising” (and related terms such as “comprise”, “comprises”, “having”, or “including”) is not intended to exclude embodiments in which, for example, any embodiment of a substance composition, composition, method, or process described herein is “composed of the described features” or “consistently composed of the described features”.

[0156] The terms "object" or "patient" include both mammals and non-mammals. Examples of mammals include, but are not limited to, any member of the mammal class: humans, non-human primates (such as chimpanzees) and other apes and monkeys; farm animals such as cattle, horses, sheep, goats, and pigs; domesticated animals such as rabbits, dogs, and cats; and laboratory animals, including rodents such as rats, mice, and guinea pigs. Examples of non-mammals include, but are not limited to, birds, fish, etc. In one embodiment of the methods and compositions provided herein, the mammal is a human.

[0157] As used herein, the terms “treatment” or “treating,” or “mitigation” or “improvement” are used interchangeably. These terms refer to methods of achieving beneficial or desired outcomes, including but not limited to therapeutic and / or preventative benefits. 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 physical symptoms associated with the underlying condition, such that improvement is observed in the patient even though the patient still has the underlying condition. For preventative benefits, the composition is administered to patients at risk of developing a specific disease, or to patients who report one or more physical symptoms of a disease, even if the disease has already been diagnosed.

[0158] "Pharmaceutically acceptable salts" include both acid and base addition salts. The term "pharmaceutically acceptable salt" for any compound described herein is intended to include any and all pharmaceutically suitable salt forms. Preferred pharmaceutically acceptable salts of the compounds described herein are pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.

[0159] "Pharmaceutically acceptable acid addition salts" refer to those salts that retain the bioavailability and properties of the free base, are not biologically or otherwise undesirable, and are formed from inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, hydroiodic acid, hydrofluoric acid, phosphorous acid, etc. It also includes salts formed from organic acids such as aliphatic monocarboxylic acids and dicarboxylic acids, phenyl-substituted alkanes, hydroxyalkanes, alkanedicosides, aromatic acids, aliphatic and aromatic sulfonic acids, etc., and includes, for example, acetic acid, trifluoroacetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc. Therefore, exemplary salts include sulfates, pyrosulfates, bisulfates, sulfites, nitrates, phosphates, monohydrophosphates, dihydrophosphates, metaphosphates, pyrophosphates, chlorides, bromides, iodides, acetates, trifluoroacetates, propionates, caprylates, isobutyrates, oxalates, malonates, succinate caprylates, sebacic acid salts, fumarates, maleates, mandelates, benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates, phthalates, benzenesulfonates, toluenesulfonates, phenylacetates, citrates, lactates, malates, tartrates, methanesulfonates, etc. Salts of amino acids, such as arginine salts, gluconates, and galacturons, are also considered (see, for example, Berge SM et al., "Pharmaceutical Salts," Journal of Pharmaceutical Science, 66:1-19 (1997)). Acid addition salts of basic compounds are prepared by contacting the free base form with a sufficient amount of the desired acid to produce a salt.

[0160] "Pharmaceutically acceptable base addition salts" refer to those salts that retain the bioavailability and properties of the free acid, and which are not biologically or otherwise undesirable. These salts are prepared by adding an inorganic or organic base to the free acid. In some embodiments, pharmaceutically acceptable base addition salts are formed from metals or amines (e.g., alkali metals and alkaline earth metals or organic amines). Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Salts derived from organic bases include, but are not limited to, salts of the following: primary amines, secondary amines, and tertiary amines; substituted amines, including naturally occurring substituted amines; cyclic amines; and basic ion exchange resins, such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, N,N-dibenzylethylenediamine, chloroprocaine, halamine, choline, betaine, ethylenediamine, ethylenediphenylamine, N-methylglucosamine, glucosamine, methylglucosamine, theobromine, purines, piperazine, piperidine, N-ethylpiperidine, polyamine resins, etc. See Berge et al. above.

[0161] As used herein, the term "pharmaceutical composition" refers to a product consisting of a mixture or combination of more than one active ingredient, and includes fixed and non-fixed combinations of active ingredients. The term "fixed combination" refers to the simultaneous administration of the active ingredients to a patient as a single entity or dose. The term "non-fixed combination" refers to the simultaneous, co-administration, or sequential administration of the active ingredients as separate entities to a patient without a specific time limit for intervention, wherein such administration provides effective levels of both compounds in the patient's body. The latter also applies to cocktail therapies, such as the administration of three or more active ingredients.

[0162] As used herein, the terms “co-administration” and the like mean “administering a selected therapeutic agent to a single patient” and “including treatment regimens in which the agent is administered via the same or different routes of administration or at the same or different times”.

[0163] The term "activator" is used in this specification to refer to any molecular substance that causes activation of the indicated receptor when the substance is applied topically, whether the substance itself binds to the receptor or a metabolite of the substance binds to the receptor. Therefore, an activator can be a ligand of the receptor, or it can be an activator metabolized into a receptor ligand, i.e., a metabolite formed in the tissue and the actual ligand.

[0164] As used in this article, the term "antagonist" refers to a small molecule agent that binds to nuclear hormone receptors and subsequently reduces agonist-induced transcriptional activity of nuclear hormone receptors.

[0165] As used in this article, the term "agonist" refers to a small molecule agent that binds to nuclear hormone receptors and subsequently increases the transcriptional activity of nuclear hormone receptors in the absence of known agonists.

[0166] As used in this article, the term "reverse agonist" refers to a small molecule agent that binds to a nuclear hormone receptor and subsequently reduces the basal level of the existing transcriptional activity of the nuclear hormone receptor in the absence of a known agonist.

[0167] As used herein, the term “regulation” refers to direct or indirect interaction with a target protein to alter its activity, including, by way of example only, inhibiting, or limiting or reducing, the activity of the target protein.

[0168] As used herein, the term "modifier" refers to a compound that alters the activity of a target. For example, a modifier can result in an increase or decrease in a specific level of activity of a target compared to the level of activity in the absence of a modifier. In some embodiments, a modifier is an inhibitor that reduces the level of one or more activities of the target. In some embodiments, an inhibitor completely blocks one or more activities of the target.

[0169] compound

[0170] In some embodiments, the P2X3 antagonist described herein is (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1) or a pharmaceutically acceptable salt thereof. Compound 1 has the following structure:

[0171]

[0172] In some embodiments, the starting material for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is

[0173] In some embodiments, the P2X3 antagonist described herein is (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1) or a pharmaceutically acceptable salt thereof. Compound 1 has the following structure:

[0174]

[0175] In some embodiments, the starting material for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is Where R 1 It is a straight-chain or branched alkyl group containing 1 to 8 carbon atoms. In some embodiments, the intermediate for synthesizing compound 1 is... In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is Where R 2 It is a branched alkyl group containing 1 to 8 carbon atoms. In some embodiments, the intermediate for synthesizing compound 1 is... In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is Where R 3 It is a straight-chain or branched alkyl group containing 1 to 8 carbon atoms. In some embodiments, the intermediate for synthesizing compound 1 is... In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is Where R 3 It is hydrogen, or a straight-chain or branched alkyl group containing 1 to 8 carbon atoms. In some embodiments, the intermediate for synthesizing compound 1 is... In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is Where X is Br, Cl or I, and R 3It is hydrogen or a straight-chain or branched alkyl group containing 1 to 8 carbon atoms. In some embodiments, the intermediate for synthesizing compound 1 is... Where X is Br, Cl, or I. In some embodiments, the intermediate for synthesizing compound 1 is... Where X is Br. In some embodiments, the intermediate for synthesizing compound 1 is... Where X is Br, Cl, or I. In some embodiments, the intermediate for synthesizing compound 1 is... Where X is Br. In some embodiments, the intermediate for synthesizing compound 1 is... Where R 3 It is hydrogen or a straight-chain or branched alkyl group containing 1 to 8 carbon atoms. In some embodiments, the intermediate for synthesizing compound 1 is... In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is Where R 3 It is a straight-chain or branched alkyl group containing 1 to 8 carbon atoms. In some embodiments, the intermediate for synthesizing compound 1 is... In some embodiments, the intermediate for synthesizing compound 1 is In some embodiments, the intermediate for synthesizing compound 1 is

[0176] Other forms of compounds

[0177] The compounds described herein may exist in certain cases as diastereomers, enantiomers, or other stereoisomers. The compounds provided herein include all diastereomers, enantiomers, and epimers, as well as suitable mixtures thereof. Separation of stereoisomers can be performed by chromatography or by forming diastereomers and separating them by recrystallization or chromatography, or any combination thereof. (Jean Jacques, Andre Collet, Samuel H. Wilen, “Enantiomers, Racemates and Resolutions”, John Wiley and Sons, Inc., 1981, incorporated herein by reference). Stereoisomers can also be obtained by stereoselective synthesis.

