WO2022078428A1 - 作为脯氨酰羟化酶抑制剂的化合物及其制备方法 - Google Patents

作为脯氨酰羟化酶抑制剂的化合物及其制备方法 Download PDF

Info

Publication number
WO2022078428A1
WO2022078428A1 PCT/CN2021/123750 CN2021123750W WO2022078428A1 WO 2022078428 A1 WO2022078428 A1 WO 2022078428A1 CN 2021123750 W CN2021123750 W CN 2021123750W WO 2022078428 A1 WO2022078428 A1 WO 2022078428A1
Authority
WO
WIPO (PCT)
Prior art keywords
compound
substituted
substituent
heterocycle
alkyl
Prior art date
Application number
PCT/CN2021/123750
Other languages
English (en)
French (fr)
Other versions
WO2022078428A8 (zh
Inventor
唐海涛
葛海涛
汤凡
王正俊
夏崇亮
Original Assignee
江苏苏中药业集团股份有限公司
江苏苏中药业研究院有限公司
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 江苏苏中药业集团股份有限公司, 江苏苏中药业研究院有限公司 filed Critical 江苏苏中药业集团股份有限公司
Priority to CN202180066355.9A priority Critical patent/CN116472266A/zh
Publication of WO2022078428A1 publication Critical patent/WO2022078428A1/zh
Publication of WO2022078428A8 publication Critical patent/WO2022078428A8/zh

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D211/00—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings
    • C07D211/04—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D211/80—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having two double bonds between ring members or between ring members and non-ring members
    • C07D211/84—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having two double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms, with at the most one bond to halogen directly attached to ring carbon atoms
    • C07D211/86—Oxygen atoms
    • C07D211/88—Oxygen atoms attached in positions 2 and 6, e.g. glutarimide
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
    • C07D405/04—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D409/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms
    • C07D409/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings
    • C07D409/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring-member bond

