A prolyl hydroxylase inhibitor and its use

Abstract

The present invention discloses a prolyl hydroxylase inhibitor, belonging to the field of medical technology. The present invention provides a pyridotriazole compound, the structure of which is shown in the following formula, which has high prolyl hydroxylase inhibitory activity and EPO induction activity, can effectively treat and prevent HIF-related and / or EPO-related diseases, and provides new ideas for diseases such as anemia, local ischemia and hypoxia.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and specifically relates to a pyridotriazole compound having prolyl hydroxylase inhibitory effect and a pharmaceutical composition thereof, and further relates to a preparation method thereof and pharmaceutical use thereof. Background Art

[0002] In cases of anemia, trauma, tissue necrosis and defect, tissues or cells are often in a state of hypoxia. Hypoxia leads to the expression of a series of transcription induction factors, which are involved in angiogenesis, iron and sugar metabolism, and cell growth and proliferation. Among them, hypoxia inducible factor (HIF) is a transcription factor activated by somatic cells under hypoxia, which responds to cellular hypoxia by mediating a series of gene regulation in biological cells. HIF is a heterodimer containing an oxygen-regulated α-subunit (HIFα) and a constitutively expressed β-subunit (HIFβ / ARNT). In oxygenated (normal oxygen content) cells, the HIFα subunit is rapidly degraded through the mechanism of ubiquitination by the von Hippel-Lindau tumor suppressor (pVHL) E3 ligase complex. Under hypoxic conditions, HIFα is not degraded and the active HIFα / β complex accumulates in the nucleus and activates the expression of various genes, including glycolytic enzymes, glucose transporters, erythropoietin (EPO) and vascular endothelial growth factor (VEGF).

[0003] Erythropoietin (EPO) is a naturally occurring hormone produced with HIFα that stimulates the production of red fat (erythrocytes) that carry oxygen throughout the body. EPO is usually secreted by the kidneys, and endogenous EPO increases under conditions of reduced oxygen (hypoxia). All types of anemia are characterized by a reduced ability of the blood to carry oxygen, and are therefore accompanied by similar signs and symptoms, including pale skin and mucous membranes, weakness, dizziness, fatigue and lethargy, leading to a decrease in quality of life. Anemia is usually associated with conditions in which there is a lack of blood in red blood cells or in hemoglobin. Common causes of anemia include iron, vitamin B12 and folic acid deficiencies, and are also complicated by chronic diseases, such as inflammatory diseases, including diseases with secondary bone marrow inflammatory suppression, etc. Anemia is also associated with renal dysfunction, and most renal failure patients who are often dialyzed suffer from chronic anemia.

[0004] Prolyl hydroxylase domain (PHD) is a key factor in regulating HIF. Under normoxic conditions, PHD can hydroxylate two key proline residues Pro402 and Pro564 of HIFα, increase its affinity with pVHL, and accelerate the degradation process. Under hypoxia and other pathological conditions, the HIF reaction catalyzed by PHD is blocked, the degradation rate of proteases is slowed down, causing HIFα to accumulate in cells, and then causing a series of adaptive responses of cells to hypoxia. Inhibiting PHD through PHD inhibitors, prolonging the action of HIF, and then increasing the expression of genes such as EPO, can effectively treat and prevent HIF-related and / or EPO-related diseases, such as anemia, local ischemia and hypoxia.

[0005] For example, US patent application US16757333 discloses a crystalline form of an alkynylpyridine aminoacyl hydroxylase inhibitor and a preparation method thereof. The alkynylpyridine aminoacyl hydroxylase inhibitor has the following structural formula:

[0006]

[0007] Due to the limited development and high demand for co-aminoacyl hydroxylase inhibitors, there is an urgent need to develop such compounds for the treatment of conditions such as anemia, local ischemia and hypoxia. Summary of the invention

[0008] The purpose of the present invention is to provide a pyridotriazole compound and a pharmaceutical composition thereof as a HIF-PHD inhibitor, which can be used to treat various HIF-related or EPO-related diseases, such as anemia.

[0009] To achieve the above-mentioned purpose of the invention, the technical solution of the present invention is as follows:

[0010] In one aspect, the present invention provides a compound represented by formula (I) or its stereoisomers, geometric isomers, tautomers, nitrogen oxides, hydrates, solvates, pharmaceutically acceptable salts or prodrugs;

[0011]

[0012] in,

[0013] R is selected from the following atoms or groups:

[0014]

[0015] L is -CH2- or -CH2O-;

[0016] The value of n is an integer between 0 and 2;

[0017] R 1 , R2 , R 3 Each is independently selected from substituted or unsubstituted alkyl, cycloalkyl, aryl, and heterocyclic groups.

[0018] Preferably, the aryl group includes an aromatic heterocycle.

[0019] In some embodiments,

[0020] L is -CH2- or -CH2O-;

[0021] The value of n is 0 or 1;

[0022] R 1 is substituted or unsubstituted C 1-4 Alkyl, phenyl, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, 5-6 membered heterocyclic group containing oxygen and / or nitrogen, the substituent is C 1-4 Alkyl, C 1-4 Alkoxy, C 1-4 At least one of a haloalkyl group, a halogen group, a cyano group, a phenyl group, a 5-6-membered aromatic heterocyclic group containing oxygen and / or nitrogen, and a 5-6-membered heterocyclic group containing oxygen and / or nitrogen;

[0023] R 2 is substituted or unsubstituted C 1-4 Alkyl, phenyl, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, 5-6 membered heterocyclic group containing oxygen and / or nitrogen, the substituent is C 1-4 Alkyl, C 1-4 Alkoxy, C 1-4 At least one of a haloalkyl group, a halogen group, a cyano group, a phenyl group, a 5-6-membered aromatic heterocyclic group containing oxygen and / or nitrogen, and a 5-6-membered heterocyclic group containing oxygen and / or nitrogen;

[0024] R 3 is substituted or unsubstituted C 1-10 Alkyl, phenyl, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, 5-6 membered heterocyclic group containing oxygen and / or nitrogen, the substituent is C 1-4 Alkyl, C 1-4 Alkoxy, C 1-4 At least one of a haloalkyl, a halogen, a cyano, a phenyl, or a 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or a 5-6 membered heterocyclic group containing oxygen and / or nitrogen.