[0178] In some cases, compounds can exist as tautomers. All tautomers are enclosed in the formulas described herein.

[0179] Pharmaceutically acceptable salts

[0180] In some embodiments, the compounds described herein are present as pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating a disease by administering such pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating a disease by administering such pharmaceutically acceptable salts as a pharmaceutical composition.

[0181] In some embodiments, the compounds described herein have acidic or basic groups, and thus react with a variety of inorganic or organic bases and any of inorganic and organic acids to form pharmaceutically acceptable salts. In some embodiments, these salts are prepared in situ during the final isolation and purification of the compounds of the invention, or by reacting the purified compound in its free form with a suitable acid or base alone and separating the resulting salt.

[0182] In some embodiments, a pharmaceutically acceptable salt of compound 1 is an acetate, benzoate, benzenesulfonate, bitartrate, carbonate, citrate, fumarate, gluconate, hydrobromide, hydrochloride, maleate, methanesulfonate, nitrate, phosphate, salicylate, succinate, sulfate, or tartrate. In some embodiments, a pharmaceutically acceptable salt of compound 1 is a monohydrochloride. In a further embodiment, a pharmaceutically acceptable salt of compound 1 is a monohydrochloride.

[0183] solvates

[0184] In some embodiments, the compounds described herein are present as solvates. The present invention provides a method for treating a disease by administering such a solvate. The present invention also provides a method for treating a disease by administering such a solvate as a pharmaceutical composition.

[0185] The solvates contain stoichiometric or non-stoichiometric amounts of solvent and, in some embodiments, are formed during a crystallization process using a pharmaceutically acceptable solvent (e.g., water, ethanol, etc.). When the solvent is water, a hydrate is formed, or when the solvent is an alcohol, an alcohol is formed. The solvates of the compounds described herein can be conveniently prepared or formed during the methods described herein. By way of example only, the hydrates of the compounds described herein are conveniently prepared by recrystallization from an aqueous / organic solvent mixture using an organic solvent (including, but not limited to, dioxane, tetrahydrofuran, or methanol). Furthermore, the compounds provided herein exist in both solvated and non-solvated forms. Generally, for the purposes of the compounds and methods provided herein, the solvated form is considered equivalent to the non-solvated form.

[0186] Labeled compounds

[0187] In some embodiments, the compounds described herein are present in their isotopically labeled form. In some embodiments, the methods disclosed herein include methods of treating a disease by administering such isotopically labeled compounds. In some embodiments, the methods disclosed herein include methods of treating a disease by administering such isotopically labeled compounds as a pharmaceutical composition. Thus, in some embodiments, the compounds disclosed herein include isotopically labeled compounds that are the same as those described herein, but in the fact that one or more atoms are replaced by atoms having atomic masses or mass numbers different from those normally found in nature. Examples of isotopes incorporated into the compounds of the present invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine, for example, respectively. 2 H, 3 H, 13 C 14 C l5 N、 18 O、 17 O、 31 P, 32 P, 35 S, 18 F, and 36 Cl. The compounds described herein, as well as pharmaceutically acceptable salts, esters, solvates, hydrates, or derivatives thereof containing the aforementioned isotopes and / or other atoms, are within the scope of this invention. Certain isotope-labeled compounds, for example, those doped with... 3 H and 14 Those radioactive isotopes of C can be used for the determination of drug and / or substrate tissue distribution. Tritium isotopes, i.e. 3 H and carbon-14 (i.e.) 14 C), is particularly preferred due to its ease of preparation and detectability. Furthermore, heavy isotopes such as deuterium (i.e., 2 H) substitution produces certain therapeutic advantages due to greater metabolic stability, such as increased in vivo half-life or reduced dose requirement. The increased deuterium incorporation level produces a detectable kinetic isotope effect (KIE) compared to compound 1 with naturally occurring deuterium levels, which may affect the pharmacokinetic, pharmacological, and / or toxicological parameters of compound 1. In some embodiments, the isotopically labeled compound or its pharmaceutically acceptable salt is prepared by any suitable method.

[0188] In some embodiments, at least one hydrogen atom in compound 1 is replaced by deuterium.

[0189] In some embodiments, the compounds described herein are labeled by other means, including but not limited to the use of chromophores or fluorescent moieties, bioluminescent labeling, or chemiluminescent labeling.

[0190] Preparation method

[0191] In some embodiments, the synthesis of the compounds described herein is accomplished using methods described in the chemical literature, methods described herein, or combinations thereof. Furthermore, the solvents, temperatures, and other reaction conditions presented herein may be varied.

[0192] In other embodiments, the starting materials and reagents used to synthesize the compounds described herein are synthesized or obtained from commercial sources, such as, but not limited to, Sigma-Aldrich, Fischer Scientific (Fischer Chemicals), and Acros Organics. In further embodiments, the compounds described herein and other related compounds with different substituents are synthesized using the techniques and materials described herein as well as techniques and materials recognized in the art, such as those described, for example, in: Fieser and Fieser's Reagents for Organic Synthesis, Volumes 1–17 (John Wiley and Sons, 1991); Rodd's Chemistry of Carbon Compounds, Volumes 1–5 and Supplements (Elsevier Science Publishers, 1989); Organic Reactions, Volumes 1–40 (John Wiley and Sons, 1991); Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1989), March, Advanced Organic Chemistry, 4th Edition (Wiley 1992); Carey and Sundberg, Advanced Organic Chemistry, 4th Edition, Volumes A and B (Plenum 2000, 2001); and Green and Wuts, Protective Groups in Organic Synthesis, 3rd Edition (Wiley 1999) (all of which are incorporated herein by reference). General methods for preparing the compounds disclosed herein are available from reactions, and the reactions can be modified by using appropriate reagents and conditions to introduce the various motifs found in the formulas provided herein.

[0193] In some embodiments, the method for preparing (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1) is as follows:

[0194] Includes having a structure:

[0195] The compound is contacted with an amide coupling agent and methylamine or a salt thereof. In some embodiments, the amide coupling agent is carbonyl diimidazole. In some embodiments, the amide coupling agent is propanephosphonic anhydride (T3P). In some embodiments, the amide coupling agent is a carbodiimide coupling agent. In some embodiments, the amide coupling agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide. In some embodiments, the amide coupling agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide with added N-hydroxybenzotriazole.

[0196] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0197] The compound is formed by including the following structure:

[0198] The compound is prepared by contacting it with 2-amino-4-methylpyridine. In some embodiments, the method includes making a compound having the following structure:

[0199] The compound is contacted with 2-amino-4-methylpyridine and sodium borohydride. In some embodiments, the method includes making a compound having the structure:

[0200] The compound is contacted with 2-amino-4-methylpyridine in the presence of a solvent. In some embodiments, the solvent is acetonitrile.

[0201] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0202] The compound is formed by including the following structure:

[0203] The compound is prepared by contacting it with a brominating agent. In some embodiments, the brominating agent is N-bromosuccinimide. In some embodiments, the brominating agent is copper(II) bromide. In some embodiments, the brominating agent is liquid bromine.

[0204] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0205] The compound is formed by including the following structure:

[0206] The compound is prepared by contacting it with methyl chloroformate and a base. In some embodiments, the base is sodium bicarbonate.

[0207] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0208] The compound is formed by including the following structure:

[0209] The compound is prepared by contacting it with hydrogen chloride in the presence of a solvent. In some embodiments, the solvent is ethyl acetate.