Definitions

  • the invention belongs to the technical field of medicine, and specifically discloses a compound as a prolyl hydroxylase inhibitor and a preparation method thereof.
  • hypoxia-inducible factor is a transcription factor initiated by somatic cells under hypoxia, and responds to cellular hypoxia by mediating a series of gene regulation in biological cells.
  • HIF is a heterodimer containing an oxygen-regulated ⁇ -subunit (HIF ⁇ ) and a constitutively expressed ⁇ -subunit (HIF ⁇ /ARNT).
  • HIF ⁇ subunit In oxygenated (normoxic) cells, the HIF ⁇ subunit is rapidly degraded by a mechanism of ubiquitination by the von Hippel-Lindaumorsuppressor (pVHL) E3 ligase complex. Under hypoxic conditions, HIF ⁇ is not degraded and the active HIF ⁇ / ⁇ complex accumulates in the nucleus and activates the expression of various genes including glycolytic enzymes, glucose transporters, erythropoietin (EPO) and blood vessels Endothelial Growth Factor (VEGF).
  • pVHL von Hippel-Lindaumorsuppressor
  • Erythropoietin is a naturally occurring hormone produced with HIF ⁇ that stimulates the production of red blood lipids (red blood cells) that carry oxygen throughout the body. EPO is normally secreted by the kidneys, and endogenous EPO increases under conditions of reduced oxygen (hypoxia). All types of anemia are characterized by a reduction in the blood's ability to carry oxygen and are thus accompanied by similar signs and symptoms, including pale skin and mucous membranes, weakness, dizziness, easy fatigue, and lethargy, resulting in a reduced quality of life. Anemia is usually associated with a condition in which blood is deficient in red blood cells or in hemoglobin.
  • anemia Common causes of anemia include iron, vitamin B12, and folic acid deficiencies, and can also be complicated by chronic diseases, such as inflammatory diseases, including those with secondary bone marrow inflammatory suppression.
  • Anemia is also associated with kidney dysfunction, and most patients with kidney failure on regular dialysis suffer from chronic anemia.
  • Prolyl hydroxylase domain is a key factor in the regulation of HIF.
  • PHD Prolyl hydroxylase domain
  • PHD can hydroxylate the two key proline residues Pro402 and Pro564 of HIF ⁇ , increasing its affinity with pVHL and accelerating the degradation process.
  • hypoxia and other pathological conditions the HIF reaction catalyzed by PHD is blocked and the degradation rate of protease is slowed down, resulting in the accumulation of HIF ⁇ in cells, which in turn causes a series of adaptive responses of cells to hypoxia.
  • Inhibiting PHD through PHD inhibitors prolongs the effect of HIF, thereby increasing the expression of genes such as EPO, and can effectively treat and prevent HIF-related and/or EPO-related disorders, such as anemia, ischemia and hypoxia. Studies have shown that deletion of the PHD2 gene can accelerate erythropoiesis.
  • Patent application 200780030720.0 discloses a prolyl hydroxylase inhibitor and its use method, the prolyl hydroxylase inhibitor is a compound with the following general formula:
  • X is N or CH;
  • R and R are each independently a unit selected from the following atoms or groups: i) hydrogen; ii) substituted or unsubstituted phenyl; and iii) substituted or unsubstituted heteroaryl group; the substituent is selected from: i) C 1 -C 4 linear, branched or cyclic alkyl; ii) C 1 -C 4 linear, branched or cyclic alkoxy; iii) C 1 -C 4 linear, branched or cyclic haloalkyl; iv) halogen; v) -CN; vi) -NHC(O)R 4 ; vii) -C(O)NR 5a R 5b ; viii) heteroaryl or ix) the two substituents may together form a fused ring having 5 to 7 atoms; R 4 is a C 1 -C 4 linear, branched or cyclic alkyl; R 5
  • Patent application 201710653887.8 discloses a pyridine derivative compound acting on prolyl hydroxylase, which has the following structure:
  • R1 is hydrogen or hydroxyl
  • R2 is independently selected from: hydrogen, halo, C 1-4 alkyl, phenyl, -O-phenyl, -O-benzyl, -O-C1-4 alkane group, phenyl optionally substituted with -C1-4 alkyl, benzyl optionally substituted with -C1-4.
  • the present invention provides a new compound as prolyl hydroxylase inhibitor and its preparation method and use, expands the types of medicines for the treatment of erythropoietin-related diseases, and can achieve good anti-proline Inhibitory activity of acyl hydroxylase.
  • the object of the present invention is to provide a compound as a prolyl hydroxylase inhibitor and its preparation method and application, expand the types of medicines for the treatment of erythropoietin-related diseases, and can achieve good p-prolyl hydroxylation Enzyme inhibitory activity.
  • the present invention provides a compound of formula I as a prolyl hydroxylase inhibitor, or a pharmaceutically acceptable salt, stereoisomer, tautomer, solvate or prodrug thereof :
  • R is selected from any one of substituted or unsubstituted aryl and heteroaryl;
  • the substituent is at least one of alkyl, alkoxy, haloalkyl, haloalkyl ether, aralkyl, halogen, cyano, phenyl, substituted or unsubstituted phenoxy.
  • aryl group is selected from phenyl
  • heteroaryl group is selected from:
  • the alkyl group is selected from C 1 -C 4 alkyl group, further selected from C 1 -C 2 alkyl group;
  • the alkoxy group is selected from C 1 -C 4 alkoxy group, further selected from C 1 -C 2 alkoxy;
  • the haloalkyl is selected from C 1 -C 4 haloalkyl, further selected from C 1 -C 2 haloalkyl;
  • the haloalkyl ether is selected from C 1 -C 4 haloalkyl ether, further selected From C 1 -C 2 haloalkyl ethers;
  • the aralkyl group is selected from C 1 -C 4 alkyl substituted 5-6-membered aryl groups, and is further selected from C 1 -C 2 alkyl substituted phenyl groups;
  • the Halogen is selected from any one of F, Cl, I and Br, and is further selected from F or Cl;
  • the compound is selected from the following four kinds:
  • R is a phenoxyphenyl substituted by at least one substituent, the substituent is independently selected from at least one of H, halogen, and C 1 -C 4 alkyl, preferably the phenoxy substituted in the meta position of benzene;
  • R is an aryl group substituted by at least one substituent independently selected from at least one of H, C 1 -C 4 haloalkyl, C 1 -C 4 haloalkyl ether, halogen, and aryl kind;
  • R is a benzo heterocycle substituted by at least one substituent, the substituent is independently at least one selected from H, C 1 -C 4 alkyl, and the heterocycle is nitrogen- or oxygen-containing 5-6 membered heterocycle;
  • R is a dibenzo heterocycle, and the heterocycle is a 5-6 membered heterocycle containing sulfur or oxygen.
  • the compound is selected from the following four kinds:
  • R is phenoxyphenyl substituted by at least one substituent independently selected from at least one of H, F, Cl, I, Br, C 1 -C 2 alkyl;
  • R is an aryl group substituted by at least one substituent independently selected from H, C 1 -C 2 haloalkyl, C 1 -C 2 haloalkyl ether, F, Cl, I, Br, at least one of phenyl;
  • R is a benzo heterocycle substituted by at least one substituent, the substituent is independently at least one selected from H, C 1 -C 2 alkyl, and the heterocycle is nitrogen- or oxygen-containing 5-6 membered heterocycle; (12-16)
  • R is a dibenzo heterocycle, and the heterocycle is a 5-6 membered heterocycle containing sulfur or oxygen.
  • the compound is selected from the following four kinds:
  • R is a phenoxyphenyl substituted by at least one substituent, and the substituent is independently selected from any one of H, Cl, and methyl;
  • R is a phenyl group substituted by at least one substituent independently selected from at least one of H, trifluoromethyl, trifluoromethyl ether, F, Cl, and phenyl;
  • R is a benzo heterocycle substituted by at least one substituent, the substituent is independently selected from at least one of H and methyl, and the heterocycle is a 5-6 membered heterocycle containing nitrogen or oxygen Ring, preferably from substituents A, B, C, D, E;
  • R is a dibenzo heterocycle, and the heterocycle is a 5-membered heterocycle containing sulfur or oxygen, preferably selected from G and H.
  • the compound is selected from any one of the following compounds:
  • R is selected from phenyl substituted with two substituents independently selected from Cl , OCF3 or CF3 .
  • R is selected from phenyl substituted by two substituents, the substituents are independently selected from Cl or CF 3 , and when the two substituents are all Cl, the positions of the two substituents are adjacent; when When the two substituents are all CF3 , the positions of the two substituents are alternate; when the two substituents are Cl and CF3 , respectively, the two substituents are adjacent in position.
  • the compound is selected from any one of the following compounds:
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising an effective dose of the above compound or a pharmaceutically acceptable salt, stereoisomer, tautomer, solvate or prodrug thereof, and a pharmaceutically acceptable carrier.
  • the present invention provides the above compounds or their pharmaceutically acceptable salts, stereoisomers, tautomers, solvates or prodrugs, or the use of the above pharmaceutical compositions in the preparation of medicines.
  • the medicament is for the treatment, prevention or pre-treatment of diseases mediated at least in part by hypoxia-inducible factor (HIF) and/or erythropoietin (EPO).
  • HIF hypoxia-inducible factor
  • EPO erythropoietin
  • the disease is selected from the group consisting of anemia disorders, renal anemia, neurological disorders and/or injuries, peripheral vascular disorders, ulcers, burns, chronic wounds and at least one of ischemia-reperfusion injury A sort of.
  • the disease can also be selected from ischemia, such as myocardial infarction, pulmonary embolism, intestinal infarction, chronic renal failure, ischemic stroke, cardiac sclerosis, transient ischemic attack, macular degeneration, peripheral arterial disease and congestive heart failure. at least one of them.
  • ischemia such as myocardial infarction, pulmonary embolism, intestinal infarction, chronic renal failure, ischemic stroke, cardiac sclerosis, transient ischemic attack, macular degeneration, peripheral arterial disease and congestive heart failure. at least one of them.
  • the present invention provides the use of the above-mentioned compound or pharmaceutical composition in the preparation of a medicament for inhibiting the activity of prolyl hydroxylase,
  • the mode of action of the use is that the prolyl hydroxylase is combined with a compound described herein or a pharmaceutically acceptable salt, stereoisomer, tautomer, solvate or prodrug thereof, or The pharmaceutical composition is contacted.
  • the present invention provides the preparation method of the above-mentioned compound, and the synthetic route is as follows:
  • R is selected from substituted or unsubstituted aryl and heteroaryl; the substituent is alkyl, alkoxy, haloalkyl, aralkyl, halogen, cyano, phenyl, oxygen-containing or at least one of nitrogen aromatic heterocycles;
  • Step 1 react compound 1 with benzyl alcohol and sodium hydride to obtain compound 2;
  • Step 2 reacting compound 2 with an alkali metal hydroxide to obtain compound 3, the alkali metal hydroxide is preferably sodium hydroxide;
  • Step 3 react compound 3 with glycine methyl ester hydrochloride in the presence of an amide coupling reagent to obtain compound 4;
  • Step 4 react compound 4 with palladium carbon under hydrogen conditions to obtain compound 5;
  • Step 5 react compound 5 with (CF 3 SO 2 ) 2 NC 6 H 5 to obtain compound 6;
  • Step 6 Compound 7 is obtained by reacting compound 6 with an arylboronic acid compound
  • Step 7 Compound 8 is obtained by reacting compound 7 with an inorganic base.
  • Step 1 The mixture of benzyl alcohol and sodium hydride was reacted in an ice-water bath for half an hour, then 1 was added, and the reaction was overnight at room temperature
  • Step 2 Compound 2 is refluxed overnight with sodium hydroxide in a mixed solution of water and 1,4-dioxane to obtain compound 3;
  • Step 3 Compartment of compound 3 with glycine methyl ester hydrochloride and PyBOP in N,N-dimethylformamide or tetrahydrofuran
  • Step 4 Compound 4 and palladium carbon are placed in alcohol solution or tetrahydrofuran solution under the atmosphere of hydrogen, and stirred at room temperature overnight
  • Step 5 Compound 5 is stirred with (CF 3 SO 2 ) 2 NC 6 H 5 in an alcoholic solution at room temperature overnight to obtain compound 6;
  • Step 6 Compound 6 and arylboronic acid compounds are dissolved in 1,4-dioxane in the presence of a palladium catalyst and an inorganic base,
  • Step 7 Compound 7 is stirred and reacted with an inorganic base in water and an organic solvent for 2 hours to obtain Compound 8;
  • the palladium catalyst can be selected from at least one of Pd(dppf)Cl 2 and Pd(PPh 3 ) 4
  • the inorganic base can be selected from at least one of cesium carbonate and sodium carbonate
  • the inorganic base can be selected from at least one of potassium hydroxide, sodium hydroxide, and lithium hydroxide
  • the organic solvent can be selected from at least one of methanol, ethanol, and tetrahydrofuran.
  • pharmaceutically acceptable salt in the present invention is intended to mean the free acid or base salt of the compound represented by formula (I), which is non-toxic, biologically tolerated, or in other words biologically Scientifically suitable for administration to a subject.
  • formula (I) the compound represented by formula (I)
  • G.S. Paulekuhn, et al. "Trendsin Active Pharmaceutical Ingredient Salt Selection based Analysis of the Orange Book Database", J. Med. Chem., 2007, 50: 6665-72, S.M.
  • Examples of pharmaceutically acceptable salts are those that are pharmacologically effective and suitable for contact with patient tissue without undue toxicity, irritation or allergic response.