[0025] L is -CH2- or -CH2O-;

[0026] The value of n is 0 or 1;

[0027] R 1 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-4 Alkyl, C 1-4 Alkoxy, C1-4 At least one of a haloalkyl group, a halogen group, a cyano group, a phenyl group, a 5-6-membered aromatic heterocyclic group containing oxygen and / or nitrogen, or a 5-6-membered heterocyclic group containing oxygen and / or nitrogen;

[0028] R 2 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-4 Alkyl, C 1-4 Alkoxy, C 1-4 At least one of a haloalkyl group, a halogen group, a cyano group, a phenyl group, a 5-6-membered aromatic heterocyclic group containing oxygen and / or nitrogen, and a 5-6-membered heterocyclic group containing oxygen and / or nitrogen;

[0029] R 3 is substituted or unsubstituted C 1-10 Alkyl, phenyl, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, 5-6 membered heterocyclic group containing oxygen and / or nitrogen, the substituent is C 1-4 Alkyl, C 1-4 At least one of an alkoxy group, a halogen group, a 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, and a 5-6 membered heterocyclic group containing oxygen and / or nitrogen.

[0030] In some embodiments,

[0031] The value of n is 0;

[0032] R 1 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-4 Alkyl, C 1-4 alkoxy, halogen, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-4 Alkyl, C 1-4 Alkoxy, oxygen and / or nitrogen-containing 5-6 membered aromatic heterocyclic group or oxygen and / or nitrogen-containing 5-6 membered heterocyclic group are para-substituted relative to the parent core structure, and halogen is ortho-, meta- or para-substituted relative to the parent core structure;

[0033] R 2 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-4 Alkyl, C 1-4 alkoxy, halogen or 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-4 Alkyl, C 1-4 The alkoxy group and the 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen or the 5-6 membered heterocyclic group containing oxygen and / or nitrogen are para-substituted relative to the parent core structure, and the halogen is ortho-, meta- or para-substituted relative to the parent core structure;

[0034] R 3 is substituted or unsubstituted C 1-5Alkyl, phenyl, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen, wherein the substituent is C 1-4 Alkyl, C 1-4 alkoxy, halogen, 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-4 Alkyl, C 1-4 The alkoxy group, the 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen, or the 5-6 membered heterocyclic group containing oxygen and / or nitrogen is para-substituted relative to the parent core structure, and the halogen is ortho-, meta- or para-substituted relative to the parent core structure.

[0035] In some embodiments,

[0036] The value of n is 0;

[0037] R 1 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-3 Alkyl, C 1-3 alkoxy, halogen, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-3 Alkyl, C 1-3 Alkoxy, oxygen and / or nitrogen-containing 5-6 membered aromatic heterocyclic group or oxygen and / or nitrogen-containing 5-6 membered heterocyclic group are para-substituted relative to the parent core structure, and halogen is ortho-, meta- or para-substituted relative to the parent core structure;

[0038] R 2 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-3 Alkyl, C 1-3 alkoxy, halogen, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-3 Alkyl, C 1-3 Alkoxy, oxygen and / or nitrogen-containing 5-6 membered aromatic heterocyclic group or oxygen and / or nitrogen-containing 5-6 membered heterocyclic group are para-substituted relative to the parent core structure, and halogen is ortho-, meta- or para-substituted relative to the parent core structure;

[0039] R 3 is substituted or unsubstituted C 1-5 Alkyl, phenyl, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen, wherein the substituent is C 1-3 Alkyl, C 1-3 alkoxy, halogen, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-3 Alkyl, C 1-3The alkoxy group, the 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen or the 5-6 membered heterocyclic group containing oxygen and / or nitrogen is para-substituted relative to the parent core structure, and the halogen is ortho-, meta- or para-substituted relative to the parent core structure.

[0040] In some embodiments,

[0041] The value of n is 0;

[0042] R 1 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-2 Alkyl, C 1-2 alkoxy, F, Cl, Br, I, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-2 Alkyl, C 1-2 The alkoxy group, the 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or the 5-6 membered heterocyclic group containing oxygen and / or nitrogen is para-substituted relative to the parent core structure, and F, Cl, Br, and I are ortho-, meta-, or para-substituted relative to the parent core structure;

[0043] R 2 is a substituted or unsubstituted phenyl group, wherein the substituent is C 1-2 Alkyl, C 1-2 alkoxy, F, Cl, Br, I, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-2 Alkyl, C 1-2 The alkoxy group, the 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or the 5-6 membered heterocyclic group containing oxygen and / or nitrogen is para-substituted relative to the parent core structure, and F, Cl, Br, and I are ortho-, meta-, or para-substituted relative to the parent core structure;

[0044] R 3 is substituted or unsubstituted C 1-4 Alkyl, phenyl, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen, wherein the substituent is C 1-2 Alkyl, C 1-2 alkoxy, F, Cl, Br, I, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen, or 5-6 membered heterocyclic group containing oxygen and / or nitrogen; preferably, the C 1-2 Alkyl, C 1-2 The alkoxy group, the 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen or the 5-6 membered heterocyclic group containing oxygen and / or nitrogen is para-substituted relative to the parent core structure, and the halogen is ortho-, meta- or para-substituted relative to the parent core structure.

[0045] In some embodiments,

[0046] The value of n is 0;

[0047] R 1 is a substituted or unsubstituted phenyl group, wherein the substituent is a methyl group, a methoxy group, a F group, a Cl group, a 5-6-membered aromatic heterocyclic group containing oxygen and / or nitrogen, or a 5-6-membered heterocyclic group containing oxygen and / or nitrogen; preferably, the methyl group, the methoxy group, the 5-6-membered aromatic heterocyclic group containing oxygen and / or nitrogen is para-substituted relative to the parent core structure, and the F group and the Cl group are ortho-, meta- or para-substituted relative to the parent core structure;

[0048] R 2 is a substituted or unsubstituted phenyl group, wherein the substituent is a methyl group, a methoxy group, a F group, a Cl group or a 5-6-membered aromatic heterocycle containing oxygen and / or nitrogen; preferably, the methyl group, the methoxy group and the 5-6-membered aromatic heterocycle containing oxygen and / or nitrogen are para-substituted relative to the parent core structure, and the F group and the Cl group are ortho-, meta- or para-substituted relative to the parent core structure;

[0049] R 3 is substituted or unsubstituted C 1-3 Alkyl, phenyl, 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen, the substituent of the substituted phenyl is methyl, methoxy, F, Cl or 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen; preferably, the methyl, methoxy and 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen are para-substituted relative to the parent core structure, and the halogen is ortho-, meta- or para-substituted relative to the parent core structure.