[0210] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0211] The compound is formed by including the following structure:

[0212] The compound is prepared by contacting it with a hydrogenation catalyst and hydrogen gas. In some embodiments, the hydrogenation catalyst is carbon-supported palladium, palladium hydroxide, carbon-supported rhodium, alumina-supported rhodium, platinum oxide, or carbon-supported platinum. In some embodiments, the hydrogenation catalyst is carbon-supported palladium.

[0213] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0214] The compound is formed by including the following structure:

[0215] Compounds and compounds with structure The compound is prepared by contacting a base with the compound. In some embodiments, the base is a mixture of potassium bicarbonate and potassium carbonate.

[0216] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0217] The compound is formed by including the following structure:

[0218] The compound was prepared by contacting it with 2,2,6,6-tetramethylpiperidine 1-oxy or T3P.

[0219] This article further discloses a method for preparing (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1), comprising:

[0220] A) Has a structure:

[0221] The compound reacts with 2,2,6,6-tetramethylpiperidine 1-oxy or T3P to produce a compound with the structure:

[0222] Compounds;

[0223] B) Then it has a structure:

[0224] Compounds with the following structures:

[0225] The compound reacts with a mixture of potassium bicarbonate and potassium carbonate to produce a compound with the structure:

[0226] Compounds;

[0227] C) Then it has a structure:

[0228] The compound reacts with carbon-supported palladium and hydrogen to produce a compound with the structure: Compounds;

[0229] D) Then it has a structure:

[0230] The compound reacts with hydrogen chloride in ethyl acetate to produce a compound with the structure:

[0231] Compounds;

[0232] E) Then it has a structure:

[0233] The compound reacts with methyl chloroformate and sodium bicarbonate to produce a compound with the structure:

[0234] Compounds;

[0235] F) Then it has a structure:

[0236] The compound reacts with N-bromosuccinimide and an acid to produce a compound with the structure:

[0237] Compounds;

[0238] G) Then it has a structure:

[0239] The compound reacts with 2-amino-4-methylpyridine and optionally sodium borohydride to produce a compound with the structure:

[0240] Compounds;

[0241] H) Then it has a structure:

[0242] The compound reacts with carbonyl diimidazole and methylamine to produce a compound with the structure: Compounds.

[0243] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0244] The compound is formed by including the following structure:

[0245] The compound is prepared by contacting it with a base and optionally sodium borohydride in the presence of a solvent. In some embodiments, the method includes making a compound having the following structure:

[0246] The compound is contacted with a base and sodium borohydride in the presence of a solvent. In some embodiments, the method includes making a compound having the following structure:

[0247] The compound is contacted with a base and without sodium borohydride in the presence of a solvent. In some embodiments, the solvent is an aqueous solution of tetrahydrofuran, dioxane, 2-methyltetrahydrofuran, an aqueous solution of methanol, an aqueous solution of ethanol, or an aqueous solution of acetonitrile. In some embodiments, the solvent is an aqueous solution of tetrahydrofuran. In some embodiments, the base is lithium hydroxide. In some embodiments, the base is sodium hydroxide. In some embodiments, the base is potassium hydroxide.

[0248] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0249] The compound is formed by including the following structure:

[0250] The compound was prepared by contacting it with 2-amino-4-methylpyridine.

[0251] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0252] The compound is formed by including the following structure:

[0253] The compound is prepared by contacting it with a brominating agent. In some embodiments, the brominating agent is copper(II) bromide. In some embodiments, the brominating agent is liquid bromine.

[0254] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0255] The compound is formed by including the following structure:

[0256] The compound is prepared by contacting it with methyl chloroformate and a base. In some embodiments, the base is sodium bicarbonate.

[0257] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0258] The compound is formed by including the following structure:

[0259] The compound was prepared by contacting it with hydrogen chloride and methanol.

[0260] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0261] The compound is formed by including the following structure:

[0262] The compound was prepared by contacting it with hydrogen chloride and ethyl acetate.

[0263] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0264] The compound is formed by including the following structure: The compound is prepared by contacting it with tert-butyl 3,5-difluorobenzoate and a base. In some embodiments, the base is lithium diisopropylamino.

[0265] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0266] The compound is formed by including the following structure:

[0267] The compound is prepared by contacting it with N,O-dimethylhydroxylamine, an amide coupling agent, and 1-hydroxybenzotriazole. In some embodiments, the amide coupling agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide.

[0268] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0269] The compound is formed by including the following structure:

[0270] The compound is prepared by contacting it with a hydrogenation catalyst and hydrogen gas. In some embodiments, the hydrogenation catalyst is carbon-supported palladium, palladium hydroxide, carbon-supported rhodium, alumina-supported rhodium, platinum oxide, or carbon-supported platinum. In some embodiments, the hydrogenation catalyst is carbon-supported palladium.

[0271] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0272] The compound is formed by including the following structure:

[0273] The compound is prepared by contacting a base with an aqueous solution of tetrahydrofuran. In some embodiments, the base is sodium hydroxide.

[0274] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0275] The compound is formed by including the following structure:

[0276] The compound was prepared by contacting it with (ethoxycarbonylmethylene)triphenylphosphine.

[0277] In some embodiments of the method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), the structure is as follows:

[0278] The compound is formed by including the following structure:

[0279] The compound was prepared by contacting it with 2,2,6,6-tetramethylpiperidine 1-oxy group.

[0280] This article further discloses a method for preparing (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1), comprising:

[0281] It has a structure:

[0282] The compound reacts with 2,2,6,6-tetramethylpiperidine 1-oxy to produce a compound with the structure:

[0283] Compounds;

[0284] B) Then it has a structure:

[0285] The compound reacts with (ethoxycarbonylmethylene)triphenylphosphine to produce a compound with the structure:

[0286] Compounds;

[0287] C) Then it has a structure:

[0288] The compound reacts with sodium hydroxide in an aqueous solution of tetrahydrofuran to produce a compound with the structure:

[0289] Compounds;

[0290] D) Then it has a structure:

[0291] The compound reacts with carbon-supported palladium and hydrogen to produce a compound with the structure: Compounds;

[0292] E) Then it has a structure:

[0293] The compound reacts with N,O-dimethylhydroxylamine, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and 1-hydroxybenzotriazole to produce a compound with the structure:

[0294] Compounds;

[0295] F) Then it has a structure:

[0296] The compound reacts with tert-butyl 3,5-difluorobenzoate and lithium diisopropylamino to produce a compound with the structure:

[0297] Compounds;

[0298] G) Then it has a structure:

[0299] The compound reacts with hydrogen chloride and ethyl acetate to produce a compound with the structure:

[0300] Compounds;

[0301] H) Then it has a structure:

[0302] The compound reacts with hydrogen chloride and methanol to produce a compound with the structure: Compounds;

[0303] I) Then it has a structure:

[0304] The compound reacts with methyl chloroformate and sodium bicarbonate to produce a compound with the structure:

[0305] Compounds;

[0306] J) Then it has a structure:

[0307] The compound reacts with copper(II) bromide to produce a compound with the structure: Compounds;

[0308] K) then has a structure:

[0309] The compound reacts with 2-amino-4-methylpyridine to produce a compound with the structure:

[0310] Compounds;

[0311] L) then has a structure:

[0312] The compound reacts with lithium hydroxide in an aqueous tetrahydrofuran solution, and then with sodium borohydride to produce a compound with the structure: Compounds;

[0313] M) then has a structure:

[0314] The compound reacts with carbonyl diimidazole and methylamine to produce a compound with the structure: Compounds.

[0315] Pharmaceutical Compositions and Administration

[0316] The administration of the P2X3 antagonist as described herein can be in any pharmacological form, including a therapeutically effective amount of the P2X3 antagonist alone or in combination with a pharmaceutically acceptable carrier.

[0317] Pharmaceutical compositions can be formulated in a conventional manner using one or more physiologically acceptable carriers, including excipients and adjuvants, which facilitate the processing of the active compound into a pharmaceutically usable formulation. The appropriate formulation depends on the chosen route of administration. Further details regarding suitable excipients for the pharmaceutical compositions described herein can be found, for example, in Remington: The Science and Practice of Pharmacy, 19th edition (Easton, Pa.: Mack Publishing Company, 1995); Hoover, John E., Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania 1975; Liberman, HA and Lachman, L., eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, NY, 1980; and Pharmaceutical Dosage Forms and Drug Delivery Systems, 7th edition (Lippincott Williams & Wilkins 1999), such disclosures being incorporated herein by reference.

