A salt of an endothelin A (ETA) receptor antagonist compound, its preparation method and medical use

Abstract

The present invention belongs to the technical field of chemical medicines and provides a salt of an endothelin A (ETA) receptor antagonist compound and a preparation method and application thereof.

Description

Technical Field

[0001] The present invention belongs to the technical field of chemical medicines and provides a salt of an endothelin A (ETA) receptor antagonist compound and a preparation method and application thereof. Background Art

[0002] Atrasentan (CAS: 173937-91-2) is a potent and selective endothelin A (ETA) receptor antagonist with the following structural formula:

[0003] Atrasentan hydrochloride (CAS: 195733-43-8) is used clinically and has the following structure:

[0004] It was previously evaluated in clinical trials for the treatment of prostate cancer and is currently being evaluated in clinical trials for the treatment of chronic kidney disease associated with type 2 diabetes. It has also been shown to reduce albuminuria in patients with diabetic nephropathy. Currently, there is no marketed product.

[0005] Nearly half of drug molecules exist in the form of salts. Salt formation can also improve certain undesirable physicochemical or biological properties of drugs. Therefore, developing salts of endothelin A (ETA) receptor antagonist compounds with superior physicochemical or pharmaceutical properties is of great significance. Summary of the Invention

[0006] In view of the problems existing in the prior art, the present invention provides a salt of an endothelin A (ETA) receptor antagonist compound with a novel structure, a preparation method and an application thereof.

[0007] Specifically, the present invention provides a salt of a compound represented by formula (I), whose structural formula is as follows Figure 1 As shown below:

[0008]

[0009] M is an inorganic acid or an organic acid, wherein n:x=1:1-2.

[0010] As a preferred technical solution of the present invention, the inorganic acid is selected from sulfuric acid, hydrochloric acid, hydrofluoric acid, hydrobromic acid, hydroiodic acid, phosphoric acid, carbonic acid or nitric acid;

[0011] The organic acid is selected from the group consisting of benzoic acid, 2,5-dihydroxybenzoic acid, 4-acetamidobenzoic acid, 4-aminobenzoic acid, oxalic acid, acetic acid, dichloroacetic acid, trichloroacetic acid, caproic acid, adipic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, caprylic acid, capric acid, cinnamic acid, citric acid, aspartic acid, gluconic acid, glutamic acid, lactic acid, malic acid, mandelic acid, pyroglutamic acid, tartaric acid, succinic acid, formic acid, fumaric acid, gentisic acid, glutaric acid, valeric acid, aspartic acid, lauric acid, camphoric acid, maleic acid, malonic acid, nicotinic acid, propionic acid, salicylic acid, 4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid, palmitic acid, pamoic acid, trifluoroacetic acid, thiocyanic acid, p-toluenesulfonic acid, and L-malic acid.

[0012] As a preferred technical solution of the present invention, the inorganic acid is selected from phosphoric acid.

[0013] As a preferred technical solution of the present invention, the inorganic acid is selected from phosphoric acid, wherein n=3, x=3-6.

[0014] As a preferred technical solution of the present invention, the inorganic acid is selected from phosphoric acid, wherein n=3, x=3; or n=3, x=4, or n=3, x=5, or n=3, x=6.

[0015] As a preferred technical solution of the present invention, the organic acid is selected from oxalic acid.

[0016] As a preferred technical solution of the present invention, the organic acid is selected from oxalic acid, wherein n=1, x=1.

[0017] The present invention further provides a pharmaceutical composition comprising a salt of the compound represented by formula (I) and one or more pharmaceutically acceptable carriers.

[0018] The present invention further provides use of a salt of the compound represented by formula (I) in the preparation of a medicament for treating and / or preventing diseases related to endothelin A (ETA) receptor antagonism.

[0019] As a preferred technical solution of the present invention, the diseases include chronic kidney disease, IgA, FSGS, Alport and hypertension.

[0020] The present invention further provides a method for preparing a salt of the compound represented by formula (I), wherein the compound A

[0021] Prepared by mixing with acid.

[0022] As a preferred technical solution of the present invention, the molar ratio of the compound to the acid molecule is 1:1-2.

[0023] As a preferred technical solution of the present invention, the following steps are included:

[0024] 1) Weigh an appropriate amount of free base and dissolve it in a benign solvent;

[0025] 2) Weigh an appropriate amount of ionic acid and add it to the reaction in step 1 to form a salt;

[0026] 3) back-titrifying the reaction solution of step 2 into a poor solvent and stirring to precipitate;

[0027] 4) Rapidly centrifuge or allow to stand to obtain the salt of the compound.