[0050] In some embodiments,

[0051] The value of n is 0;

[0052] R 1 is a substituted or unsubstituted phenyl group, wherein the substituent is a methyl group or a 5-6-membered aromatic heterocycle containing oxygen and / or nitrogen; preferably, the methyl group, the 5-6-membered aromatic heterocycle containing oxygen and / or nitrogen is para-substituted relative to the parent core structure;

[0053] R 2 is a substituted or unsubstituted phenyl group, wherein the substituent is a methyl group or a 5-6-membered aromatic heterocycle containing oxygen and / or nitrogen; preferably, the methyl group, the 5-6-membered aromatic heterocycle containing oxygen and / or nitrogen is para-substituted relative to the parent core structure;

[0054] R 3 is substituted or unsubstituted C 1-3 Alkyl, phenyl, 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen, the substituent is methyl, methoxy, F, Cl or 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen; preferably, the methyl, methoxy and 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen are para-substituted relative to the parent core structure, and the halogen is ortho-, meta- or para-substituted relative to the parent core structure.

[0055] In some embodiments,

[0056] The value of n is 0;

[0057] R 1 is phenyl;

[0058] R 2 is phenyl;

[0059] R 3 is substituted or unsubstituted C 1-3 Alkyl, phenyl, 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen, the substituent is methyl, methoxy, halogen or 5-6 membered aromatic heterocycle containing oxygen and / or nitrogen; preferably, the methyl, methoxy, 5-6 membered aromatic heterocyclic group containing oxygen and / or nitrogen or 5-6 membered heterocyclic group containing oxygen and / or nitrogen is para-substituted relative to the parent core structure, and the halogen is ortho-, meta- or para-substituted relative to the parent core structure.

[0060] The term "core structure" refers to the remaining main structure of the compound structure represented by formula (I) except the R group.

[0061] In some embodiments, the compound is selected from the following structures:

[0062]

[0063] In some embodiments, the compound is selected from the following structures:

[0064]

[0065]

[0066] In some embodiments, the compound is selected from the following structures:

[0067]

[0068] In some embodiments, the compound is selected from the following structures:

[0069]

[0070] In the present invention, FG-4592 is rosuvastatin.

[0071] Unless otherwise specified, the term "alkyl" as used herein includes branched and straight chain saturated aliphatic hydrocarbon groups having a specified number of carbon atoms, including all isomers. Common abbreviations for alkyl groups, such as methyl, can be represented by "Me" or CH3, ethyl, can be represented by "Et" or CH2CH3, propyl, can be represented by "Pr" or CH2CH2CH3, butyl, can be represented by "Bu" or CH2CH2CH2CH3, etc. For example, "C 1-4 "C1-C4 alkyl" refers to a straight or branched chain alkyl group having a specified number of carbon atoms, including all isomers.1-4 Alkyl includes n-, iso-, sec- and tert-butyl, n- and iso-propyl, ethyl and methyl. 1-10 "Alkyl" and the like have similar meanings.

[0072] The term "alkoxy" refers to straight and branched chain alkyl groups having the indicated number of carbon atoms attached through an oxygen bridge.

[0073] The term "halogen" (or "halo") refers to fluorine, chlorine, bromine, and iodine (alternatively referred to as fluoro (F), chloro (Cl), bromo (Br), and iodo (I)).

[0074] The term "aryl" refers to aromatic mono- and polycyclic carbon ring systems wherein the individual carbon rings in the polycyclic system are fused or linked to each other by single bonds. Typical aryl groups include phenyl, naphthyl and biphenylene.

[0075] The term "heterocycle" refers to a ring structure composed of carbon atoms and non-carbon atoms, such as nitrogen, oxygen and sulfur. Typical heterocyclic groups include pyridine, quinoline, tropane, phenothiazine, benzodiazepine, furan, pyrazolone and pyrimidine.

[0076] The term "aromatic heterocycle" refers to a 5- or 6-membered monocyclic aromatic ring or a 7- to 12-membered bicyclic ring consisting of carbon atoms and one or more heteroatoms selected from N, O and S. Examples of aromatic heterocycles include pyridyl, pyrrolyl, pyrazinyl, pyrimidinyl, pyridazinyl, thienyl (or thiophenyl), thiazolyl, furanyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, isothiazolyl and thiadiazolyl, benzotriazolyl, indolyl, isoindolyl, indazolyl, indolinyl, isoindolinyl, quinoxalinyl, quinazolinyl, cinnolinyl, chromanyl, isochromanyl, tetrahydroquinolyl, quinolyl, tetrahydroisoquinolyl, isoquinolyl, 2,3-dihydrobenzofuranyl, 2,3-dihydrobenzo-1,4-dienyl, imidazo(2,1-b)(1,3)thiazole and benzo-1,3-dioxolyl.

[0077] The aryl group in the term "substituted aryl" is as defined above. When the substituent of the substituted aryl group is not specified, the substituent may be selected from the following groups, including but not limited to: halogen, C1-C 20 Alkyl, CF3, NH2, N(C1-C6 alkyl)2, NO2, oxo, CN, N3, -OH, -O(C1-C6 alkyl), C3-C 10 Cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, (C0-C6 alkyl)S(O) 0-2 -, Aryl-S(O) 0-2 -, (C0-C6 alkyl) S (O) 0-2(C0-C6 alkyl)-, (C0-C6 alkyl)C(O)NH-, H2N-C(NH)-, -O(C1-C6 alkyl)CF3, (C0-C6 alkyl)C(O)-, (C0-C6 alkyl)OC(O)-, (C0-C6 alkyl)2NC(O)-(C0-C6 alkyl)O(C1-C6 alkyl)-, (C0-C6 alkyl)C(O) 1-2 (C0-C6 alkyl)-, (C0-C6 alkyl)OC(O)NH-, aryl, aralkyl, heteroaryl, heterocyclylalkyl, halogen-aryl, halogen-aralkyl, halogen-heterocycle, halogen-heterocyclylalkyl, cyano-aryl, cyano-aralkyl, cyano-heterocycle and cyano-heterocyclylalkyl. The term "substituted phenyl" has a similar definition.

[0078] Unless explicitly stated, all ranges listed herein are inclusive. For example, "the value of n is an integer between 0 and 2" means that n can be 0, 1 or 2.