  • Compounds of formula (I) may have sufficiently acidic groups, sufficiently basic groups, or both types of functional groups to react with a variety of inorganic or organic bases, as well as inorganic and organic acids, to form pharmaceutically acceptable Salt.
  • Examples of pharmaceutically acceptable salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, phosphate, monohydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, chloride , Bromide, Iodide, Acetate, Propionate, Caprate, Caprylate, Acrylate, Formate, Isobutyrate, Caproate, Heptanoate, Propiolate, Oxalic Acid Salt, malonate, phopoate, suberate, sebacate, fumarate, maleate, butyne-1,4-dioate, hexyne-1,6- Diacid salts, benzoates, chlorobenzoates, methyl benzoates, dinitrobenzoates, hydrocarbyl benzoates, oxybenzoates, phthalates , sulfonate, xylene sulfonate, phenylacetate, phenylpropionate,
  • Compounds of formula (I) contain a basic nitrogen, and the desired pharmaceutically acceptable salts can be prepared by any suitable method available in the art, such as treating the free base with a mineral acid such as hydrochloric acid, hydrobromic acid, Sulfuric acid, sulfamic acid, nitric acid, boric acid, phosphoric acid, etc., or treating the free base with an organic acid such as acetic acid, phenylacetic acid, propionic acid, stearic acid, lactic acid, ascorbic acid, maleic acid, hydroxy horse Leric acid, isethionic acid, succinic acid, valeric acid, fumaric acid, malonic acid, pyruvic acid, oxalic acid, glycolic acid, salicylic acid, oleic acid, palmitic acid, lauric acid, pyranosic acid (eg glucuronic acid or galacturonic acid), alpha-hydrocarbon acids (eg mandelic acid, citric acid or tartaric acid), amino acids
  • Compounds of formula (I) also contain carboxylic acids, and the desired pharmaceutically acceptable salts can be prepared by any suitable method, for example by treating the free acid with an inorganic or organic base such as an amine (primary). amines, secondary or tertiary amines), alkali metal hydroxides, alkaline earth metal hydroxides, any compatible mixture of bases such as those given as examples herein, and are considered equivalent according to the ordinary level of skill in the art or any other bases and mixtures thereof that can accept substitutions.
  • an inorganic or organic base such as an amine (primary).
  • suitable salts include organic salts derived from amino acids (eg, N-methyl-D-glucosamine, lysine, choline, glycine, and arginine), ammonia, carbonates , bicarbonate, primary, secondary, tertiary, and cyclic amines (such as tromethamine, benzylamine, pyrrolidine, piperidine, morpholine, and piperazine), and inorganic salts derived from sodium, Calcium, potassium, magnesium, manganese, iron, copper, zinc, aluminum and lithium.
  • amino acids eg, N-methyl-D-glucosamine, lysine, choline, glycine, and arginine
  • ammonia e.g, N-methyl-D-glucosamine, lysine, choline, glycine, and arginine
  • ammonia e.g, ammonia, carbonates , bicarbonate, primary, secondary, tertiary, and cycl
  • Prodrugs described herein may include compounds having an amino acid residue covalently linked to a carboxylic acid group of formula (I) through an amide or ester bond or two or more (eg two, three or four) polypeptide chains of amino acid residues.
  • amino acid residues include the twenty naturally occurring amino acids commonly identified by a three-letter symbol, as well as 4-hydroxyproline, hydroxylysine, demosine, isodemosine, 3-methyl Histidine, norvaline, beta-alanine, gamma-aminobutyric acid, citrulline homocysteine, homoserine, ornithine and methionine sulfone.
  • prodrugs can be prepared by derivatizing the free carboxylic acid of formula (I) as an amide or alkyl ester.
  • amides include those derived from ammonia, C1-6 alkyl primary amines, and di(C 1-6 alkyl) secondary amines.
  • Secondary amines include 5- or 6-membered heterocycloalkyl or heteroaryl ring moieties.
  • acid amines include those amides derived from ammonia. Ci - 3 alkyl primary amines and di(C 1-2 alkyl) amines.
  • Examples of the lipids of the present invention include C 1-7 alkyl esters, C 5-7 cycloalkyl esters, phenyl esters and (C 1-6 alkyl)phenyl esters.
  • the lipids include methyl esters.
  • Prodrugs are prepared by derivatizing the free hydroxyl groups with groups including hemisuccinate, phosphate, dimethylaminoacetate, and phosphoryloxymethoxycarbonyl.
  • Carbamate derivatives of hydroxyl and amino groups can also yield prodrugs.
  • Carbonate derivatives of hydroxyl groups, sulfonates and sulfates can also provide prodrugs.
  • the acyl groups are derivatized to (acyloxy) methyl esters and (acyloxy) ethyl esters, where the acyl group can be an alkyl ester optionally substituted with one or more ether, amine, or acid functional groups, or where the acyl group is As described above for amino acid lipids, this can also be used to generate prodrugs.
  • Prodrugs of this type can be described as Robinson et al., J Med Chem.
  • Free amines can also be derivatized as phthalamides, sulfonamides, or phosphonamides. All of these prodrug moieties can incorporate groups including ether, amine and carboxylic acid functionalities.
  • the invention provides a new compound as prolyl hydroxylase inhibitor, a preparation method and application thereof, expands the types of medicines for the treatment of erythropoietin-related diseases, and can achieve good p-prolyl hydroxylation Enzyme inhibitory activity.
  • the inhibitory effect of some compounds of the present invention on PHD2 enzyme has reached a level comparable to that of the marketed drug roxadustat (FG-4592), indicating that the compounds of the present invention have good biological activity on prolyl hydroxylase, It can be used to prepare medicines that inhibit the activity of prolyl hydroxylase.
  • the operation methods used are conventional operation methods
  • the equipment used are conventional equipment.
  • the starting materials may be appropriately selected such that the ultimately desired substituents are protected or unprotected as desired throughout the reaction scheme. Carry protected to obtain the desired product. Alternatively, it may be necessary or desirable to replace the ultimately desired substituent with a suitable group, which can be used throughout the reaction scheme, and then substituted with the desired substituent where appropriate.
  • the various variables are as defined above for formula (I).
  • the reaction can be carried out between the melting point of the solvent and the reflux temperature, preferably between 0°C and the reflux temperature of the solvent.
  • the reaction can also be carried out in a closed pressure vessel above the normal reflux temperature of the solvent.
  • these examples should not be construed as limiting the scope of the present invention.
  • 3,5-Bis-benzyloxy-pyridine-2-carbonitrile (2) (43.3 g) was dissolved in ethanol (360 mL), then 30% NaOH aqueous solution (690 mL) was added to the reaction flask, and then the temperature was raised to 90 °C React overnight. TLC detected, after the reaction of the raw materials was complete, the reaction was closed and cooled to room temperature, and the pH was adjusted to between 1-2 with 4N HCl, and solid appeared, filtered, and dried to obtain 40.2g of solid.
  • the reaction flask was operated without water and oxygen, replaced with N, and then DMF (700 mL) 3,5-bis-benzyloxy-pyridine- 2 -carboxylic acid (3) (80.4 g), EDCI (63.7 mL), HOBt ( 6.5 g), DIPEA (127 mL), and glycine methyl ester hydrochloride (45.2 g) were successively added to the reaction flask under an ice-water bath, and then stirred at room temperature for 3 days. After TLC detected that the reaction of the raw materials was complete, water was added to the reaction flask under stirring, followed by stirring for 1 hour, filtering, and drying to obtain 80 g of white solids.
  • Methyl ⁇ [5-(3-phenoxyphenyl)-3-hydroxypyridine-2-carbonyl]amino ⁇ acetate (7) (180 mg) was dissolved in MeOH (4 mL) followed by the addition of 30% aqueous NaOH ( 4 mL), heated to 40 °C and stirred for two hours. After TLC detects that the reaction of the raw materials is complete, the reaction is stopped, and after cooling to room temperature, the organic solvent is removed by concentrating under reduced pressure, and then the pH is adjusted to between 1 and 2 with 4N HCl, and a solid appears, which is filtered to obtain 104.3 mg of a white solid, which is the present invention. of compounds that act as prolyl hydroxylase inhibitors.
  • step 6 3-(4-chlorophenoxy)benzeneboronic acid (363.7mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 150.5mg of white solid, namely It is a compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 3-(3-chlorophenoxy)benzeneboronic acid (372.0mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 240.3mg of brown liquid, namely It is a compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 3-(2-chlorophenoxy)phenylboronic acid (372.0mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 241.3mg of white solid, namely It is a compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 3-(3,4-dichlorophenoxy)benzeneboronic acid (424.3mg) was used to replace 3-phenoxybenzeneboronic acid, and the rest were the same to obtain a white solid 112.5 mg, that is, the compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 3-(4-methylphenoxy)benzeneboronic acid (342.7mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 123.5mg of white solid, That is, the compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 4-trifluoromethoxybenzeneboronic acid (308.9mg) was used to replace 3-phenoxybenzeneboronic acid, and the rest were the same to obtain 150.5mg of white solid, which is the present invention of compounds that act as prolyl hydroxylase inhibitors.
  • step 6 replace 3-phenoxybenzeneboronic acid with 4-trifluoromethylbenzeneboronic acid (284.9mg), and the rest are the same to obtain 75.3mg of white solid, which is the present invention Compounds that are inhibitors of prolyl hydroxylase.
  • HPLC-MS m/z 341.07 [M+H]
  • step 6 replace 3-phenoxyphenylboronic acid with 3,4-dichlorophenylboronic acid (286.2mg), and the rest are the same to obtain 50.5mg of white solid, which is the present invention Compounds that are inhibitors of prolyl hydroxylase.
  • step 6 replace 3-phenoxyphenylboronic acid with 3,5-bis-trifluoromethylphenylboronic acid (386.9mg), and the rest are the same to obtain 120.5mg of white solid, which is Compounds of the present invention that are prolyl hydroxylase inhibitors.
  • step 6 replace 3-phenoxyphenylboronic acid with 4-phenylphenylboronic acid (296.7mg), and the rest are the same to obtain 111.4mg of white solid, which is the product of the present invention.
  • step 6 replace 3-phenoxyphenylboronic acid with 5-isoquinoline boronic acid (259.4mg), and the rest are the same to obtain 167.5mg of white solid, which is the Compounds that are inhibitors of aminoacyl hydroxylases.
  • step 6 5-(2,3-dihydrobenzofuran)boronic acid (246.3mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 143.5mg of white solid , which is the compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 replace 3-phenoxyphenylboronic acid with 3-quinoline boronic acid (259.5mg), and the rest are the same to obtain white solid 30.1mg, which is the proline of the present invention
  • Compounds that are acyl hydroxylase inhibitors are acyl hydroxylase inhibitors.
  • step 6 6-N-methylindoleboronic acid (262.5mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 130.2mg of white solid, which is the present invention of compounds that act as prolyl hydroxylase inhibitors.
  • HPLC-MS m/z 326.13 [M+H]
  • step 6 4-[b,d]-dibenzothiopheneboronic acid (342.2mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 207.9mg of white solid, That is, the compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 4-[b,d]-dibenzofuranboronic acid (318.7mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 134.7mg of white solid, That is, the compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 3-phenoxyphenylboronic acid was replaced with 3-trifluoromethyl-4-chlorophenylboronic acid (336.6 mg), and the rest were the same to obtain 54.7 mg of white solid, namely It is a compound of the present invention as a prolyl hydroxylase inhibitor.
  • step 6 replace 3-phenoxyphenylboronic acid with 3,5-dichlorophenylboronic acid (286.1mg), and the rest are the same to obtain 130.1mg of white solid, which is the invention Compounds that are inhibitors of prolyl hydroxylase.
  • step 6 2,5-dichlorophenylboronic acid (286.1mg) was used to replace 3-phenoxyphenylboronic acid, and the rest were the same to obtain 90.5mg of white solid, which is the present invention.
  • step 6 replace 3-phenoxyphenylboronic acid with 3-chloro-4-fluorophenylboronic acid (261.5mg), and the rest are the same to obtain 103.5mg of white solid, which is the present invention of compounds that act as prolyl hydroxylase inhibitors.
  • HPLC-MS m/z 339.08 [M+H]
  • Test Example 1 Detection of PHD2 Enzyme Activity Inhibition
  • PHD2 enzyme purchased from Activemotif; ⁇ -ketoglutarate sodium salt: purchased from Sigma; FITC-HIF1 ⁇ : purchased from GL; Multiwell plate Nunc TM 384: purchased from ThermoScientific.
  • 3Abcam ELISA (Abcam, ab119522) used 50 ⁇ L of cell supernatant sample, and the detailed procedure was carried out according to the kit instructions. Finally, through the absorbance value of the sample (OD 450nm -OD 620nm ) and the standard curve, the EPO expression-promoting content (mIU/mL) of the test substance was calculated. The results are shown in the following table:
  • the biological activity test results show that the compounds of the present invention have a more significant promoting effect on intracellular EPO expression than under hypoxic conditions.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Epidemiology (AREA)
  • Urology & Nephrology (AREA)
  • Vascular Medicine (AREA)
  • Cardiology (AREA)
  • Heart & Thoracic Surgery (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Pyrrole Compounds (AREA)