[0318] As used herein, a pharmaceutical composition refers to a mixture of Compound 1 described herein with other chemical components (such as carriers, stabilizers, diluents, dispersants, suspending agents, thickeners, and / or excipients). The pharmaceutical composition facilitates the administration of the compound to a living organism. When performing the treatment or methods of use provided herein, a therapeutically effective amount of the compound described herein is administered in the form of a pharmaceutical composition to a mammal suffering from a disease, condition, or ailment to be treated. In some embodiments, the mammal is a human. The therapeutically effective amount can vary considerably depending on the severity of the disease, the age and relative health status of the subject, the efficacy of the compound used, and other factors. Compound 1 may be used alone or in combination with one or more therapeutic agents (as a component of a mixture) (e.g., in combination therapy).

[0319] The pharmaceutical formulations described herein can be administered to the subject via a variety of routes of administration, including but not limited to oral, parenteral (e.g., intravenous, subcutaneous, intramuscular), intranasal, buccal, topical, rectal, or transdermal administration. Furthermore, the pharmaceutical compositions described herein, including compound 1, can be formulated into any suitable dosage form, including but not limited to aqueous oral dispersions, liquids, gels, syrups, elixirs, slurries, suspensions, aerosols, controlled-release formulations, rapidly melting formulations, effervescent formulations, lyophilized formulations, tablets, powders, pills, sugar-coated pills, capsules, delayed-release formulations, sustained-release formulations, pulsatile-release formulations, multi-microparticle formulations, and a mixture of immediate-release and controlled-release formulations.

[0320] In some embodiments, compound 1 is formulated as a tablet. In some embodiments, compound 1 is formulated as a capsule. In some embodiments, compound 1 is formulated as a suspension. In some embodiments, compound 1 is formulated as a capsule-powder formulation. In some embodiments, compound 1 is formulated as a bottle-packaged powder for recombining into a suspension.

[0321] Pharmaceutical compositions including the compounds described herein can be prepared in conventional ways, such as by way of conventional mixing, dissolving, granulation, sugar-coated pelleting, grinding, emulsification, encapsulation, embedding, or compression processes, by way of example only.

[0322] Depending on the pharmacokinetic parameters of the dosage form and the route of administration, it can be repeatedly administered.

[0323] Formulating compositions in unit dosage form is particularly advantageous for ease of administration and dosage uniformity. As used herein, unit dosage form refers to a single, physically discrete unit suitable for use as a mammalian subject to be treated; each unit contains a predetermined amount of active compound calculated to produce the desired therapeutic effect associated with the desired drug carrier. The specification of unit dosage form is determined by and directly depends on: (a) the unique characteristics of Compound 1 and the specific therapeutic effect to be achieved, and (b) the inherent limitations in the technology of compounding such active compound for individual therapeutic sensitivity. The specific dosage can be readily calculated by those skilled in the art, for example, based on the patient's approximate weight or body surface area or volume of body space to be occupied. The dosage will also be calculated based on the specific route of administration chosen. Those skilled in the art typically refine the calculations required to determine the appropriate therapeutic dosage. The precise dosage is determined in conjunction with standard dose-response studies. It should be understood that the amount of composition actually administered will be determined by the physician based on relevant circumstances, including one or more conditions to be treated, the choice of composition to be administered, the individual patient's age, weight and response, the severity of the patient's symptoms, and the chosen route of administration.

[0324] Administration method and treatment regimen

[0325] The compounds described herein can be used to prepare medicaments for regulating P2X3, or for treating diseases or conditions that at least partially benefit from P2X3 regulation. Furthermore, methods for treating any of the diseases or conditions described herein in subjects requiring such treatment involve administering to the subject a therapeutically effective amount of a pharmaceutical composition containing at least one compound described herein, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate or hydrate thereof.

[0326] Compositions containing one or more of the compounds described herein may be used for preventative and / or therapeutic treatment. In therapeutic application, the composition is administered to a patient already suffering from the disease or condition in an amount sufficient to cure or at least partially stop the symptoms of the disease or condition. The effective amount for this use will depend on the severity and course of the disease or condition, prior treatment, the patient's health status, weight, and response to the drug, as well as the judgment of the treating physician.

[0327] In prophylactic use, a composition containing the compounds described herein is administered to a patient who is susceptible to or otherwise at risk of a particular disease, condition, or illness. This amount is defined as the “preventative effective amount or dose.” In this use, the precise amount also depends on the patient’s health condition, weight, etc. When used on a patient, the effective amount for this purpose will depend on the severity and duration of the disease, condition, or illness, prior treatment, the patient’s health condition and response to the drug, and the judgment of the treating physician.

[0328] If a patient’s condition does not improve, the compound may be administered for an extended period of time, including throughout the patient’s life, as determined by the physician, to improve or otherwise control or limit the symptoms of the patient’s disease or condition.

[0329] If the patient's condition does improve, the medication may continue to be administered at the physician's discretion; alternatively, the dosage of the medication being administered may be temporarily reduced or temporarily suspended for a period of time (i.e., a "medication holiday"). The length of a medication holiday can vary between 2 days and 1 year, and examples include 2, 3, 4, 5, 6, 7, 10, 12, 15, 20, 28, 35, 50, 70, 100, 120, 150, 180, 200, 250, 280, 300, 320, 350, or 365 days. Dosage reductions during medication holidays can range from about 10% to about 100%, and by way of example only, include about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100%.

[0330] Once the patient's condition improves, a maintenance dose may be administered if necessary. Subsequently, depending on the symptoms, the dose or frequency, or both, may be reduced to a level that maintains the improvement of the disease, symptoms, or condition. However, in the event of any recurrence of symptoms, the patient may require long-term intermittent treatment.

[0331] The amount of a given agent corresponding to such a quantity will vary depending on factors such as the specific compound, the disease or condition and its severity, and the attributes of the person or host requiring treatment (e.g., weight), but can still be determined in a manner recognized in the art based on the specific circumstances surrounding the case, including, for example, the specific agent being administered, the route of administration, the condition being treated, and the person or host being treated. However, generally, the dosage for adult treatment typically ranges from about 0.01 mg / day to about 5000 mg / day, and in some embodiments, from about 1 mg / day to about 1500 mg / day. The desired dosage can be conveniently provided as a single dose or as concurrently administered fractional doses (or over a short period of time) or at appropriate intervals, such as as sub-dose twice, three, four, or more times daily.

[0332] The pharmaceutical compositions described herein may be unit dosage forms suitable for precise single-dose administration. In a unit dosage form, the formulation is divided into unit doses containing an appropriate amount of one or more compounds. A unit dose may be a packaging form containing discrete quantities of the formulation. Non-limiting examples include packaged tablets or capsules, and powders in vials, capsules, bottles, or ampoules. Aqueous suspension compositions may be packaged in single-dose, non-resealable containers. Alternatively, multi-dose, resealable containers may be used, in which case preservatives are typically included in the composition. By way of example only, parenteral injection formulations may be provided in unit dosage forms, including but not limited to ampoules, or in multi-dose containers with added preservatives.

[0333] Example

[0334] All chemicals, reagents and solvents were purchased from commercial sources (where available) and were ready for use without further purification.

[0335] Example 1: Synthesis of (S)-2-(3-(2,6-difluoro-4-(methoxycarbonyl)phenyl)-3-oxypropyl)morpholine-4-carboxylic acid methyl ester (compound J)

[0336]

[0337] Step 1: Under biphasic conditions (dichloromethane-water), the hydroxymethyl group of A (106 kg, 487.9 mol) was oxidized to the corresponding aldehyde B by reaction with sodium bromide, sodium bicarbonate, catalytic TEMPO (2,2,6,6-tetramethyl-1-piperidinoxy, free radical), and sodium hypochlorite (added dropwise over approximately 10 hours while maintaining a temperature of -3°C to 1.5°C) at a temperature of -3°C to 1.5°C. After an additional 2 hours of stirring, the reaction was quenched using sodium thiosulfate at -5°C to 0°C, followed by stirring for 30 minutes.