[0079] The term "pharmaceutically acceptable salt" refers to a salt prepared from a pharmaceutically acceptable non-toxic base or acid. When the compound of the present invention is acidic, its corresponding salt can be easily prepared from an inorganic base or an organic base. Salts derived from such inorganic bases include salts of aluminum, ammonium, calcium, copper (copper and cuprous), iron, ferrous, lithium, magnesium, manganese (manganese and manganous), potassium, sodium, zinc, etc. Preferred are salts of ammonium, calcium, magnesium, potassium and sodium. Salts prepared from organic bases include primary, secondary and tertiary amines derived from natural and synthetic sources. Pharmaceutically acceptable organic non-toxic bases that can form salts include arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucosamine, glucosamine, histidine, hydrabamine, isopropylamine, dicyclohexylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, etc. When the compound of the present invention is basic, its corresponding salt can be easily prepared from inorganic or organic acids. Such acids include, for example, acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethanesulfonic acid, fumaric acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid, isethionic acid, lactic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, mucic acid, nitric acid, pamoic acid, pantothenic acid, phosphoric acid, succinic acid, sulfuric acid, tartaric acid, p-toluenesulfonic acid, and the like.

[0080] The term "solvate" refers to a complex of variable stoichiometry formed by a solute (i.e., a compound of Formula I) or a pharmaceutically acceptable salt thereof and a solvent that does not interfere with the biological activity of the solute. Examples of solvents include, but are not limited to, water, ethanol, and acetic acid. When the solvent is water, the solvate is referred to as a hydrate. Hydrates include, but are not limited to, hemihydrates, monohydrates, one and a half hydrates, dihydrates, and trihydrates.

[0081] The term "prodrug" is a functional derivative of the compounds of the invention which is readily convertible in vivo into the desired compound.

[0082] On the other hand, the present invention provides a method for preparing the above-mentioned compound, comprising the following three synthetic routes:

[0083] Route 1:

[0084]

[0085] Route 2:

[0086]

[0087] Route 3:

[0088]

[0089] Among them, R 1 , R 2 , R 3 Has the same definition as above.

[0090] In another aspect, the present invention provides a pharmaceutical composition comprising a compound represented by formula (I) or its stereoisomers, geometric isomers, tautomers, nitrogen oxides, hydrates, solvates, pharmaceutically acceptable salts or prodrugs, and one or more pharmaceutically acceptable carriers, diluents, and excipients.

[0091] In pharmaceutical compositions, the term "composition" includes products comprising active ingredients and inert ingredients constituting carriers (pharmaceutically acceptable excipients), as well as any product obtained directly or indirectly by the combination, complexation or aggregation of two or more ingredients, or the decomposition of one or more ingredients, or other types of reactions or interactions of one or more ingredients. Therefore, the pharmaceutical compositions of the present invention include any composition prepared by mixing a compound of formula I, other active ingredients and a pharmaceutically acceptable excipient.

[0092] The pharmaceutical composition of the present invention comprises the compound shown in the formula I as active ingredient (or its pharmaceutically acceptable salt or solvate), pharmaceutically acceptable carrier and optional other therapeutic ingredients or adjuvant.The pharmaceutical composition includes compositions suitable for oral, rectal, topical and parenteral (including subcutaneous, intramuscular and intravenous) administration, although the most suitable approach in any particular case depends on the nature and severity of the disease to which the active ingredient is given.The pharmaceutical composition can be prepared by any method known to the field of pharmacy.

[0093] The active ingredient can be administered orally in a solid dosage form or a liquid dosage form, such as capsules, tablets, lozenges, lozenges, granules and powders, and the liquid dosage form is such as elixirs, syrups, emulsions, dispersions and suspensions. The active ingredient can also be administered parenterally in a sterile liquid dosage form such as a dispersion, suspension or solution. Other dosage forms that can be used to administer the active ingredient include ointments, creams, drops, transdermal patches or powders for topical administration; ophthalmic solutions or suspension forms for eye administration, i.e., eye drops; sprays or powder compositions for inhalation or intranasal administration, or creams, ointments, sprays or suppositories for rectal or vaginal administration. Gelatin capsules contain active ingredients and powdered carriers, such as lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, etc. Similar diluents can be used to prepare compressed tablets. Both tablets and capsules can be prepared as sustained-release products to provide sustained release of drugs over several hours. Compressed tablets can be coated with sugar or film to cover any unpleasant taste and protect the tablet from air, or can be enteric coated for selective disintegration in the gastrointestinal tract. Liquid dosage forms for oral administration can include colorants and flavoring agents to increase patient acceptance. Generally speaking, water, suitable oil, saline, dextrose (glucose) aqueous solution and related sugar solution and glycols such as propylene glycol or polyethylene glycol are suitable carriers of parenteral solutions. The solution for parenteral administration preferably includes water-soluble salts of active ingredients, suitable stabilizers and buffer substances used as required. Antioxidants such as sodium bisulfite, sodium sulfite or ascorbic acid alone or in combination are suitable stabilizers. Citric acid and its salts and sodium EDTA can also be used. In addition, parenteral solutions can also include preservatives, such as benzalkonium chloride, methylparaben or propylparaben and chlorobutanol. For inhalation administration, the compound of the present invention can be conveniently delivered in aerosol form from a pressurized package or a sprayer. The compound can also be delivered in the form of a powder for preparation, and the powder composition can be sucked with the help of a powder inhaler device. The preferred delivery system for suction is a metered dose inhalation (MDI) aerosol, which can be formulated into a suspension or solution of the compound of formula I in a suitable propellant, such as a fluorocarbon or hydrocarbon. For eye administration, ophthalmic preparations can be prepared with a solution or suspension of a suitable weight percentage of the compound of formula I in a suitable ophthalmic carrier, thereby keeping the compound in contact with the surface of the eye enough time to allow the compound to penetrate into the cornea and the inner area of ​​the eye.

[0094] Useful pharmaceutical dosage forms for administering the compounds of the present invention include, but are not limited to, hard and soft gelatin capsules, tablets, parenteral injection solutions, and oral suspensions.

[0095] When the compounds of the present invention are administered stepwise or in combination with other therapeutic agents, the same dosage forms as described above may be used. When the drugs are administered in physical combination, the dosage form and route of administration should be selected based on the compatibility of the combined drugs. The compounds of the present invention may be administered as the sole active ingredient or in combination with a second active ingredient, including active ingredients known to be useful for increasing erythropoietin levels in patients.

[0096] In another aspect, the present invention provides a compound represented by formula (I) or its stereoisomers, geometric isomers, tautomers, nitrogen oxides, hydrates, solvates, pharmaceutically acceptable salts or prodrugs, or the use of the above-mentioned pharmaceutical composition in the preparation of a drug for treating diseases mediated by the inhibition of co-aminoacyl hydroxylases by inhibiting co-aminoacyl hydroxylases.

[0097] In some embodiments, the disease mediated by inhibition of coenzyme A hydroxylase is anemia, ischemia, or hypoxia.

[0098] In some embodiments, the anemia is renal anemia.