Abstract

提供一种作为脯氨酰羟化酶抑制剂的化合物及其制备方法，属于医药技术领域。所述化合物如式I所示，具有脯氨酰羟化酶抑制活性，可用于治疗由脯氨酰羟化酶活性介导的疾病，如肾性贫血等。

Description

作为脯氨酰羟化酶抑制剂的化合物及其制备方法 技术领域
本发明属于医药技术领域，具体公开了一种作为脯氨酰羟化酶抑制剂的化合物及其制备方法。
背景技术
在贫血、外伤、组织坏死及缺损等情况下，组织或细胞常处于低氧状态。低氧导致一系列转录诱导因子的表达，它们参与了血管形成、铁和糖代谢及细胞的生长和增殖。其中，缺氧诱导性因子(hypoxiainduciblefactor，HIF)是体细胞在缺氧状态下启动的一种转录因子，在生物细胞内通过介导一系列基因调节来响应细胞缺氧状态。HIF是含有氧调控的α-亚单位(HIFα)和组成型表达的β-亚单位(HIFβ/ARNT)的杂二聚体。在含氧(含氧量正常的)细胞中，HIFα亚单位通过逢希伯-林道肿瘤抑制因子(vonHippel-Lindautumorsuppressor，pVHL)E3连接酶复合物泛素化(ubiquitination)的机制被快速降解。在缺氧条件下，HIFα不被降解，且活性HIFα/β复合物在细胞核内积累，并活化各种基因的表达，包括糖酵解酶，葡萄糖转运蛋白，促红细胞生成素(EPO)和血管内皮生长因子(VEGF)。
促红细胞生成素(EPO)是随HIFα而产生的一种自然存在的激素，其剌激运载氧气贯穿全身的红细脂(红血球)的产生。EPO通常由肾分泌，且内源性EPO在氧减少(缺氧)的条件下增加。所有类型贫血的特征在于血液运载氧的能力减少，并因而伴有类似体征与症状，包括皮肤及粘膜苍白、虚弱、头晕、易疲劳和嗜睡，导致生活质量的下降。贫血通常与红细胞中或血红蛋白中血液缺乏的病况有关。贫血的普遍原因包括铁、维生素B12和叶酸缺乏，也会与慢性疾病并发，例如炎性疾病，包括具有继发骨髓炎性抑制的疾病等。贫血也与肾功能障碍有关，经常透析的大多数肾衰竭患者患有慢性贫血。
脯氨酰羟化酶(prolylhydroxylasedomain,PHD)是调节HIF的关键因子。PHD有三种亚型，即PHD1、PHD2和PHD3，所有亚型的活性位点都具有高度序列同源性。在常氧状态下，PHD可以羟基化HIFα的两个关键脯氨酸残基Pro402和Pro564，增加其与pVHL的亲和力，加速降解的过程。在缺氧及其他病理状态下，PHD催化的HIF反应受阻，蛋白酶降解速度减慢，造成HIFα在细胞内积聚，进而引起细胞对低氧的一系列适应性反应。通过PHD抑制剂抑制PHD，延长HIF的作用，进而增加EPO等基因的表达，可以有效治疗和预防HIF相关和/或 EPO相关的病症，如贫血、局部缺血和缺氧的病症。研究表明PHD2基因缺失能够加速红细胞生成。
专利申请200780030720.0公开了一种脯氨酰羟化酶抑制剂及使用方法，所述脯氨酰羟化酶抑制剂为具有如下通式的化合物：
Figure PCTCN2021123750-appb-000001
其中，X为N或CH；R和R 1各自独立地为选自以下原子或基团的单元：i)氢；ii)取代或未取代的苯基；以及iii)取代或未取代的杂芳基；所述取代基选自：i)C 1-C 4直链、支链或环状烷基；ii)C 1-C 4直链、支链或环状烷氧基；iii)C 1-C 4直链、支链或环状卤代烷基；iv)卤素；v)-CN；vi)-NHC(O)R 4；vii)-C(O)NR 5aR 5b；viii)杂芳基；或者ix)两个取代基可一起形成一个具有5至7个原子的稠环；R 4为C 1-C 4直链、支链或环状烷基；R 5a和R 5b各自独立地为：i)氢；ii)C 1-C 4直链、支链或环状烷基；或者iii)R 5a和R 5b可一起形成一个具有3至7个原子的环；R 2选自：i)-OR 6；或者ii)-NR 7aN 7b；R 6为氢或者C 1-C 4直链、支链或环状烷基；R 7a和R 7b各自独立地为：i)氢；ii)C 1-C 4直链、支链或环状烷基；或者iii)R 7a和R 7b可一起形成一个具有3至7个环原子的环；Y选自：i)氢；ii)-OR 3；R 3为氢、甲基或乙基；L为具有下式的连接单元：-[C(R 8aR 8b)] n-R 8a和R 8b各自独立地为氢、甲基或乙基；下标n选自1至3；R 9为氢或甲基。
专利申请201710653887.8公开了一种作用于脯氨酰羟化酶的吡啶衍生化合物，具有以下结构：
Figure PCTCN2021123750-appb-000002
其中：R1为氢或羟基；R2为独立地选自：氢、卤代基、C 1-4烷基、苯基、-O-苯基、-O-苄基、-O-C1-4烷基、任选地被-C1-4烷基取代的苯基、任选地被-C 1-4取代的苄基。
目前仍然需要以HIF为靶点的新的有效治疗促红细胞生成素相关疾病的化合物。有鉴于此，本发明提供一种新的作为脯氨酰羟化酶抑制剂的化合物及其制备方法、用途，拓展促红 细胞生成素相关疾病治疗的药物种类，并能实现很好的对脯氨酰羟化酶的抑制活性。
发明内容
本发明的目的是提供一种作为脯氨酰羟化酶抑制剂的化合物及其制备方法、用途，拓展促红细胞生成素相关疾病治疗的药物种类，并能实现很好的对脯氨酰羟化酶的抑制活性。
为实现上述发明目的，本发明的技术方案如下：
首先，本发明提供一种如式I所示的作为脯氨酰羟化酶抑制剂的化合物，或其药学上可接受的盐、立体异构体、互变异构体、溶剂化物或前药：
Figure PCTCN2021123750-appb-000003
其中，R选自被取代基取代或未被取代的芳基、杂芳基中的任一种；
所述取代基为烷基、烷氧基、卤代烷基、卤代烷基醚、芳烷基、卤素、氰基、苯基、取代或未被取代的苯氧基中的至少一种。
进一步地，所述芳基为选自苯基，所述杂芳基选自：
Figure PCTCN2021123750-appb-000004
进一步地，所述的烷基选自C 1-C 4烷基，进一步选自C 1-C 2烷基；所述烷氧基选自C 1-C 4烷氧基，进一步选自C 1-C 2烷氧基；所述卤代烷基选自C 1-C 4卤代烷基，进一步选自C 1-C 2卤代烷基；所述卤代烷基醚选自C 1-C 4卤代烷基醚，进一步选自C 1-C 2卤代烷基醚；所述芳烷 基选自C 1-C 4烷基取代的5-6元芳基，进一步选自C 1-C 2烷基取代的苯基；所述卤素选自F、Cl、I和Br中的任一种，进一步选自F或Cl；所述取代或未被取代的苯氧基选自被C 1-C 4烷基和/或卤素取代或未被取代的苯氧基，进一步选自被C 1-C 2烷基、F、Cl、I和Br中至少一种取代或未被取代的苯氧基。
进一步地，所述化合物选自以下四种：
(1)R为被至少一个取代基取代的苯氧基苯基，所述取代基独立的选自H、卤素、C 1-C 4烷基中的至少一种，优选所述苯氧基取代在苯的间位；
(2)R为被至少一个取代基取代的芳基，所述取代基独立的选自H、C 1-C 4卤代烷基、C 1-C 4卤代烷基醚、卤素、芳基中的至少一种；
(3)R为被至少一个取代基取代的苯并杂环，所述取代基独立的选自H、C 1-C 4烷基中的至少一种，所述杂环为含氮或氧的5-6元杂环；
(4)R为二苯并杂环，所述杂环为含硫或氧的5-6元杂环。
更进一步地，所述化合物选自以下四种：
(1)R为被至少一个取代基取代的苯氧基苯基，所述取代基独立的选自H、F、Cl、I、Br、C 1-C 2烷基中的至少一种；
(2)R为被至少一个取代基取代的芳基，所述取代基独立的选自H、C 1-C 2卤代烷基、C 1-C 2卤代烷基醚、F、Cl、I、Br、苯基中的至少一种；
(3)R为被至少一个取代基取代的苯并杂环，所述取代基独立的选自H、C 1-C 2烷基中的至少一种，所述杂环为含氮或氧的5-6元杂环；(12-16)
(4)R为二苯并杂环，所述杂环为含硫或氧的5-6元杂环。
再进一步地，所述化合物选自以下四种：
(1)R为被至少一个取代基取代的苯氧基苯基，所述取代基独立的选自H、Cl、甲基中的任一种；
(2)R为被至少一个取代基取代的苯基，所述取代基独立的选自H、三氟甲基、三氟甲基醚、F、Cl、苯基中的至少一种；
(3)R为被至少一个取代基取代的苯并杂环，所述取代基独立的选自H、甲基中的至少一种，所述杂环为含氮或氧的5-6元杂环,优选自取代基A、B、C、D、E；
(4)R为二苯并杂环，所述杂环为含硫或氧的5元杂环，优选自G、H。
再进一步地，所述的化合物选自以下化合物中的任一种：
Figure PCTCN2021123750-appb-000005
Figure PCTCN2021123750-appb-000006
Figure PCTCN2021123750-appb-000007
作为优选，R选自被两个取代基取代的苯基，所述取代基独立地选自Cl、OCF 3或CF 3。
进一步优选的，R选自被两个取代基取代的苯基，所述取代基独立地选自Cl或CF 3，且当两个取代基全为Cl时，两个取代基位置相邻；当两个取代基全为CF 3时，两个取代基位置相间；当两个取代基分别为Cl和CF 3时，两个取代基位置相邻。
最优选的，所述的化合物选自以下化合物中的任一种：
Figure PCTCN2021123750-appb-000008
另一方面，本发明提供一种药物组合物，包括有效剂量的上述化合物或其药学上可接受的盐、立体异构体、互变异构体、溶剂化物或前药，及药用载体。
再一方面，本发明提供上述化合物或其药学上可接受的盐、立体异构体、互变异构体、溶剂化物或前药，或上述药物组合物在制备药物方面的应用。
进一步地，所述药物用于治疗、预防或预先治疗至少部分由缺氧诱导因子(HIF)和/或促红细胞生成素(EPO)调介的疾病。
进一步地，所述疾病选自由以下病症组成的群组：贫血病症、肾性贫血、神经学病症和/或损伤、周围血管病症、溃疡、烧伤、慢性伤口和缺血-再灌注损伤中的至少一种。
进一步地，疾病还可以选自局部缺血，如心肌梗塞、肺栓塞、肠梗塞、慢性肾衰竭、缺 血性中风、心脏硬化、瞬时缺血发作、黄斑退化、外周动脉疾病和充血性心脏衰竭中的至少一种。
再一方面，本发明提供一种上述的化合物或药物组合物在制备用于抑制脯氨酰羟化酶的活性的药物中的用途，
进一步地，所述用途的作用方式为所述脯氨酰羟化酶与本文所述化合物或其药学上可接受的盐、立体异构体、互变异构体、溶剂化物或前药，或所述药物组合物接触。
最后，本发明提供上述化合物的制备方法，合成路线如下：
Figure PCTCN2021123750-appb-000009
其中，R选自被取代基取代或未被取代的芳基、杂芳基；所述取代基为烷基、烷氧基、卤代烷基、芳烷基、卤素、氰基、苯基、含氧或氮的芳杂环中的至少一种；
包括以下步骤：
步骤一：将化合物1与苄醇和氢化钠反应得到化合物2；
步骤二：将化合物2与碱金属氢氧化物反应得到化合物3，所述碱金属氢氧化物优选氢氧化钠；
步骤三：将化合物3与甘氨酸甲酯盐酸盐在酰胺偶联试剂存在的条件下反应得到化合物4；
步骤四：将化合物4与钯碳在氢气条件下反应得到化合物5；
步骤五：将化合物5与(CF 3SO 2) 2NC 6H 5反应得到化合物6；
步骤六：将化合物6与芳基硼酸类化合物反应得到化合物7；
步骤七：将化合物7与无机碱反应得到化合物8。
进一步地，包括以下步骤：
步骤一：将苄醇和氢化钠的混合物在冰水浴下反应半小时，然后加入1，室温下反应过夜
得到化合物2；
步骤二：化合物2与氢氧化钠在水和1,4-二氧六环的混合溶液中回流过夜得到化合物3；
步骤三：化合物3与甘氨酸甲酯盐酸盐和PyBOP在N，N-二甲基甲酰胺或四氢呋喃中室
温下搅拌过夜得到化合物4；
步骤四：化合物4与钯碳置于醇溶液或四氢呋喃溶液中在氢气的氛围下，室温搅拌过夜
得到化合物5；
步骤五：化合物5与(CF 3SO 2) 2NC 6H 5在醇溶液中室温搅拌过夜得到化合物6；
步骤六：化合物6与芳基硼酸类化合物在钯催化剂、无机碱存在下，溶于1,4-二氧六环，
在85℃的条件下，搅拌过夜反应，得到化合物7；
步骤七：化合物7与无机碱在水和有机溶剂中搅拌反应2小时，得到化合物8；
进一步地，步骤六中，所述钯催化剂可以选自Pd(dppf)Cl 2、Pd(PPh 3) 4中的至少一种，所述无机碱可以选自碳酸铯、碳酸钠中的至少一种，
进一步地，步骤七中，所述无机碱可以选自氢氧化钾、氢氧化钠、氢氧化锂中的至少一种，所述有机溶剂可以选自甲醇、乙醇、四氢呋喃中的至少一种。
本发明所称“药学上可接受的盐”，旨在表示式(I)表示的化合物的游离酸或碱的盐，其是无毒的、生物学耐受的，或换句话说是在生物学上适于施用给受试者。一般来讲，参见G.S.Paulekuhn,etal.，“Trendsin Active Pharmaceutical Ingredient Salt Selection based Analysis of the Orange Book Database”，J.Med.Chem.,2007,50：6665～72,S.M.Berge,etal.，“PharmaceuticalSalts”，JPharmSci.，1977,66：1-19,and Handbook of Pharmaceutical Salts,Properties,Selection,and Use,Stahland Wermuth,Eds.,Wiley-VC Hand VHCA,Zurich，2002(G.S.Paulekuhn等人，“根据橙皮书数据库的分析选择活性药物成分盐的趋势”，《药物化学杂志》，2007年，第50卷，第6665-6672页；S.M.Berge等人“，药用盐”,《药物科学杂志》，1977年，第66卷，第1-19页；以及《药用盐：特性、选择与应用》，Stahl和Wermuth编辑，Wiley-VCHandVHCA,Zurich，2002年)。可药用盐的例子是那些药理学有效且适于与患者组织接触而不会有不当毒性、刺激或变应性反应的盐。式(I)的化合物可以具有足够酸性的基团、足够碱性的基团或这两种类型的官能团，从而与多种无机碱或有机碱，以及无机酸和有机酸反应，形成可药用盐。