[0338] Step 2: The two-phase system containing aldehyde B was treated in batches with commercially available (ethoxycarbonylmethylene)triphenylphosphine at 5°C to 10°C. After stirring for 1 hour at 8°C to 15°C, water was added, the mixture was stirred for 30 minutes, the layers were separated, and the aqueous layer was extracted with additional dichloromethane. The combined organic layers containing unsaturated ester C were washed with brine and concentrated to remove most of the dichloromethane. A mixture of petroleum ether / THF was added, and the resulting mixture was stirred at 20°C for 1 hour. The mixture was then filtered to remove triphenylphosphine oxide, and the filter cake was washed with additional petroleum ether / THF. The filtrate containing C was concentrated and THF was added. The mixture was concentrated again and fresh THF was added. In the following steps, the solution of C was used "as is". The yield of C was 58.2 kg.

[0339] Step 3: Treat the C solution in THF dropwise with 3M NaOH solution over 2 hours at 15°C to 25°C. Then warm the mixture to 25°C to 35°C and stir for 8 hours. Cool the mixture to 20°C to 25°C, add MTBE, and separate the layers. Extract the organic layer with water while keeping the temperature below 15°C, and slowly acidify the combined aqueous layer containing the sodium salt of D with 3N HCl until the pH is 10–11. Then wash the aqueous mixture with dichloromethane to remove any residual triphenylphosphine oxide, and then slowly acidify with 3N HCl to pH 5 while keeping the temperature below 15°C. Extract the resulting mixture with dichloromethane and concentrate the organic extract containing D. Then add THF and evaporate. Dissolve the crude product D in THF and use it directly in the next step.

[0340] Step 4: The solution of D in THF (measured at 43.7 kg) is loaded into the hydrogenation reactor. A THF slurry of Pd / C (2.90 kg) is added, and the resulting mixture is stirred at 25°C to 49°C under hydrogen (approximately 145 psi) for 12 hours. The mixture is filtered under nitrogen, the filter cake is washed with THF, and the filtrate is concentrated. Dichloromethane is added and the mixture is concentrated to remove THF; this process is repeated. Fresh dichloromethane is added to the mixture, and the resulting E solution (based on a measured amount of 43.5 kg) is used directly in the next step.

[0341] Step 5: Treat the solution of E in dichloromethane at 10-15°C with N-hydroxybenzotriazole (HOBT), N,O-dimethylhydroxylamine hydrochloride, and triethylamine. Then, add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI) in portions. Stir the mixture at 15-25°C for 12 hours. Add water, stir the resulting mixture for 12 hours, and separate the layers. Separate the aqueous layer and extract with fresh dichloromethane. Wash the combined organic layers with sodium bicarbonate solution to remove HOBT and dry. Concentrate the dichloromethane, add n-heptane, and concentrate the mixture to remove dichloromethane. Add fresh n-heptane and stir the mixture at 15°C for 10 hours. Filter the solid and dry to give 38.6 kg of F.

[0342] Step 6: The solution of tert-butyl 3,5-difluorobenzoate in THF was cooled to -65°C under nitrogen and treated dropwise with 1.5 equivalents of LDA solution. The mixture was stirred at -60°C to -65°C for 1 hour, and then treated dropwise with a solution of compound F (37 kg) in THF. The reaction was stirred between -65°C and -60°C for 6 hours, and then quenched at -65°C with a solution of acetic acid in THF. The temperature was raised to -33°C, and the mixture was stirred for 30 minutes. Ethyl acetate was added, and the mixture was diluted with brine. The layers were separated, the organic layer was washed with brine, and then concentrated to obtain a solution of compound G in ethyl acetate, which was used directly in the next step.

[0343] Step 7: Bubbling HCl gas (60.4 kg) into ethyl acetate (360 kg) between -6°C and 0°C. Adding compound G to the mixture over 2 hours at a temperature between 20°C and 25°C. The reaction mixture is then stirred for 16 hours, filtered, and the product is washed with ethyl acetate and MTBE, and dried under vacuum to obtain H.

[0344] Step 8: Methanol was loaded into the reactor at 26°C and cooled to -7°C. Then, HCl was bubbled into the methanol over 8 hours at -7°C to 0°C. Compound H (28.8 kg) was added at 2°C, and the mixture was heated to 40°C to 50°C and stirred for 6 hours. The reaction was then concentrated, and the residual dichloromethane solvent was first exchanged for heptane (heptane added, then concentrated), and then exchanged for THF (THF added, then concentrated). The resulting compound I solution was used directly in the next step.

[0345] Step 9: Dilute the solution of compound I (approximately 24.6 kg) in THF with water, cool the mixture to -5°C to 0°C, and adjust the pH to 7-8 using a sodium bicarbonate solution (2.5 equivalents of bicarbonate). Add another 2 equivalents of sodium bicarbonate, and add methyl chloroformate (1.2 equivalents) dropwise over 1.5 hours, stirring at -5°C to 0°C for 1.5 hours. Add water, ethyl acetate, and 2N HCl, separate the layers, wash the organic layer with brine, and then concentrate. Add additional ethyl acetate and evaporate to obtain an ethyl acetate solution of J. While stirring at 55°C, add heptane (4 volumes), cool the mixture to 10°C, and stir for 6 hours. Filter the product, wash with ethyl acetate:heptane (1:4), and dry to give J (20.6 kg).

[0346] The product was further purified by dissolving J (20.6 kg) in ethyl acetate at 28 °C and filtering through a silica gel pad (25 kg). The filtrate was concentrated to approximately 50 L at 40-50 °C, and 50 kg of ethyl acetate:heptane (1:3) was added at 55 °C. After stirring for 1 hour, the mixture was cooled to 10 °C and stirred for 6 hours. The product was filtered, washed with ethyl acetate:heptane (1:3), and dried to give J (18.5 kg).

[0347] Synthesis of tert-butyl 3,5-difluorobenzoate

[0348] A solution of 3,5-difluorobenzoic acid (75 kg) in tert-butanol was treated with DMAP (5.8 kg) and triethylamine (67.2 kg) and cooled to 5 °C. Di-tert-butyl dicarbonate (124 kg, 1.2 equivalents) was added in portions over 3 hours, and the mixture was stirred at 20-25 °C for 12 hours. The mixture was diluted with MTBE and water and stirred for 30 minutes. The organic layer was cooled to 0 °C, acidified with 1.5 M HCl (470 kg), and stirred for 30 minutes. The organic layer was then washed with brine, concentrated to approximately 150 L, and THF (90 kg) was added, followed by concentration. This step was repeated (adding 90 kg of THF and evaporating), and the resulting solution of tert-butyl 3,5-difluorobenzoate in THF was used directly in step 6 (above) to prepare compound G.

[0349] Example 2: Synthesis of (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid methyl ester (compound 1)

[0350] Ethyl acetate was loaded into the reactor and degassed. Compound J (80 g) was added, followed by CuBr2 (101 g). The resulting mixture was stirred at 65°C to 75°C for 15 to 24 hours. If HPLC indicated incomplete reaction, additional CuBr2 was added. The reaction was stirred at 65°C to 75°C for another 3 to 5 hours, then stirred again for 3 to 5 hours, cooled to 20°C to 30°C, water was added, followed by sodium bicarbonate. The resulting mixture was filtered through diatomaceous earth, and the filter cake was washed with ethyl acetate. The organic matter was then washed with 5% disodium EDTA solution to remove copper residues, followed by washing with 1% sodium bicarbonate solution, and then with water to give compound K, which was used directly (“as is”) in the following steps.