[0099] The beneficial effects of the present invention are:

[0100] A new compound is proposed, which can be used as an inhibitor of aminoacyl hydroxylase, has high aminoacyl hydroxylase inhibitory activity and EPO induction activity, can effectively treat and prevent HIF-related and / or EPO-related diseases, and provides new ideas for diseases such as anemia, local ischemia and hypoxia. DETAILED DESCRIPTION

[0101] The following non-limiting examples can enable those skilled in the art to more fully understand the present invention, but do not limit the present invention in any way. The following content is merely an exemplary description of the scope of protection claimed in this application, and those skilled in the art can make various changes and modifications to the invention of this application based on the disclosed content, and they should also fall within the scope of protection claimed in this application.

[0102] The present invention is further described below by way of specific examples. Unless otherwise specified, the various chemical reagents used in the examples of the present invention are obtained through conventional commercial channels.

[0103] Example 1

[0104] Preparation of (7-hydroxy-5-(phenoxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid

[0105]

[0106] Step 1 Preparation of 5-(phenoxy)-7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester 5-iodo-7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester (1.0 g), phenol (310 mg), 2-picolinic acid (55 mg), cuprous iodide (42 mg), potassium phosphate (942 mg), dimethyl sulfoxide (5.0 mL) were added to the reaction bottle in sequence, and nitrogen was replaced three times. The temperature was raised to 65 ° C and reacted overnight until the reaction of the raw materials was complete as monitored by TLC. After cooling to room temperature, the mixture was extracted with ethyl acetate, washed with saturated brine, the organic phase was separated, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and column chromatography was performed to obtain 450 mg of a white solid.

[0107] MS m / z(ESI)[M+H] + :418.30

[0108] Step 2 Preparation of 5-(phenoxy)-7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid

[0109] Dissolve tert-butyl 5-(phenoxy)-7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylate (450 mg) in a mixed solution of toluene and tetrahydrofuran (10 mL) in a volume ratio of 1:1, then add methanesulfonic acid (0.5 mL) and stir, keeping the reaction temperature between 60-70°C. Stop the reaction until the reaction of the raw material is complete as monitored by TLC. Cool to room temperature, add a small amount of ethyl acetate, filter, and obtain 440 mg of a white solid.

[0110] MS m / z(ESI)[M+H] + :362.22

[0111] Step 3 Preparation of [(7-(benzyloxy)-5-(phenoxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid methyl ester

[0112] 5-(Phenoxy)-7-(Benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid (440 mg), EDCI (280 mg), HoBt (197 mg), N,N-dimethylformamide (5.0 mL), triethylamine (247 mg) were added to the reaction bottle in sequence, stirred at room temperature for 10 minutes, then glycine methyl ester hydrochloride (306 mg) was added to the system, and the temperature was raised to 60°C for 1.5 hours. The reaction was stopped, cooled to room temperature, saturated sodium bicarbonate aqueous solution was added to quench the reaction, and 300 mg of yellow solid was obtained by suction filtration.

[0113] MS m / z(ESI)[M+H] +:433.31

[0114] Step 4 Preparation of [(7-hydroxy-5-(phenoxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid methyl ester

[0115] Methyl [(7-(benzyloxy)-5-(phenoxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetate (300 mg) and palladium on carbon (30 mg) were dissolved in a mixed solvent (4.0 mL) of methanol and tetrahydrofuran in a volume ratio of 1:1, and stirred at room temperature under a hydrogen atmosphere until the reaction of the raw material was complete as monitored by TLC, and the reaction was stopped. The solvent was removed by concentration under reduced pressure, and the product was separated by column chromatography to obtain 170 mg.

[0116] MS m / z(ESI)[M+H] + :343.22

[0117] Step 5 Preparation of [(7-hydroxy-5-(phenoxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid

[0118] Methyl [(7-hydroxy-5-(phenoxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetate (170 mg) was dissolved in methanol (2.0 mL), followed by addition of 30% aqueous sodium hydroxide solution (1.0 mL). The mixture was stirred at room temperature until the reaction of the raw material was complete as monitored by TLC. The pH was adjusted to between 1 and 2 with dilute hydrochloric acid and filtered to obtain 150 mg of the product.

[0119] 1H NMR (400MHz, d-DMSO) δ: 14.52 (s, 1H), 12.95 (brs, 1H), 9.71 (s, 1H), 8.59 (s, 1H),7.59–7.53(m,2H),7.44–7.41(m,3H),5.83(s,1H),4.02(d,J=5.6Hz,2H)

[0120] MS m / z(ESI):329.09

[0121] Example 2

[0122] Preparation of (5-(2-chlorophenoxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0123]

[0124] According to the method of Example 3, 428.1 mg of 2-chlorophenol was used to replace phenol to obtain 108.9 mg of the product.

[0125] 1H NMR (400MHz, d-DMSO) δ: 14.58 (s, 1H), 12.97 (brs, 1H), 9.71 (t, J = 5.6Hz, 1H), 8.63 (s, 1H), 7.75 (d, J = 7.6Hz ,1H),7.61(d,J=8.0Hz,1H),7.54(t,J=8.0Hz,1H),7.47(t,J=7.2Hz,1H),5.89(s,1H),4.21(d,J=5.6Hz,2H)

[0126] MS m / z(ESI)[M+H] + :363.17

[0127] Example 3

[0128] Preparation of (5-(3-chlorophenoxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0129]

[0130] According to the method of Example 3, 428.1 mg of 3-chlorophenol was used to replace phenol to obtain 201.6 mg of product.

[0131] 1H NMR (400MHz, d-DMSO) δ: 14.54(s,1H),12.97(brs,1H),9.72(s,1H),8.58(s,1H),7.62(s,1H),7.5 7(t,J=8.4Hz,1H),7.47(d,J=8.4Hz,1H),7.41(d,J=8.4Hz,1H),6.10(s,1H),4.21(d,J=5.6Hz,2H)

[0132] MS m / z(ESI)[M+H] + :363.14

[0133] Example 4

[0134] Preparation of (5-(4-chlorophenoxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0135]

[0136] According to the method of Example 3, 428.1 mg of 4-chlorophenol was used to replace phenol to obtain 182.1 mg of product.

[0137] 1H NMR (400MHz, d-DMSO) δ: 14.54 (s, 1H), 12.96 (s, 1H), 9.72 (t, J = 5.6Hz, 1H), 8.60 (s, 1H),7.62(d,J=8.4Hz,2H),7.48(d,J=8.4Hz,2H),6.02(s,1H),4.22(d,J=5.6Hz,2H)

[0138] MS m / z(ESI)[M+H] + :363.14

[0139] Example 5

[0140] Preparation of (5-(p-methylphenyloxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0141]

[0142] According to the method of Example 3, 356.9 mg of 4-methylphenol was used to replace phenol to obtain 87.7 mg of the product.