可药用盐的例子包括硫酸盐、焦硫酸盐、硫酸氢盐、亚硫酸盐、亚硫酸氢盐、磷酸盐、磷酸一氢盐、磷酸二氢盐、偏磷酸盐、焦磷酸盐、氯化物、溴化物、碘化物、乙酸盐、丙酸盐、癸酸盐、辛酸盐、丙烯酸盐、甲酸盐、异丁酸盐、己酸盐、庚酸盐、丙炔酸盐、草酸盐、 丙二酸盐、唬泊酸盐、辛二酸盐、癸二酸盐、富马酸盐、马来酸盐、丁炔一1,4一二酸盐、己炔一1,6一二酸盐、苯甲酸盐、氯代苯甲酸盐、甲基苯甲酸盐、二硝基苯甲酸盐、烃基苯甲酸盐、氧基苯甲酸盐、邻苯二甲酸盐、磺酸盐、二甲苯磺酸盐、苯乙酸盐、苯丙酸盐、苯丁酸盐、檬酸盐、乳酸盐、Y一轻丁酸盐、乙醇酸盐、酒石酸盐、甲磺酸盐、丙磺酸盐、蔡-1-磺酸盐、茶-2-磺酸盐和扁桃酸盐。
式(I)的化合物含有碱性氮，所需的可药用盐可通过本领域可用的任何合适方法制备，例如用无机酸处理该游离碱，所述无机酸例如是盐酸、氢溴酸、硫酸、氨基磺酸、硝酸、硼酸、磷酸等，或者用有机酸处理该游离碱，所述有机酸例如是乙酸、苯乙酸、丙酸、硬脂酸、乳酸、抗坏血酸、马来酸、羟基马来酸、羟乙磺酸、琥珀酸、戊酸、富马酸、丙二酸、丙酮酸、草酸、乙醇酸、水杨酸、油酸、棕桐酸、月桂酸、砒喃糖昔基酸(如葡糖醛酸或半乳糖醛酸)、α-烃酸(如扁桃酸、柠檬酸或酒石酸)、氨基酸(如天冬氨酸、戊二酸或谷氨酸)、芳族酸(如苯甲酸、2-乙酸氧基苯甲酸、蔡甲酸或肉桂酸)、磺酸(如月桂基磺酸、对甲苯磺酸、甲磺酸、乙磺酸)、诸如本文作为例子给出的那些酸的任何相容混合物、和根据本技术领域的普通技能水平被认为是等效物或可接受取代物的任何其他酸及其混合物。
式(I)的化合物也含有羧酸，所需的可药用盐可通过任何合适的方法制备，例如通过用无机或有机碱处理该游离酸来制备，所述无机或有机碱例如胺(伯胺、仲胺或叔胺)、碱金属氢氧化物、碱土金属氢氧化物、诸如本文作为例子给出的那些碱的任何相容混合物、和根据本技术领域的普通技能水平被认为是等效物或可接受取代物的任何其他碱及其混合物。合适的盐的示例性例子包括衍生自下述物质的有机盐：氨基酸(例如N-甲基-D-葡糖胺、赖氨酸、胆碱、甘氨酸和精氨酸)、氨、碳酸盐、碳酸氢盐、伯胺、仲胺、叔胺和环胺(例如氨基丁三醇、苄胺、吡咯烷、哌啶、吗啉和哌嗪)，以及衍生自下列物质的无机盐：钠、钙、钾、镁、猛、铁、铜、锌、铝和锂。
本文所述的前药可以包括这样的化合物，其具有通过酰胺键或酯键共价连接到式(I)羧酸基团的氨基酸残基或者两个或更多个(例如两个、三个或四个)氨基酸残基的多肽链。氨基酸残基的例子包括通常用三个字母符号标识的二十种天然存在的氨基酸以及4-羟脯氨酸、羟基赖氨酸、锁链素(demosine)、异锁链素(isodemosine)、3-甲基组氨酸、正缬氨酸、β-丙氨酸、γ-氨基丁酸、瓜氨酸高半胱氨酸、高丝氨酸、鸟氨酸和甲硫氨酸砜。
其他类型的前药可以通过将式(I)结构的游离羧酸衍生为酰胺或烷基酯来制备。酰胺的例子包括衍生自氨、C 1-6烷基伯胺和二(C 1-6烷基)仲胺的那些。仲胺包括5-元或6-元杂环烷基或杂芳基环部分。酸胺的例子包括衍生自氨.C 1-3烷基伯胺和二(C l-2烷基)胺的那些酰胺。本 发明的脂的例子包括C 1-7烷基酯,C 5-7环烷基酯、苯酯和(C 1-6烷基)苯酯。优选地，脂包括甲酯。还可以按照诸如Fleisheretal.,Adv.Drug Delivery Rev.1996,19,115-130(Fleisher等人，《药物递送进展综述》，1996年，第19卷，第115-130页)中所描述的那些方法，通过用包括半琥珀酸脂、磷酸脂、二甲基氨基乙酸脂、和磷酰氧基甲氧基羰基在内的基团使游离羟基衍生化来制备前药。羟基基和氨基的氨基甲酸酝衍生物也可产生前药。羟基的碳酸脂衍生物、磺酸脂和硫酸脂也可以提供前药。将经基衍生化为(酰氧基)甲脂和(酰氧基)乙脂，其中酰基可为任选被一个或多个醚、胺或浚酸官能团取代的烷基脂，或者其中酰基为如上所述的氨基酸脂，这也可用于产生前药。该类型的前药可如Robinsoneta1.，J Med Chem.1996,39(1)，10-18(Robinson等人，《药物化学杂志》，1996年，第39卷第1期，第10-18页)中所述制备。游离胺也可衍生化为酞胺、磺酞胺或磷酞胺。所有这些前药部分都可掺入包括醚、胺和羧酸官能团在内的基团。
本发明的有益效果为：
本发明提供了一种新的作为脯氨酰羟化酶抑制剂的化合物及其制备方法、用途，拓展促红细胞生成素相关疾病治疗的药物种类，并能实现很好的对脯氨酰羟化酶的抑制活性。本发明部分化合物对PHD2酶的抑制效果达到了与上市药物罗沙司他(FG-4592)相当的水平，说明本发明所述的化合物对脯氨酰羟化酶表现出较好的生物活性，可用于制备抑制脯氨酰羟化酶的活性的药物。
具体实施方式
为了使本发明实现的技术手段、创作特征、达成目的与功效易于明白了解，下面结合具体实施例，进一步阐明本发明，但下述实施例仅为本发明的优选实施例，并非全部。基于实施方式中的实施例，本领域技术人员在没有做出创造性劳动的前提下所获得其它实施例，都属于本发明的保护范围。
下述实施例中，若无特殊说明，所用的操作方法均为常规操作方法，所用设备均为常规设备。
下面的实施例可以对本发明做进一步地描述，技术人员会认识到，为获得本文的各种化合物，可适当地选择原料，使得最终需要的取代基会在整个反应方案中根据需要进行保护或不经保护的情况下携带，以得到需要的产物。或者，可能需要或者希望采用合适的基团代替该最终需要的取代基，该合适的基团可得以贯穿反应方案，然后在适当情况下用需要的取代基取代。除非另外指明，否则各种变量如上文针对式(I)所定义。反应可在溶剂的熔点和回流温度之间进行，优选在0℃和溶剂回流温度之间进行。反应还可在密闭压力容器中在溶剂 的正常回流温度以上进行。然而，这些实施例不应作为对本发明的范围的限制。
实施例1
{[5-(3-苯氧苯基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000010
步骤1)3,5-双-苄氧基-吡啶-2-腈(2)的制备：
将DMF(500mL)，60％氢化钠(26.4g)依次加入在反应瓶中，然后置于冰水浴中，缓慢滴加苄醇(68.3mL)，用时约20分钟。继续搅拌十分钟后，将3,5-二氯-2-氰基吡啶(51.6g)一次性加入反应瓶中，然后自然升至室温搅拌过夜。TLC检测原料反应完全后，搅拌状态下向反应瓶中加水，然后搅拌1小时，过滤，烘干得到86.5g黄色固体。
HPLC-MS：m/z 317[M+H]
步骤2)3,5-双-苄氧基-吡啶-2-羧酸(3)的制备：
将3,5-双-苄氧基-吡啶-2-腈(2)(43.3g)溶于乙醇(360mL)中，然后向反应瓶中加入30％NaOH水溶液(690mL),随后升温至90℃反应过夜。TLC检测，原料反应完全后，关闭反应冷却至室温，用4N HCl调节pH至1-2之间，出现固体，过滤，烘干得到40.2g固体。
HPLC-MS：m/z 336[M+H]
步骤3)[(3,5-双-苄氧基-吡啶-2-羰基)-氨基]乙酸甲酯(4)的制备：
反应瓶无水无氧操作，N 2置换，然后将DMF(700mL)3,5-双-苄氧基-吡啶-2-羧酸(3)(80.4g)，EDCI(63.7mL)，HOBt(6.5g)，DIPEA(127mL)，甘氨酸甲酯盐酸盐(45.2g)在冰水浴下依次加入反应瓶中，然后室温搅拌3天。TLC检测原料反应完全后，搅拌状态下向反应瓶中加水，然后搅拌1小时，过滤，烘干得到80g白色固体。
HPLC-MS：m/z 407[M+H]
步骤4)[(3,5-二羟基-吡啶-2-羰基)-氨基]乙酸甲酯(5)的制备：
反应瓶无水无氧操作，N 2置换一次，接着H 2置换三次，然后将[(3,5-双-苄氧基-吡啶-2-羰基)-氨基]乙酸甲酯(4)(80.4g)加入反应瓶中，并溶于MeOH(800mL)中，随后加入10％的Pd/C(8.0g)，室温下搅拌过夜。TLC检测原料反应完全后，停止反应，减压浓缩除去部分MeOH，接着往反应体系中加水，直至出现白色固体，过滤，烘干得到类白色固体33.2g。
HPLC-MS：m/z 227[M+H]
步骤5)[(3-羟基-5-三氟甲磺酰氧基-吡啶-2-羰基)-氨基]乙酸甲酯(6)的制备：
将[(3,5-二羟基-吡啶-2-羰基)-氨基]乙酸甲酯(5)(34.1g)，二异丙基乙胺(DIPEA)(32.1mL)溶于MeOH(300mL)中，冰水浴下加入N-苯基三氟甲磺酰亚胺(61.9g)，缓慢升至室温搅拌过夜。TLC检测原料反应完全后，停止反应，过滤得到灰色固体37.1g。
HPLC-MS：m/z 359[M+H]
步骤6){[5-(3-苯氧苯基)-3-羟基吡啶-2-羰基]氨基}乙酸甲酯(7)的制备：
反应瓶无水无氧操作，N 2置换一次，然后将[(3-羟基-5-三氟甲磺酰氧基-吡啶-2-羰基)-氨基]乙酸甲酯(6)(358mg)，3-苯氧基苯硼酸(323.1mg)，Pd(dppf)Cl 2(73.2mg)，K3PO4(636.9mg)，1,4-二氧六环(10mL)依次加入反应瓶中，封口85℃反应过夜。TLC检测原料反应完全后，停止反应，冷却到室温后乙酸乙酯萃取，水洗，分析有机相，浓缩，柱层析得白色固体约200mg。
步骤7){[5-(3-苯氧苯基)-3-羟基吡啶-2-羰基]氨基}乙酸(8)的制备：
将{[5-(3-苯氧苯基)-3-羟基吡啶-2-羰基]氨基}乙酸甲酯(7)(180mg)，溶于MeOH(4mL)中，随后加入30％NaOH水溶液(4mL)，升温至40℃搅拌两小时。TLC检测原料反应完全后，停止反应，冷却到室温后减压浓缩旋除有机溶剂，然后用4N HCl调节pH至1-2之间，出现固体，过滤，得到白色固体104.3mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 365.23[M+H]
实施例2
{[5-(3-(4-氯苯氧基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000011
按实施例1的方法，在步骤6)中，用3-(4-氯苯氧基)苯硼酸(363.7mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体150.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 399.28[M+H]
实施例3
{[5-(3-(3-氯苯氧基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000012
按实施例1的方法，在步骤6)中，用3-(3-氯苯氧基)苯硼酸(372.0mg)替换3-苯氧基苯 硼酸，其余皆相同，得棕色液体240.3mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 397.21[M-H]
实施例4
{[5-(3-(2-氯苯氧基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000013
按实施例1的方法，在步骤6)中，用3-(2-氯苯氧基)苯硼酸(372.0mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体241.3mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 397.23[M-H]
实施例5
{[5-(3-(3,4-二氯苯氧基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000014
按实施例1的方法，在步骤6)中，用3-(3,4-二氯苯氧基)苯硼酸(424.3mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体112.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 433.13[M+H]
实施例6
{[5-(3-(4-甲基苯氧基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000015
按实施例1的方法，在步骤6)中，用3-(4-甲基苯氧基)苯硼酸(342.7mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体123.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 379.24[M+H]
实施例7
{[5-(4-三氟甲氧基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000016