[0351] The ethyl acetate solution from above was exchanged with acetonitrile to produce a solution of K (100 g) in acetonitrile. 2-Amino-4-methylpyridine (72.06 g) was added to the solution, and the mixture was stirred under nitrogen at 75°C to 85°C for 30 to 40 hours. The mixture was concentrated to 1–2 volumes below 40°C and diluted with dichloromethane. Water was added, the mixture was cooled to 0°C to 10°C, and acidified to pH 4–5 with 2N HCl. The layers were separated, and the aqueous layer was extracted with additional dichloromethane. The combined organic layers were washed with water at 0°C to 10°C, a 7% sodium bicarbonate solution at 0°C to 10°C, and then washed with water. The organic layers were treated with silica gel, and the mixture was concentrated to dryness below 35°C. The residue was transferred to a silica gel pad eluted with dichloromethane-ethyl acetate (1V / 9V), the fraction containing compound L was concentrated, and diluted with THF. This process was repeated until the residual ethyl acetate was ≤1%.

[0352] Cool the L of THF solution to 15°C to 25°C and treat with 10% LiOH solution, stirring the mixture for 2 to 5 hours. Then, add NaBH4 (2.11 g) in portions to the mixture at 15°C to 25°C and stir the reaction for 2 to 4 hours. Add water dropwise at 0°C to 10°C and dilute the mixture with MTBE. Separate the layers and wash the aqueous layer with fresh MTBE. Cool the aqueous layer to 0°C to 10°C, treat with dichloromethane-methanol (approximately 6-1), and adjust the pH to 4-5 with 2N HCl. Filter the mixture through diatomaceous earth and extract the aqueous layer with fresh dichloromethane-methanol (6-1). Concentrate the combined organic layers to 1-2V at below 35°C and add ethanol (2-3V). Concentrate the solution to 1-2V, treat with ethyl acetate, and concentrate the resulting mixture to 1-2V. Add additional ethanol-ethyl acetate and heat the mixture to 70°C to 85°C for 10 to 30 minutes. The mixture was cooled and stirred at -15°C to 5°C for 2 to 8 hours. The mixture was filtered to give compound M, which was washed with ethyl acetate. Compound M was prepared into a slurry in ethyl acetate and stirred at -15°C to 5°C for 1 to 3 hours. The mixture was filtered, compound M was washed with additional ethyl acetate and dried.

[0353] A solution of M (26 g) in THF (115 mL) was stirred at 20°C to 30°C, and 19 g of CDI (1,1-carbonyldiimidazole) was added. The mixture was stirred for 0.5 h to 1 h, then 7.9 g of methylamine hydrochloride was added in portions, followed by dropwise addition of 18.9 g of diisopropylethylamine. The reaction was stirred at 20°C to 30°C for 2 to 34 h, then diluted dropwise with water while maintaining the temperature. Dichloromethane (173 g) was added, the mixture was stirred, and the layers were separated. The aqueous layer was extracted with fresh dichloromethane, and the combined organic layers were washed twice with 27% ammonium chloride solution, followed by twice with water. The organic layer containing compound 1 was circulated through activated carbon for 1 to 3 h using a CUNO filter. The filtrate was concentrated to 1-2 V at below 35°C, and dichloromethane was exchanged with ethyl acetate by a continuous addition / evaporation operation until the residual dichloromethane was ≤1%. Ethyl acetate (2-4 V) was added. The mixture of compound 1 in ethyl acetate was stirred at 45°C to 55°C for 1 to 2 hours, cooled to 20°C to 30°C, and then stirred for 1 to 2 hours. Compound 1 was filtered and washed with ethyl acetate, and then dried.

[0354] Recrystallization of compound 1

[0355] Compound 1 was treated with water / methanol (1V / 7V; 6.5-7.9 kg) and stirred under nitrogen at 47°C to 55°C for 0.5 to 3 hours until a clear solution was obtained. The solution was polished and filtered, and the original reactor was rinsed with methanol / water. The mixture was heated to 47°C to 55°C and stirred for 10 to 30 minutes to obtain a clear solution. Water (8 kg) was added under nitrogen while maintaining the temperature, and the mixture was allowed to crystallize. The mixture was stirred under nitrogen at 47°C to 55°C for 3 to 6 hours, and then cooled to 22°C to 27°C over 5 hours. The mixture was stirred for 12 to 24 hours and filtered under nitrogen. Compound 1 was washed with water / methanol (2 / 1.8) and dried at 47°C to 53°C.

[0356] Example 3: Substitutional synthesis of methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1)

[0357] A solution of starting material A (30 g, 0.138 mol) in ethyl acetate (100 mL) was treated with DMSO (129 g, 1.66 mol, 117 mL; 12 equivalents) while maintaining a temperature <30 °C. The mixture was cooled to -10 °C under nitrogen atmosphere, and a solution of propanephosphonic anhydride (T3P) (0.221 mol; 1.6 equivalents) in ethyl acetate was added dropwise. The resulting mixture was stirred at -10 °C to 0 °C for 1 hour, and then treated dropwise with N,N-diisopropylethylamine (39.2 g, 0.304 mol; 2.2 equivalents). The resulting mixture was stirred at -10 °C to 0 °C for 18 hours.

[0358] A slurry of potassium bicarbonate (9 equivalents relative to compound A) in water (8 volumes) was cooled to -5°C. The reaction mixture containing aldehyde B was quenched in the bicarbonate solution while maintaining the temperature at -5°C to 5°C. At -5°C to 5°C, a solution of intermediate V (48.7 g, 0.145 mol; 1.05 equivalents), THF (6 V), and 2.5 equivalents of K₂CO₃ (47.7 g, 0.345 mol) in 3 volumes of water was added sequentially. The reaction was stirred at -5°C to 5°C for 5 hours, then at 20°C to 30°C for 5 hours. The reaction was cooled to -5°C to 5°C, and while maintaining the temperature, a 20% aqueous solution of potassium persulfate preparation (oxone, 1 equivalent) was added. The mixture was then stirred for 1 hour while maintaining the temperature at -5°C to 5°C. The pH was then adjusted to 3–5 using 85% H₃PO₄. The reaction mixture was filtered, and the filter cake was washed with fresh ethyl acetate (10 V). The layers were separated, and the aqueous phase was extracted with ethyl acetate (2 x 20 volumes). The organic layers were combined and washed with water (2 x 10 volumes). The organic solution was concentrated to give compound N, which was recrystallized from acetonitrile at -15°C to 5°C to give N (the overall yield of the two steps starting from compound A was 64%).

[0359] A solution of N (25 g) recrystallized under nitrogen atmosphere in 100 mL of THF was stirred. Wet Pd / C (1.25 g) was added, and the resulting mixture was stirred at 40–50 psi H2 for 18 hours at room temperature. The catalyst was filtered and washed with fresh THF. The filtrate was concentrated, and the residual THF solvent was exchanged several times with acetonitrile. A solution of reduced product P in acetonitrile (75 mL) was cooled to -15°C to -5°C and stirred. The crystallized product was filtered and washed with approximately 10 mL of acetonitrile to give P (21.1 g, yield 84%).

[0360] Compound P (130 g) in ethyl acetate was treated with 4 M HCl (5 equivalents of HCl) in ethyl acetate. The reaction was stirred at 15 °C to 25 °C and monitored by HPLC to ensure completion (approximately 2 hours). The solid was filtered, washed with ethyl acetate, and dried at 50 °C for 10 hours to give compound H (108.2 g).

[0361] A solution of compound H (20 g, 0.06 mol) was placed in a flask, followed by the addition of THF (100 mL) and water (30 mL). The mixture was cooled to -5 °C, and sodium bicarbonate (22.5 g, 0.27 mol; 4.5 equivalents) was added. A solution of Moc-Cl (6.77 g, 0.072 mol; 1.2 equivalents) in THF (40 mL) was added dropwise to the mixture over 30 minutes while maintaining the temperature between -5 °C and 0 °C. After the initial 6 was exhausted, the resulting solid was filtered and washed with water (approximately 40 mL), then dried at 45 °C to 55 °C to give the sodium salt of Q (29.55 g). The sodium salt was converted to a carboxylic acid by adding it to water and adjusting the pH to 3 using 3 M HCl. The resulting mixture was extracted with ethyl acetate, and the organic extract was concentrated. Product Q was recrystallized from ethyl acetate:methylcyclohexane (1.5 V: 10 V).