[0143] 1H NMR (400MHz, d-DMSO) δ: 14.51 (s, 1H), 12.95 (s, 1H), 9.69 (t, J = 5.6Hz, 1H), 8.59 (s, 1H), 7. 37(d,J=8.4Hz,2H),7.31(d,J=8.4Hz,2H),5.77(s,1H),4.20(d,J=5.6Hz,2H),2.37(s,3H)

[0144] MS m / z(ESI)[M+H] + :343.16

[0145] Example 6

[0146] Preparation of (5-(p-methoxyphenoxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0147]

[0148] According to the method of Example 3, 4-methoxyphenol (409.2 mg) was used to replace phenol to obtain 223.7 mg of the product.

[0149] 1H NMR (400MHz, d-DMSO) δ: 14.51 (s, 1H), 12.96 (s, 1H), 9.70 (t, J = 5.2Hz, 1H), 8.60 (s, 1H), 7. 39(d,J=8.8Hz,2H),7.10(d,J=8.8Hz,2H),5.72(s,1H),4.21(d,J=5.6Hz,2H),3.82(s,3H)

[0150] MS m / z(ESI)[M+H] + :359.22

[0151] Example 7

[0152] Preparation of (5-(3-morpholinophenoxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0153]

[0154] According to the method of Example 3, 3-morpholinophenol (590 mg) was used to replace phenol to obtain 110.7 mg of the product.

[0155] 1H NMR (400MHz, d-DMSO) δ: 14.51 (s, 1H), 12.95 (brs, 1H), 9.70 (t, J = 5.2Hz, 1H), 8.59 (s, 1H), 7.39 (d, J = 8.0Hz, 2H), 6.99 ( d,J=12.0Hz,2H),6.80(d,J=7.6Hz,2H),5.82(s,1H),4.21(d,J=5.6Hz,2H),3.73(t,J=4.8Hz,4H),3.18(t,J=4.8Hz,4H)

[0156] MS m / z(ESI)[M+H] + :414.23

[0157] Example 8

[0158] Preparation of (5-(4-fluorophenoxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0159]

[0160] According to the method of Example 3, 4-fluorophenol (369.6 mg) was used to replace phenol to obtain 239.3 mg of the product.

[0161] 1H NMR (400MHz, d-DMSO) δ: 14.53 (s, 1H), 12.98 (brs, 1H), 9.71 (s, 1H), 8.60 (s, 1H),7.53–7.49(m,2H),7.43–7.38(m,2H),5.86(s,1H),4.20(d,J=5.6Hz,2H)

[0162] MS m / z(ESI)[M+H] + :347.14

[0163] Example 9

[0164] Preparation of (5-(Propoxy)-7-hydroxy-[1,2,4]triazolyl[1,5-a]pyridine-8-carbonyl)glycine

[0165]

[0166] According to the method of Example 3, phenol was replaced by n-propanol (2 ml) to obtain 20.1 mg of the product.

[0167] 1H NMR (400MHz, d-DMSO) δ: 14.51 (s, 1H), 12.92 (brs, 1H), 9.65 (t, J = 5.6Hz, 1H), 8.47 (s, 1H), 6.4 4(s,1H),4.39(t,J=6.4Hz,2H),4.18(d,J=5.6Hz,2H),1.91–1.82(m,2H),1.03(t,J=7.2Hz,3H)

[0168] MS m / z(ESI)[M+H] + :295.12

[0169] Example 10

[0170] Preparation of [(7-hydroxy-5-(phenylethynyl)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid

[0171]

[0172] Step 1 Preparation of tert-butyl 7-hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylate

[0173] In a reaction flask, tert-butyl 7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylate (15 g) was dissolved in a mixed solution of methanol and tetrahydrofuran (25 mL / 50 mL), and stirred at room temperature under a hydrogen atmosphere until the reaction of the raw material was complete as monitored by TLC, and the reaction was stopped. The palladium carbon in the solution was removed by filtration, and the filtrate was then concentrated under reduced pressure to obtain 11.7 g of a crude product.

[0174] MS m / z(ESI):[M+H] + :236.24

[0175] Step 2 Preparation of tert-butyl 7-(pivaloyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylate

[0176] 7-Hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester (10.7g) was placed in a reaction bottle and tetrahydrofuran (65mL) was added, followed by triethylamine (8.0mL) and pivaloyl chloride (9.1g). Nitrogen was replaced three times and the reaction bottle was sealed. The reaction mixture was heated to 60°C and stirred overnight. The reaction was stopped until the reaction of the raw materials was complete as monitored by TLC. After the reaction solution was cooled to room temperature, the reaction solution was extracted with ethyl acetate, rinsed with saturated sodium bicarbonate aqueous solution, saturated ammonium chloride aqueous solution, and saturated brine in turn, and then concentrated under reduced pressure to obtain 12.0g of a yellow solid.

[0177] MS m / z(ESI):[M+H] + :320.36

[0178] Step 3 Preparation of tert-butyl 5-iodo-7-(pivaloyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylate

[0179] 7-(pivaloyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester (10.0g) was dissolved in tetrahydrofuran (50mL) solution, and then iodine (8.75g) was added, and the reaction temperature was kept below -60 degrees. After 5 minutes, hexamethyldisiloxane lithium (62.7mL, 1.0M) was added in batches and stirred until the raw material reaction was complete as monitored by TLC. Subsequently, hydrochloric acid ethyl acetate solution was added to quench the reaction, ethyl acetate was extracted, and sodium sulfite aqueous solution was washed. The organic phase was separated and washed with saturated ammonium chloride aqueous solution, saturated sodium bicarbonate aqueous solution, and saturated brine in turn. The organic phase was separated and concentrated under reduced pressure, and the crude product was column chromatographed to obtain 5.9g of the product.

[0180] 1H NMR (400MHz, CDCl3) δ: 8.36 (d, J = 5.2Hz, 1H), 7.45 (d, J = 5.2Hz, 1H), 2.31 (s, 6H)

[0181] MS m / z(ESI):[M+H] + :446.26

[0182] Step 4 Preparation of tert-butyl 5-(phenylethynyl)-7-hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylate

[0183] 5-iodo-7-(pivaloyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester (5.9 g), cuprous iodide (70 mg), Pd(dppf)Cl2 (285 mg), tetrahydrofuran (30 mL), triethylamine (6.6 g), and phenylacetylene (2.1 g) were added to the reaction bottle in sequence. Nitrogen was replaced three times, and the temperature was raised to 50°C for reaction until the reaction of the raw materials was complete as monitored by TLC, and ammonia water was added to quench the reaction. The organic phase was separated after extraction with ethyl acetate. The ammonia layer was washed with hydrochloric acid and extracted with ethyl acetate, and then the organic phases were combined, concentrated under reduced pressure, and purified by column chromatography to obtain 1.9 g of a red solid.