按实施例1的方法，在步骤6)中，用4-三氟甲氧基苯硼酸(308.9mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体150.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 357.18[M+H]
实施例8
{[5-(4-三氟甲基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000017
按实施例1的方法，在步骤6)中，用4-三氟甲基苯硼酸(284.9mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体75.3mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。HPLC-MS：m/z 341.07[M+H]
实施例9
{[5-(3,4-二氯基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000018
按实施例1的方法，在步骤6)中，用3,4-二氯苯硼酸(286.2mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体50.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 341.10[M+H]
实施例10
{[5-(3,5-双三氟甲基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000019
按实施例1的方法，在步骤6)中，用3,5-双三氟甲基苯硼酸(386.9mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体120.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合 物。
HPLC-MS：m/z 409.12[M+H]
实施例11
{[5-(4-苯基)苯基-3-羟基吡啶-2-羰基]氨基}乙酸制备
Figure PCTCN2021123750-appb-000020
按实施例1的方法，在步骤6)中，用4-苯基苯硼酸(296.7mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体111.4mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 349.21[M+H]
实施例12
{[5-(5-异喹啉)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000021
按实施例1的方法，在步骤6)中，用5-异喹啉硼酸(259.4mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体167.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 324.07[M+H]
实施例13
{[5-(2,3-二氢苯并呋喃基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000022
按实施例1的方法，在步骤6)中，用5-(2,3-二氢苯并呋喃)硼酸(246.3mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体143.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 315.11[M+H]
实施例14
{[5-(3-喹啉基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000023
按实施例1的方法，在步骤6)中，用3-喹啉硼酸(259.5mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体30.1mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 324.12[M+H]
实施例15
{[5-(2-苯并呋喃基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000024
按实施例1的方法，在步骤6)中，用2-苯并呋喃硼酸(243.7mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体195.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。HPLC-MS：m/z 313.10[M+H]
实施例16
{[5-(6-N-甲基吲哚)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000025
按实施例1的方法，在步骤6)中，用6-N-甲基吲哚硼酸(262.5mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体130.2mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。HPLC-MS：m/z 326.13[M+H]
实施例17
{[5-(4-[b,d]-二苯并噻吩)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000026
按实施例1的方法，在步骤6)中，用4-[b,d]-二苯并噻吩硼酸(342.2mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体207.9mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 379.20[M+H]
实施例18
{[5-(4-[b,d]-二苯并呋喃)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000027
按实施例1的方法，在步骤6)中，用4-[b,d]-二苯并呋喃硼酸(318.7mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体134.7mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 363.20[M+H]
实施例19
{[5-(3-三氟甲基-5-氯苯基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000028
按实施例1的方法，在步骤6)中，用3-三氟甲基-4-氯苯硼酸(336.6mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体54.7mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 375.16[M+H]
实施例20
{[5-(3,5-二氯苯基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000029
按实施例1的方法，在步骤6)中，用3,5-二氯苯硼酸(286.1mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体130.1mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 341.04[M+H]
实施例21
{[5-(2,5-二氯苯基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000030
按实施例1的方法，在步骤6)中，用2,5-二氯苯硼酸(286.1mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体90.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。
HPLC-MS：m/z 341.06[M+H]
实施例22
{[5-(3-氯-4-氟苯基)-3-羟基吡啶-2-羰基]氨基}乙酸的制备
Figure PCTCN2021123750-appb-000031
按实施例1的方法，在步骤6)中，用3-氯-4-氟苯硼酸(261.5mg)替换3-苯氧基苯硼酸，其余皆相同，得白色固体103.5mg，即为本发明的作为脯氨酰羟化酶抑制剂的化合物。HPLC-MS：m/z 339.08[M+H]
生物学测试
试验例1：PHD2酶活抑制检测
试剂及耗材：
PHD2酶：从Activemotif处购买；α-酮戊二酸钠盐：从Sigma处购买；FITC-HIF1α：从GL处购买；多孔板Nunc TM384：从ThermoScientific处购买。
试验方法
(1)配制1×Assaybuffer检测溶液(10mM HEPES，150mM NaCl，0.05％Tween20，pH7.4)；(2)化合物浓度梯度的配制：受试化合物测试浓度为10μM起始，3倍稀释，10个浓度，复孔测试；在384孔板中稀释成100倍终浓度的溶液，然后用Echo550转移100nL到384反应板中备用。阴性对照孔和阳性对照孔中分别加100nL的100％DMSO；(3)用1×Assaybuffer配制2倍终浓度的酶溶液；(4)在化合物孔和阳性对照孔分别加5μL的2倍终浓度的酶溶液；(5)在阴性对照孔中加5μL的1×Assaybuffer；(6)1000rpm离心30秒，振荡混匀后孵育15min；(7)用1×Assaybuffer配制2倍终浓度的tracer溶液。加入5.0μL的2倍终浓度的tracer溶液，起始反应；(8)将384孔板1000rpm离心30秒，振荡混匀60min，从Envision(MultimodePlateReader，PerkinElmer)上读数得到mP值，导出数据并进行处理，得到待测化合物的抑制率，结果如下表所示：
表1.
化合物编号 IC 50(nM)
FG-4592 65.6
1 61.6
2 50.5
3 86.9
4 59.6
5 138.8
6 76.7
7 104.2
8 90.8
9 25.3
10 26.4
11 96.6
12 118.0
13 87.9
14 54.2
15 99.7
16 106.2
17 142.4
18 87.9
19 53.0
22 23.0
由上表可知，本发明实施例化合物具有很好的HIF-脯氨酰羟化酶抑制活性。
试验例2：细胞实验
①消化收集Hep-3B细胞，用含10％FBS(Gibco，10270-106)的完全培养液(含NEAA的MEM基础培养基，上海中乔新舟生物科技有限公司，ZQ-300)，调整细胞密度为4×10 4个/mL，按100μL每孔接种于PDL(终浓度10μg/mL)预处理1h的2块96孔板，放置常氧培养箱(5％CO 2，21％O 2，37℃)6h使贴壁。其中1块板用于考察化合物两个浓度在常氧条件 下培养24h对EPO产生的影响，同时铺设常氧溶剂对照孔(DMSO，终浓度0.1％)，另1块板用于缺氧条件下(5％CO 2，1％O 2，37℃)培养24h的缺氧溶剂对照孔(DMSO，终浓度0.1％)。
②细胞在常氧下培养6小时后，吸弃原培养液，用PBS润洗3遍，分别加入DMSO(终浓度0.1％)、待测化合物(终浓度10μM)，每孔200μL，在无血清培养基中常氧培养24h。缺氧下诱导的细胞不经过药物处理，加入DMSO(终浓度0.1％)在无血清培养基中培养24h。吸取上清液(约200μL)到0.2mL EP管，冻存在-80℃冰箱备用等待ELISA检测，下层细胞加入MTT液(终浓度0.5mg/mL，常氧培养箱中继续孵育约4h)，待细胞中出现蓝紫色结晶，停止孵育，弃上清，加入150μL DMSO/孔，200rpm震荡溶解10min，于570nm检测吸光度，650nm作为参比波长，以溶剂对照孔细胞活力为100％评价各实验组细胞活力变化。
③Abcam ELISA(Abcam，ab119522)使用50μL细胞上清液样品，详细流程按照试剂盒说明书进行。最后通过样品的吸光度值(OD 450nm-OD 620nm)和标准曲线计算出受试物促EPO表达含量(mIU/mL)，结果如下表所示：
表2.
化合物编号 IC 50(nM)
FG-4592 65.6
1 61.6
2 50.5
3 86.9
4 59.6
5 138.8
6 76.7
7 104.2
8 90.8
9 25.3
10 26.4
11 96.6
12 118.0
13 87.9
14 54.2
15 99.7
16 106.2
17 142.4
18 87.9
19 23.0
生物活性测试结果显示本发明化合物相比缺氧条件下，对细胞内EPO表达具有更为显著的促进作用。
以上所述仅为本发明的较佳实施例而已，并不用以限制本发明，凡在本发明的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本发明的保护范围之内。