[0362] A solution of compound Q (80 g, based on the starting sodium salt, 0.224 mol) in dichloromethane (400 mL) was prepared. N-bromosuccinimide (NBS; 39.6 g, 0.224 mol) was added at room temperature, and the resulting mixture was cooled to -5 °C. Trifluoromethanesulfonic acid (1 equivalent) was added dropwise, and the mixture was stirred at 25 °C to 30 °C. Additional NBS (0.02 equivalent) was added, and the mixture was stirred for another 1 hour. The reaction was cooled to -5 °C and transferred to a 7% cold aqueous solution of sodium bicarbonate. The resulting pH was 5–6. Ascorbic acid (0.01 equivalent) was added, the layers were separated, and the aqueous layer was extracted with dichloromethane (2 x 250 mL). The combined organic layers were washed with water, and the resulting solution of compound R (with a 101% yield for the determination of the two diastereomers) was used directly in the following steps.

[0363] The solution of R in DCM was concentrated to a small volume, and the DCM was exchanged for acetonitrile (total 10V of acetonitrile). 2-Amino-4-methylpyridine (5 equivalents relative to R) was added, and the reaction was stirred at 50°C for 24 hours, followed by stirring at 80°C for 16 hours. The reaction mixture was concentrated to remove acetonitrile, and a mixture of dichloromethane (8 volumes) and water (3 volumes) was added. The mixture was cooled to 0–10°C, and the pH was adjusted to 2.5 using 2N HCl. The aqueous layer was extracted with dichloromethane, the organic layer was concentrated, and the solvent was exchanged for THF. A 10% aqueous solution of NaOH (1.5 equivalents of NaOH) was added, followed by NaBH4 (0.45 equivalents), and the mixture was stirred at 20°C for 2–5 hours. Water (4 volumes) was added, and the mixture was washed with MTBE. Dichloromethane (5 volumes) was added to the aqueous layer containing the sodium salt of the product, and the pH was adjusted to 5.6–5.8 using 2N HCl. The aqueous layer was extracted with dichloromethane, concentrated, and the solvent was exchanged with ethanol, and concentrated to approximately 0.4 volumes. Two volumes of ethyl acetate were added, the mixture was stirred at 0°C, and filtered to give compound M. Compound M was recrystallized by heating in methanol (5 volumes) at 55–60°C, then adding seed crystals, and then cooling to 0–5°C over 1 hour. Water (10 volumes) was added dropwise at -5 to 5°C, the mixture was stirred for 15 hours, filtered, washed with 1:2 methanol-water, and dried under vacuum at 55–60°C.

[0364] As described in Example 2, compound 1 was prepared from compound M.

[0365] Synthesis of intermediate V

[0366]

[0367] A solution of carboxylic acid S (100 g, 0.387 mol) in 2-methyltetrahydrofuran (400 mL) and DMF (1.61 g, 0.022 mol) was cooled to 10°C to 20°C. The solution was then treated dropwise with oxalyl chloride (61.45 g, 0.48 mol) while maintaining the temperature at 10°C to 20°C, and the resulting mixture was stirred at 10°C to 20°C for 1–3 hours. The resulting acyl chloride T solution was used directly in the next step.

[0368] MgCl2 (46.46 g, 0.488 mol) was placed in a flask containing T, followed by the addition of 2-methyltetrahydrofuran (400 mL). Then, ethyl 2-diethoxyphosphonate (101.06 g, 0.4686 mol) was added to the flask. While maintaining the temperature at 10°C to 25°C, triethylamine (196 g, 1.9365 mol) was added dropwise. The reaction was then stirred at 10°C to 25°C for 1–3 hours. The temperature was then adjusted to 0°C to 10°C, and water (250 mL) was added dropwise while maintaining the temperature below 10°C. The pH was then adjusted to 2.0–4.0 using approximately 275 g of 18% sulfuric acid (while maintaining the temperature below 10°C). Another 250 mL of water was added, and the organic layer was separated. The organic layer was washed with 7% sodium bicarbonate solution, and then concentrated to 2–3 volumes while maintaining the temperature below 40°C. Add toluene (170 mL) and concentrate the resulting solution to 2-3 volumes while maintaining the temperature <40°C. Add acetic acid (944 mL), and use the resulting U-containing solution directly in the next step.

[0369] Trifluoroacetic acid (220.8 g, 1.937 mol) was added dropwise to the U solution prepared in the previous step at <40 °C. The resulting mixture was stirred at 85 °C to 95 °C for 12–24 hours. The mixture was then concentrated to 2–3 volumes at <70 °C. While maintaining the temperature at 20 °C to 30 °C, water (1.44 L) was added dropwise, and the resulting mixture was stirred for 2–4 hours. The product was filtered and washed with water (approximately 140 mL). The product was treated with methyl tert-butyl ether (133 mL), and the resulting slurry was stirred at -20 °C to -5 °C for 1–2 hours. The product was filtered and washed with methyl tert-butyl ether, and then dried at 50 °C to 60 °C to give intermediate V (115 g).

[0370] Example 4: Efficacy and selectivity of human P2X3 and P2X2 / 3 receptors

[0371] The ability of compound 1 described in this article to act as a P2X3 and P2X2 / P2X3 channel antagonist (encoded by the human P2RX2 and P2RX3 genes and stably expressed in HEK293 cells) was evaluated using the Fluo-8 Calcium Kit. Compound 1 was evaluated at twelve concentrations.

[0372] For antagonist efficacy evaluation, cells were pre-incubated with compound 1 for 20 minutes, then stimulated with the P2X3 and P2X2 / P2X3 agonists α,β-methylene ATP (meATP) at final concentrations of 3 μM and 30 μM, respectively. Four minutes and 50 seconds after meATP addition, ionomycin was added at a final concentration of 5 μM to obtain the maximum possible calcium influx and fluorescence signal in the cells. Fluorescence was continuously recorded for 10 minutes, starting 10 seconds before meATP addition. The IC50 values ​​obtained using the above method were... 50This indicates that compound 1 is a selective P2X3 antagonist (P2X3 IC50). 50 =11nM; P2X2 / 3 IC 50 >30μM).

[0373] The embodiments and implementations described herein are for illustrative purposes only, and in some implementations, various modifications or changes will be included within the scope and range of the appended claims.

Claims

1. A method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1): (Compound 1), Includes having a structure: The compound is contacted with an amide coupling agent and methylamine or methylamine salt.

2. The method according to claim 1, wherein the amide coupling agent is carbonyl diimidazole.

3. The method according to claim 1, wherein the amide coupling agent is propanephosphonic anhydride (T3P).

4. The method according to any one of claims 1-3, wherein the method has the following structure: The compound is formed by including the following structure: The compound is prepared by contacting 2-amino-4-methylpyridine with, optionally, sodium borohydride.

5. The method according to claim 4, wherein the structure is: The compound is formed by including the following structure: The compound is prepared by contacting it with a brominating agent.

6. The method according to claim 5, wherein the brominating agent is N-bromosuccinimide in the presence of an acid.

7. The method according to claim 5, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with methyl chloroformate and a base.

8. The method of claim 6, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with methyl chloroformate and a base.

9. The method according to claim 7, wherein the base is sodium bicarbonate.

10. The method according to claim 8, wherein the base is sodium bicarbonate.

11. The method according to any one of claims 7-10, wherein the method has the following structure: The compound is formed by including the following structure: The compound was prepared by contacting it with hydrogen chloride in the presence of a solvent.

12. The method of claim 11, wherein the solvent is ethyl acetate.

13. The method of claim 11, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with a hydrogenation catalyst and hydrogen gas.

14. The method of claim 12, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with a hydrogenation catalyst and hydrogen gas.

15. The method according to claim 13, wherein the hydrogenation catalyst is carbon-supported palladium, palladium hydroxide, carbon-supported rhodium, alumina-supported rhodium, platinum oxide, or carbon-supported platinum.

16. The method according to claim 14, wherein the hydrogenation catalyst is carbon-supported palladium, palladium hydroxide, carbon-supported rhodium, alumina-supported rhodium, platinum oxide, or carbon-supported platinum.