[0184] MS m / z(ESI):[M+H] + :336.36

[0185] Step 5 Preparation of 5-(phenylethynyl)-7-hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid

[0186] Dissolve 5-(phenylethynyl)-7-hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester (1.8 g) in a mixed solution of toluene and ethyl acetate (24 mL) in a volume ratio of 2:1, then add methanesulfonic acid (2.1 g) and stir, keeping the reaction temperature between 60-70 ° C. Stop the reaction until the raw material reaction is complete as monitored by TLC. After the reaction solution is cooled to room temperature, add ethyl acetate (10 mL), precipitate a large amount of solid, and filter to obtain a green solid. Subsequently, dissolve the solid with a small amount of N, N-dimethylacetamide, add ethyl acetate to the system, precipitate the solid, and filter to obtain 1.0 g of crude product.

[0187] MS m / z(ESI):[M+H] + :280.26

[0188] Step 6 Preparation of [(7-hydroxy-5-(phenylethynyl)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid methyl ester

[0189] In reaction flask A, 5-(phenylethynyl)-7-hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid (0.8 g) was dissolved in dichloromethane (20 mL), oxalyl chloride (0.76 g) was added dropwise, and stirred at room temperature. In reaction flask B, glycine methyl hydrochloride (0.68 g), dichloromethane (20 mL), and triethylamine (3.0 mL) were added in sequence and stirred at 0°C. TLC monitored that the raw material in reaction flask A was completely reacted, the stirring in reaction flask A was stopped, and the material in reaction flask A was added to reaction flask B, stirred at 0°C until the raw material reaction was completely monitored by TLC, and the reaction was quenched with water. Dichloromethane extraction, saturated ammonium chloride aqueous solution, saturated sodium bicarbonate aqueous solution were used to wash the organic phase in sequence, the organic phase was separated and combined, and after reduced pressure concentration, column chromatography was performed to obtain 0.8 g of yellow solid.

[0190] MS m / z(ESI):[M+H] + :351.33

[0191] Step 7 Preparation of [(7-hydroxy-5-(phenylethynyl)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid

[0192] Methyl [(7-hydroxy-5-phenylethynyl[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetate (0.8 g) was dissolved in ethanol (10 mL), and then an aqueous sodium hydroxide solution (18 mL, 2.0 M) was added and stirred at room temperature until the reaction of the raw material was complete as monitored by TLC. The pH was adjusted to 4-5 with dilute hydrochloric acid and filtered to obtain 0.42 g of a light yellow solid.

[0193] 1H NMR (400MHz, d-DMSO) δ: 14.33(s,1H),13.04(s,1H),9.90(s,1H),8.64(s,1 H),7.74–7.72(m,2H),7.61–7.53(m,3H),7.37(s,1H),4.24(d,J=5.6Hz,2H)

[0194] MS m / z(ESI):[M+H] + :337.31

[0195] Embodiment 11

[0196] Preparation of [(7-hydroxy-5-(phenylvinyl)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid

[0197]

[0198] Step 1 Preparation of tert-butyl 5-(phenylvinyl)-7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylate

[0199] Dissolve 5-iodo-7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester (500 mg), Pd(PPh3)2Cl2 (70.2 mg), sodium carbonate (360 mg), and styrene boronic acid (200 mg) in a mixed solvent (30 mL) of ethylene glycol dimethyl ether and degassed water in a volume ratio of 1:1, and replace nitrogen three times. Then stir at 80°C until the raw material reaction is complete as monitored by TLC, and stop heating. After the reaction solution is cooled to room temperature, extract with ethyl acetate, and wash the organic phase with saturated brine. Separate the organic phase, concentrate under reduced pressure, and separate by column chromatography to obtain 340 mg of crude product.

[0200] Step 2 Preparation of 5-(phenylvinyl)-7-hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid

[0201] Dissolve 5-(phenylvinyl)-7-(benzyloxy)-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid tert-butyl ester (290 mg) in dichloromethane (10 mL) solution and cool to -65°C, then add dichloromethane solution of boron tribromide (3.4 mL, 1.0 M) to the reaction system and stir until the reaction of the raw material is complete as monitored by TLC. Stop the reaction, concentrate the solution under reduced pressure to remove, then add the residue to a mixed solvent of water (10 mL) and methanol (3.0 mL) and stir at room temperature for 1 hour, and filter to obtain 250 mg of solid.

[0202] MS m / z(ESI):[M+H] + :282.04

[0203] Step 3 Preparation of [(7-hydroxy-5-(phenylvinyl)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid methyl ester

[0204] 5-(Vinyl)-7-hydroxy-[1,2,4]triazolo[1,5-a]pyridine-8-carboxylic acid (143 mg), N,N-dimethylformamide (10 mL), DIPEA (329 mg), PyBOP (395 mg), and glycine methyl hydrochloride (178 mg) were added to the reaction bottle in sequence, stirred at room temperature overnight, and the solid was filtered out to obtain 50 mg of crude product.

[0205] MS m / z(ESI):[M+H] + :353.28

[0206] Step 4 Preparation of [(7-hydroxy-5-(phenylvinyl)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetic acid

[0207] Methyl [(7-hydroxy-5-(phenylvinyl)-[1,2,4]triazolo[1,5-a]pyridine-8-carbonyl)amino]acetate (50 mg) was dissolved in methanol (2.0 mL), followed by addition of 30% aqueous sodium hydroxide solution (1.0 mL). The mixture was stirred at room temperature until the reaction of the raw material was complete as monitored by TLC. The pH was adjusted to between 1 and 2 with dilute hydrochloric acid and filtered to obtain 10 mg of the product.

[0208] 1H NMR (400MHz, d-DMSO) δ: 14.26 (s, 1H), 12.98 (s, 1H), 9.90 (t, J = 16.4Hz, 1H), 8.63 (s, 1 H),8.22(d,J=16.4Hz,1H),7.76–7.71(m,3H),7.50–7.40(m,4H),4.23(d,J=5.6Hz,2H)

[0209] MS m / z(ESI):[M+H] + :339.21

[0210] Biological tests

[0211] 1. PHD2 enzyme activity inhibition assay

[0212] 1.1 Reagents and consumables

[0213] PHD2 enzyme: purchased from Active motif;

[0214] α-Ketoglutarate sodium salt: purchased from Sigma;

[0215] FITC-HIF1α: purchased from GL;

[0216] Multi-well plate Nunc TM 384: Purchased from Thermo Scientific.