Claims (10)

  1. 一种如式I所示的作为脯氨酰羟化酶抑制剂的化合物，或其药学上可接受的盐、立体异构体、互变异构体、溶剂化物或前药：
    Figure PCTCN2021123750-appb-100001
    其中，R选自被取代基取代或未被取代的芳基、杂芳基中的任一种；
    所述取代基为烷基、烷氧基、卤代烷基、卤代烷基醚、芳烷基、卤素、氰基、苯基、取代或未被取代的苯氧基中的至少一种。
  2. 根据权利要求1所述的化合物，其特征在于，所述芳基为选自苯基，所述杂芳基选自：
    Figure PCTCN2021123750-appb-100002
  3. 根据权利要求1所述的化合物，其特征在于，所述的烷基选自C 1-C 4烷基，进一步选自C 1-C 2烷基；所述烷氧基选自C 1-C 4烷氧基，进一步选自C 1-C 2烷氧基；所述卤代烷基选自C 1-C 4卤代烷基，进一步选自C 1-C 2卤代烷基；所述卤代烷基醚选自C 1-C 4卤代烷基醚，进一步选自C 1-C 2卤代烷基醚；所述芳烷基选自C 1-C 4烷基取代的5-6元芳基，进一步选自C 1-C 2烷基取代的苯基；所述卤素选自F、Cl、I和Br中的任一种，进一步选自F或Cl；所述取代或未被取代的苯氧基选自被C 1-C 4烷基和/或卤素取代或未被取代的苯氧基，进一步选自被C 1-C 2烷基、F、Cl、I和Br中至少一种取代或未被取代的苯氧基。
  4. 根据权利要求1所述的化合物，其特征在于，所述化合物选自以下四种：
    (1)R为被至少一个取代基取代的苯氧基苯基，所述取代基独立的选自H、卤素、C 1-C 4烷基中的至少一种，优选所述苯氧基取代在苯的间位；
    (2)R为被至少一个取代基取代的芳基，所述取代基独立的选自H、C 1-C 4卤代烷基、C 1-C 4卤代烷基醚、卤素、芳基中的至少一种；
    (3)R为被至少一个取代基取代的苯并杂环，所述取代基独立的选自H、C 1-C 4烷基中的至少一种，所述杂环为含氮或氧的5-6元杂环；
    (4)R为二苯并杂环，所述杂环为含硫或氧的5-6元杂环，
    进一步地，所述化合物选自以下四种：
    (1)R为被至少一个取代基取代的苯氧基苯基，所述取代基独立的选自H、F、Cl、I、Br、C 1-C 2烷基中的至少一种；
    (2)R为被至少一个取代基取代的芳基，所述取代基独立的选自H、C 1-C 2卤代烷基、C 1-C 2卤代烷基醚、F、Cl、I、Br、苯基中的至少一种；
    (3)R为被至少一个取代基取代的苯并杂环，所述取代基独立的选自H、C 1-C 2烷基中的至少一种，所述杂环为含氮或氧的5-6元杂环；
    (4)R为二苯并杂环，所述杂环为含硫或氧的5-6元杂环，
    更进一步地，所述化合物选自以下四种：
    (1)R为被至少一个取代基取代的苯氧基苯基，所述取代基独立的选自H、Cl、甲基中的任一种；
    (2)R为被至少一个取代基取代的苯基，所述取代基独立的选自H、三氟甲基、三氟甲基醚、F、Cl、苯基中的至少一种；
    (3)R为被至少一个取代基取代的苯并杂环，所述取代基独立的选自H、甲基中的至少一种，所述杂环为含氮或氧的5-6元杂环,优选自取代基A、B、C、D、E；
    (4)R为二苯并杂环，所述杂环为含硫或氧的5元杂环，优选自G、H。
  5. 根据权利要求4所述的化合物，其特征在于，所述的化合物选自以下化合物中的任一种：
    Figure PCTCN2021123750-appb-100003
    Figure PCTCN2021123750-appb-100004
    Figure PCTCN2021123750-appb-100005
    Figure PCTCN2021123750-appb-100006
  6. 根据权利要求1所述的化合物，其特征在于，R选自被两个取代基取代的苯基，所述取代基独立地选自Cl、OCF 3或CF 3。
  7. 根据权利要求6所述的化合物，其特征在于，R选自被两个取代基取代的苯基，所述取代基独立地选自Cl或CF 3，且当两个取代基全为Cl时，两个取代基位置相邻；当两个取代基全为CF 3时，两个取代基位置相间；当两个取代基分别为Cl和CF 3时，两个取代基位置相邻。
  8. 根据权利要求7所述的化合物，其特征在于，所述的化合物选自以下化合物中的任一种：
    Figure PCTCN2021123750-appb-100007
  9. 权利要求1-8任一项所述化合物的制备方法，其特征在于，合成路线如下：
    Figure PCTCN2021123750-appb-100008
    包括以下步骤：
    步骤一：将化合物1与苄醇和氢化钠反应得到化合物2；
    步骤二：将化合物2与碱金属氢氧化物反应得到化合物3；
    步骤三：将化合物3与甘氨酸甲酯盐酸盐在酰胺偶联试剂存在的条件下反应得到化合物4；
    步骤四：将化合物4与钯碳在氢气条件下反应得到化合物5；
    步骤五：将化合物5与(CF 3SO 2) 2NC 6H 5反应得到化合物6；
    步骤六：将化合物6与芳基硼酸类化合物反应得到化合物7；
    步骤七：将化合物7与无机碱反应得到化合物8。
  10. 根据权利要求9所述的制备方法，其特征在于，具体包括以下步骤：
    步骤一：将苄醇和氢化钠的混合物在冰水浴下反应半小时，然后加入1，室温下反应过夜得到化合物2；
    步骤二：化合物2与氢氧化钠在水和1,4-二氧六环的混合溶液中回流过夜得到化合物3；
    步骤三：化合物3与甘氨酸甲酯盐酸盐和PyBOP在N，N-二甲基甲酰胺或四氢呋喃中室温下搅拌过夜得到化合物4；
    步骤四：化合物4与钯碳置于醇溶液或四氢呋喃溶液中在氢气的氛围下，室温搅拌过夜得到化合物5；
    步骤五：化合物5与(CF 3SO 2) 2NC 6H 5在醇溶液中室温搅拌过夜得到化合物6；
    步骤六：化合物6与芳基硼酸类化合物在钯催化剂、无机碱存在下，溶于1,4-二氧六环，在85℃的条件下，搅拌过夜反应，得到化合物7；
    步骤七：化合物7与无机碱在水和有机溶剂中搅拌反应2小时，得到化合物8。
PCT/CN2021/123750 2020-10-16 2021-10-14 作为脯氨酰羟化酶抑制剂的化合物及其制备方法 WO2022078428A1 (zh)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202180066355.9A CN116472266A (zh) 2020-10-16 2021-10-14 作为脯氨酰羟化酶抑制剂的化合物及其制备方法