17. The method of claim 15, wherein the hydrogenation catalyst is a carbon-supported palladium.

18. The method of claim 16, wherein the hydrogenation catalyst is a carbon-supported palladium.

19. The method according to any one of claims 13-18, wherein the method has the following structure: The compound is formed by including the following structure: Compounds with the following structures: The compound is prepared by contacting a base with the compound.

20. The method according to claim 19, wherein the base is a mixture of potassium bicarbonate and potassium carbonate.

21. The method of claim 19, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with 2,2,6,6-tetramethylpiperidine 1-oxy or T3P.

22. The method of claim 20, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with 2,2,6,6-tetramethylpiperidine 1-oxy or T3P.

23. The method according to any one of claims 1-3, wherein the method has the following structure: Compounds acquire their structure by including them in the presence of a solvent: The compound is prepared by contacting a base, and optionally by contacting sodium borohydride.

24. The method according to claim 23, wherein the solvent is an aqueous solution of tetrahydrofuran, dioxane, 2-methyltetrahydrofuran, an aqueous solution of methanol, an aqueous solution of ethanol, or an aqueous solution of acetonitrile.

25. The method of claim 23, wherein the solvent is an aqueous solution of tetrahydrofuran.

26. The method of claim 23, wherein the base is lithium hydroxide.

27. The method of claim 24, wherein the base is lithium hydroxide.

28. The method of claim 25, wherein the base is lithium hydroxide.

29. The method of claim 23, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with 2-amino-4-methylpyridine.

30. The method according to any one of claims 24-28, wherein the method has the following structure: The compound is formed by including the following structure: The compound was prepared by contacting it with 2-amino-4-methylpyridine.

31. The method of claim 29, wherein the structure is: The compound is formed by including the following structure: The compound is prepared by contacting it with a brominating agent.

32. The method of claim 30, wherein the structure is: The compound is formed by including the following structure: The compound is prepared by contacting it with a brominating agent.

33. The method of claim 31, wherein the brominating agent is copper(II) bromide.

34. The method of claim 32, wherein the brominating agent is copper(II) bromide.

35. The method of claim 31, wherein the brominating agent is liquid bromine.

36. The method of claim 32, wherein the brominating agent is liquid bromine.

37. The method according to any one of claims 31-36, wherein the method has the following structure: The compound is formed by including the following structure: The compound was prepared by contacting it with methyl chloroformate and a base.

38. The method of claim 37, wherein the base is sodium bicarbonate.

39. The method of claim 38, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with hydrogen chloride and methanol.

40. The method of claim 39, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with hydrogen chloride in the presence of a solvent.

41. The method of claim 40, wherein the solvent is ethyl acetate.

42. The method according to claim 40 or 41, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with tert-butyl 3,5-difluorobenzoate and a base.

43. The method according to claim 42, wherein the base is lithium diisopropylamino.

44. The method of claim 42, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with N,O-dimethylhydroxylamine, an amide coupling agent and 1-hydroxybenzotriazole.

45. The method of claim 43, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with N,O-dimethylhydroxylamine, an amide coupling agent and 1-hydroxybenzotriazole.

46. ​​The method according to claim 44, wherein the amide coupling agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide.

47. The method according to claim 45, wherein the amide coupling agent is 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide.

48. The method according to any one of claims 44-47, wherein the method has the following structure: The compound is formed by including the following structure: The compound was prepared by contacting it with a hydrogenation catalyst and hydrogen gas.

49. The method according to claim 48, wherein the hydrogenation catalyst is carbon-supported palladium, palladium hydroxide, carbon-supported rhodium, alumina-supported rhodium, platinum oxide, or carbon-supported platinum.

50. The method of claim 49, wherein the hydrogenation catalyst is a carbon-supported palladium.

51. The method of claim 48, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting a base with an aqueous solution of tetrahydrofuran.

52. The method of claim 49, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting a base with an aqueous solution of tetrahydrofuran.

53. The method of claim 50, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting a base with an aqueous solution of tetrahydrofuran.

54. The method according to any one of claims 51-53, wherein the base is sodium hydroxide.

55. The method according to any one of claims 51-53, wherein the method has the following structure: The compound is formed by including the following structure: The compound was prepared by contacting it with (ethoxycarbonylmethylene)triphenylphosphine.

56. The method of claim 54, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with (ethoxycarbonylmethylene)triphenylphosphine.

57. The method of claim 55, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with 2,2,6,6-tetramethylpiperidine 1-oxy group.

58. The method of claim 56, wherein the structure is: The compound is formed by including the following structure: The compound was prepared by contacting it with 2,2,6,6-tetramethylpiperidine 1-oxy group.

59. A method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), comprising: A) Has a structure: The compound reacts with 2,2,6,6-tetramethylpiperidine 1-oxy or T3P to produce a compound with the structure: Compounds; B) Then the structure described is: Compounds and having the following structures: The compound reacts with potassium carbonate and potassium bicarbonate to produce a compound with the structure: Compounds; C) Then the structure described is: The compound reacts with carbon-supported palladium and hydrogen to produce a compound with the structure: Compounds; D) Then the structure described is: The compound reacts with hydrogen chloride in ethyl acetate to produce a compound with the structure: Compounds; E) Then the structure described is: The compound reacts with methyl chloroformate and sodium bicarbonate to produce a compound with the structure: Compounds; F) Then the structure described is: The compound reacts with N-bromosuccinimide and an acid to produce a compound with the structure: Compounds; G) Then the structure is described as follows: The compound reacts with 2-amino-4-methylpyridine and optionally with sodium borohydride to produce a compound having the structure: Compounds; H) Then the structure described is: The compound reacts with carbonyl diimidazole and methylamine to produce a compound with the structure: Compounds.

60. A method for preparing methyl (S)-2-((2-(2,6-difluoro-4-(methylcarbamoyl)phenyl)-7-methylimidazo[1,2-a]pyridin-3-yl)methyl)morpholine-4-carboxylic acid (compound 1), comprising: A) Has a structure: The compound reacts with 2,2,6,6-tetramethylpiperidine 1-oxy to produce a compound with the structure: Compounds; B) Then the structure described is: The compound reacts with (ethoxycarbonylmethylene)triphenylphosphine to produce a compound with the structure: Compounds; C) Then the structure described is: The compound reacts with sodium hydroxide in an aqueous solution of tetrahydrofuran to produce a compound with the structure: Compounds; D) Then the structure described is: The compound reacts with carbon-supported palladium and hydrogen to produce a compound with the structure: Compounds; E) Then the structure described is: The compound reacts with N,O-dimethylhydroxylamine, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and 1-hydroxybenzotriazole to produce a compound with the structure: Compounds; F) Then the structure described is: The compound reacts with tert-butyl 3,5-difluorobenzoate and lithium diisopropylamino to produce a compound with the structure: Compounds; G) Then the structure is described as follows: The compound reacts with hydrogen chloride in ethyl acetate to produce a compound with the structure: Compounds; H) Then the structure described is: The compound reacts with hydrogen chloride and methanol to produce a compound with the structure: Compounds; I) Then the structure described is: The compound reacts with methyl chloroformate and sodium bicarbonate to produce a compound with the structure: Compounds; J) Then the structure described is: The compound reacts with copper(II) bromide to produce a compound with the structure: Compounds; K) Then the structure described is: The compound reacts with 2-amino-4-methylpyridine to produce a compound with the structure: Compounds; L) Then the structure described is: The compound reacts with lithium hydroxide in an aqueous tetrahydrofuran solution, and then with sodium borohydride to produce a compound with the structure: Compounds; M) Then the structure described is: The compound reacts with carbonyl diimidazole and methylamine to produce a compound with the structure: Compounds.

61. A compound having the following structure: or its pharmaceutically acceptable salt.

62. A compound having the following structure: or its pharmaceutically acceptable salt.

63. A compound having the following structure: or its pharmaceutically acceptable salt.

64. A compound having the following structure: or its pharmaceutically acceptable salt.

65. A compound having the following structure: or its pharmaceutically acceptable salt.

66. A compound having the following structure: or its pharmaceutically acceptable salt.

67. A compound having the following structure: or its pharmaceutically acceptable salt.

68. A compound having the following structure: or its pharmaceutically acceptable salt.

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