[0217] 1.2 Test methods

[0218] (1) Prepare 1× Assay buffer;

[0219] (2) Preparation of compound concentration gradient: The test compound concentration is 10 μM, 3-fold dilution, 10 concentrations, and duplicate testing; dilute to 100-fold final concentration solution in a 384-well plate, and then transfer 100 nL to a 384-well reaction plate using Echo550 for later use. Add 100 nL of 100% DMSO to the negative control well and the positive control well respectively;

[0220] (3) Prepare an enzyme solution with 2 times the final concentration using 1× Assay buffer;

[0221] (4) Add 5 μL of enzyme solution at 2 times the final concentration to the compound wells and positive control wells respectively;

[0222] (5) Add 5 μL of 1× Assay buffer to the negative control well;

[0223] (6) Centrifuge at 1000 rpm for 30 seconds, shake to mix, and incubate for 15 minutes;

[0224] (7) Prepare a tracer solution with a 2-fold final concentration using 1× Assay buffer, add 5.0 μL of the tracer solution with a 2-fold final concentration to start the reaction;

[0225] (8) The 384-well plate was centrifuged at 1000 rpm for 30 seconds and oscillated for 60 minutes. The mP value was read on Envision (Multimode Plate Reader, Perkin Elmer). The data was exported and processed to obtain the inhibition rate of the test compound. Compound numbers 1-9 correspond to the compounds prepared in Examples 1-9, respectively. The results are shown in the following table:

[0226] Table 1.

[0227] Compound No. <![CDATA[IC 50 (nM)]]> FG-4592 89.2 Enarodustat 21.9 Example 1 Compound 19.8 Example 2 Compound 34.8 Example 3 Compound 30.2 Example 4 Compound 33.3 Example 5 Compound 44.1 Example 6 Compound 47.6 Example 7 Compound 34.1 Example 8 Compound 35.4 Example 9 Compound 34.0

[0228] It can be seen from the above table that the compounds of the embodiments of the present invention have good HIF-prolyl hydroxylase inhibitory activity, and it can be seen that compounds 1-9 have good inhibitory activity.

[0229] 2. In vitro erythropoietin (EPO) induction activity test of the compounds of the present invention

[0230] The in vitro erythropoietin (EPO) induction activity of the compounds of Examples 10 and 11 of the present invention was evaluated using the cell line Hep3B (ATCC) derived from human liver cancer. Hep3B cells were cultured in EMEM (Eagle's Minimum Essential Medium) at 37°C in the presence of 10% fetal bovine serum (FBS). The experimental steps are as follows:

[0231] (1) Cell preparation: Cells were seeded in a 96-well plate at a volume of 2*10^4 cells per well and 100 μL per well;

[0232] (2) Preparation of compound concentration gradient: 100 μM, 20 μM, 2 replicate wells. Prepare a solution with a final concentration of 200 times in a 96-well plate, dilute the compound 200 / 3 times with cell culture medium, and then take 50 μL to administer to the cells. Add 50 μL of culture medium containing DMSO to the negative control well to make its final concentration contain 5‰ DMSO, and add 50 μL of the highest concentration of positive compound to the positive control well, and incubate at 37°C for 24 hours;

[0233] (3) Wash the reaction plate twice with approximately 400 μL of 1X Wash Buffer per well;

[0234] (4) Add 100 μL of diluted standard (including standard blank control) to appropriate wells;

[0235] (5) Add 50 μL of sample and 50 μL of Sample Diluent to the sample well;

[0236] (6) Add 50 μL of 1X Biotin Conjugated Antibody to all wells and incubate at room temperature for 1 hour;

[0237] (7) Wash the reaction plate six times with approximately 400 μL 1X Wash Buffer per well;

[0238] (8) Add 100 μL 1X Streptavidin-HRP to each well. Incubate at room temperature for 15 minutes.

[0239] (9) Wash the reaction plate six times with approximately 400 μL 1X Wash Buffer per well;

[0240] (10) Add 100 μL TMB Substrate Solution to each well. Incubate at room temperature for 10 minutes.

[0241] (11) Add 100 μL Stop Solution to each well;

[0242] (12) Read OD450 using EnSight.

[0243] The EPO induction activity of each compound is expressed as half maximal effect concentration (EC50). The experimental results are shown in the following table:

[0244] Table 2.

[0245] serial number <![CDATA[EC 50 (μM)]]> FG-4592 13.79 Enarodustat 15.06 Example 10 Compound 6.46 Example 11 Compound 1.79

[0246] As can be seen from the above table, the compounds of the present invention have higher EPO induction activity. It can be seen that the compounds obtained in Examples 10 and 11 have good induction activity.

[0247] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A compound represented by formula (I) or a pharmaceutically acceptable salt thereof, （I） in, R is selected from the following atoms or groups: or , L is -CH2- or -CH2O-, The value of n is 0; R 1 is a substituted or unsubstituted phenyl group, wherein the substituent is a C1-4 alkyl group, a C1-4 alkoxy group, or a halogen group; R 2 is a substituted or unsubstituted phenyl group, wherein the substituent is a C1-4 alkyl group, a C1-4 alkoxy group or a halogen group.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: The compound is selected from the following structures: 、 。 3. The compound according to claim 2 or a pharmaceutically acceptable salt thereof, characterized in that: The compound is selected from the following structures: 。 4. A method for preparing the compound according to any one of claims 1 to 3, characterized in that: There are two synthetic routes: Route 1: Route 2: Among them, R 1 , R 2 Have the same definitions as in the recited claims.

5. A pharmaceutical composition, characterized in that The invention comprises the compound represented by formula (I) according to claim 1 or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents and excipients.

6. Use of the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof and the pharmaceutical composition according to claim 5 in the preparation of a medicament for treating diseases mediated by inhibition of coenzyme aminoacyl hydroxylase by inhibiting coenzyme aminoacyl hydroxylase.

7. The use according to claim 6, characterized in that The diseases mediated by the inhibition of co-aminoacyl hydroxylase are anemia, local ischemia, and hypoxia.

8. The use according to claim 7, characterized in that The disease mediated by inhibition of co-aminoacyl hydroxylase is renal anemia.

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