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN202011114555 2020-10-16
CN202011114555.0 2020-10-16

Publications (2)

Publication Number Publication Date
WO2022078428A1 true WO2022078428A1 (zh) 2022-04-21
WO2022078428A8 WO2022078428A8 (zh) 2023-03-30

Family

ID=81207690

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2021/123750 WO2022078428A1 (zh) 2020-10-16 2021-10-14 作为脯氨酰羟化酶抑制剂的化合物及其制备方法

Country Status (2)

Country Link
CN (1) CN116472266A (zh)
WO (1) WO2022078428A1 (zh)

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101506149A (zh) * 2006-06-26 2009-08-12 宝洁公司 脯氨酰羟化酶抑制剂及使用方法
CN107417605A (zh) * 2017-08-02 2017-12-01 江苏艾立康药业股份有限公司 作用于脯氨酰羟化酶的吡啶衍生化合物
WO2019028150A1 (en) * 2017-08-01 2019-02-07 Akebia Therapeutics, Inc. COMPOSITIONS FOR USE IN METHODS OF TREATING HEMOGLOBINOPATHIES
CN109593084A (zh) * 2019-01-23 2019-04-09 中国药科大学 脯氨酰羟化酶小分子光敏前药及其制备方法与应用
CN111349077A (zh) * 2019-02-02 2020-06-30 杭州华东医药集团新药研究院有限公司 一种哒嗪衍生物及其制备方法和医药用途

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101506149A (zh) * 2006-06-26 2009-08-12 宝洁公司 脯氨酰羟化酶抑制剂及使用方法
WO2019028150A1 (en) * 2017-08-01 2019-02-07 Akebia Therapeutics, Inc. COMPOSITIONS FOR USE IN METHODS OF TREATING HEMOGLOBINOPATHIES
CN107417605A (zh) * 2017-08-02 2017-12-01 江苏艾立康药业股份有限公司 作用于脯氨酰羟化酶的吡啶衍生化合物
CN109593084A (zh) * 2019-01-23 2019-04-09 中国药科大学 脯氨酰羟化酶小分子光敏前药及其制备方法与应用
CN111349077A (zh) * 2019-02-02 2020-06-30 杭州华东医药集团新药研究院有限公司 一种哒嗪衍生物及其制备方法和医药用途

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
DEMETRIADES, MARINA ET AL.: "Dynamic Combinatorial Chemistry Employing Boronic Acids/Boronate Esters Leads to Potent Oxygenase Inhibitors", ANGEW.CHEM.INT.ED., vol. 51, 25 May 2012 (2012-05-25), pages 6672 - 6675, XP055272020, DOI: 10.1002/anie.201202000 *
LEI, YONGHUA ET AL.: "An improved and scalable synthesis of N-(5-(4-cyanophenyl)-3- hydroxypicolinoyl)glycine, a promising PHD2 inhibitor for the treatment of anemia", TETRAHEDRON LETTERS, vol. 56, 9 July 2015 (2015-07-09), pages 5017 - 5019, XP029248344, DOI: 10.1016/j.tetlet.2015.07.013 *

Also Published As

Publication number Publication date
CN116472266A (zh) 2023-07-21
WO2022078428A8 (zh) 2023-03-30

Similar Documents

Publication Publication Date Title
EP0385237B1 (en) 2-(1-piperazinyl)-4-phenylcycloalkanopyridine derivatives, processes for the production thereof, and pharmaceutical composition containing the same
JP4615826B2 (ja) 置換インドールおよびインテグリンアンタゴニストとしてのそれらの使用
AU2014232308B2 (en) Multisubstituted aromatic compounds as serine protease inhibitors
JP5284268B2 (ja) Ｃｆｔｒモジュレーターとしてのアザインドール誘導体
EP3083554B1 (en) Dimethylbenzoic acid compounds
KR101274879B1 (ko) 1-시클로프로필메틸-4-[2-(3,3,5,5-테트라메틸시클로헥실)페닐]피페라진의 염 및 결정
WO2023046158A1 (zh) 氮杂喹啉酮类化合物及其医药用途
JP2010520889A (ja) 蛋白キナーゼの阻害剤として有用なアミノピリミジン
IL223978A (en) Bipyridyl derivatives useful for the treatment of kinase-induced diseases
EP3080125A1 (en) Novel azaindole derivatives as selective histone deacetylase (hdac) inhibitors and pharmaceutical compositions comprising the same
KR20090122931A (ko) 오르니틴 유도체
JP2001512103A (ja) 第ｘａ因子を阻害する複素環誘導体
NO20023293L (no) Piperidin- og piperazinderivater som virker som 5-HT2A- reseptorantagonister
JP4805166B2 (ja) アロイルフランおよびアロイルチオフェン
JP2018508563A (ja) Ｕｓｐ７阻害剤化合物及び使用方法
Li et al. Design, synthesis and biological evaluation of diamino substituted cyclobut-3-ene-1, 2-dione derivatives for the treatment of drug-resistant tuberculosis
JP2007308402A (ja) シンナモイル化合物及びその用途
WO2008053863A1 (fr) Nouveau composé ayant un squelette de 1,4-benzothiazin-3-one ou un squelette de 3,4-dihydroquinolin-2-one
US8350026B2 (en) VEGF-2 receptor and protein tyrosine kinase inhibitors and pharmaceutical use thereof
WO2022078428A1 (zh) 作为脯氨酰羟化酶抑制剂的化合物及其制备方法
WO2016104451A1 (ja) 新規複素環誘導体
SK11462001A3 (sk) Heterocyklické deriváty ako inhibítory faktora xa, farmaceutická kompozícia, ktorá ich obsahuje a ich použitie
TW202220961A (zh) Ｐｈｄ抑制劑化合物、組成物及使用方法
KR20210080347A (ko) 트롬빈 억제제, 제제, 및 그의 용도
SK8652003A3 (en) Piperazinylcarbonylquinoline and piperazinylcarbonylisoquinoline derivative, process for the preparation thereof, use and pharmaceutical composition comprising same

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 21879471

Country of ref document: EP

Kind code of ref document: A1

WWE Wipo information: entry into national phase

Ref document number: 202180066355.9

Country of ref document: CN

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 21879471

Country of ref document: EP

Kind code of ref document: A1