[0028] The acid is selected from M, which is an inorganic acid or an organic acid. The inorganic acid is selected from sulfuric acid, hydrochloric acid, hydrofluoric acid, hydrobromic acid, hydroiodic acid, phosphoric acid, carbonic acid or nitric acid. The organic acid is selected from the group consisting of benzoic acid, 2,5-dihydroxybenzoic acid, 4-acetamidobenzoic acid, 4-aminobenzoic acid, oxalic acid, acetic acid, dichloroacetic acid, trichloroacetic acid, caproic acid, adipic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, caprylic acid, capric acid, cinnamic acid, citric acid, aspartic acid, gluconic acid, glutamic acid, lactic acid, malic acid, mandelic acid, pyroglutamic acid, tartaric acid, succinic acid, formic acid, fumaric acid, gentisic acid, glutaric acid, valeric acid, aspartic acid, lauric acid, camphoric acid, maleic acid, malonic acid, nicotinic acid, propionic acid, salicylic acid, 4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid, palmitic acid, pamoic acid, trifluoroacetic acid, thiocyanic acid, p-toluenesulfonic acid, and L-malic acid.

[0029] The solvent used for salt formation in the present invention is selected from at least one of ethyl acetate, methanol, n-propanol, isopropanol, isopropyl ether, tetrahydrofuran, isopropyl acetate, acetone, methyl tert-butyl ether, acetonitrile, ethanol, 1,4-dioxane, n-hexane, and isopropyl ether.

[0030] Furthermore, in an optional embodiment, the method for preparing the aforementioned pharmaceutically acceptable salt further comprises the steps of volatilizing the solvent or stirring for crystallization, filtering, drying, and the like.

[0031] Unless otherwise indicated, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase should not be considered as undefined or unclear in the absence of a specific definition, but should be understood according to its ordinary meaning. When a trade name appears in this article, it is intended to refer to its corresponding commercial product or its active ingredient. The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions and / or dosage forms that are suitable for use in contact with human and animal tissues within the scope of sound medical judgment without excessive toxicity, irritation, allergic reaction or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0032] As used herein, "pharmaceutically acceptable salts" pertain to derivatives of the compounds of the present invention wherein the parent compound is modified by forming a salt with an acid or a base.

[0033] Prodrugs of the compounds described herein readily undergo chemical changes under physiological conditions to convert to the compounds of the invention. Additionally, prodrugs can be converted to the compounds of the invention by chemical or biochemical methods in an in vivo environment.

[0034] Certain compounds of the present invention may exist in unsolvated forms as well as solvated forms, including hydrates. In general, the solvated forms are equivalent to the unsolvated forms and are encompassed within the scope of the present invention.

[0035] The atoms of the molecules of the compounds of the present invention are isotopes, and isotope derivatization can generally extend half-life, reduce clearance, enhance metabolic stability and improve in vivo activity. In addition, an embodiment is included in which at least one atom is replaced by an atom having the same atomic number (proton number) and a different mass number (protons and neutrons). Examples of isotopes included in the compounds of the present invention include hydrogen atoms, carbon atoms, nitrogen atoms, oxygen atoms, phosphorus atoms, sulfur atoms, fluorine atoms, chlorine atoms, which respectively include 2 H. 3 H. 13 C. 14 C. 15 N. 17 O. 18 O. 31 P. 32 P. 35 S. 18 F. 36 Cl. In particular, radioactive isotopes that emit radiation as they decay, such as 3H or 14C, can be used in pharmaceutical preparations or for local anatomical testing of compounds in vivo. Stable isotopes do not decay or change with their amount, nor are they radioactive, and therefore can be used safely. When the atoms constituting the molecules of the compounds of the invention are isotopes, the isotopes can be converted according to general methods by replacing the reagents used in the synthesis with reagents containing the corresponding isotopes.

[0036] The compounds of the present invention may contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute the compound. For example, the compounds may be labeled with radioactive isotopes, such as deuterium ( 2 H), iodine-125( 125 I) or C-14( 14 C). All isotopic variations of the compounds of the present invention, whether radioactive or not, are included within the scope of the present invention. Furthermore, the compounds of the present invention may have one or more hydrogen atoms replaced by the isotope deuterium ( 2H) substitution, the compounds of the present invention, after deuteration, have the effects of extending half-life, reducing clearance rate, enhancing metabolic stability and improving in vivo activity. The preparation method of the isotopic derivatives generally includes: phase transfer catalysis method. For example, the preferred deuteration method uses a phase transfer catalyst (e.g., tetraalkylammonium salt, NBu4HSO4). The use of a phase transfer catalyst to exchange the methylene protons of the diphenylmethane compound results in the introduction of higher deuterium than reduction with deuterated silane (e.g., triethyldeuterated monosilane) in the presence of an acid (e.g., methanesulfonic acid) or with a Lewis acid such as aluminum trichloride using sodium deuterated borate.

[0037] The term "pharmaceutically acceptable carrier" refers to any formulation carrier or medium that can deliver an effective amount of the active substance of the present invention, does not interfere with the biological activity of the active substance, and has no toxic side effects on the host or patient. Representative carriers include water, oils, vegetables and minerals, cream bases, lotion bases, ointment bases, etc. These bases include suspending agents, viscosity increasing agents, transdermal enhancers, etc. Their preparations are well known to those skilled in the art of cosmetics or topical medicine. For additional information about carriers, reference can be made to Remington: The Science and Practice of Pharmacy, 21st Ed., Lippincott, Williams & Wilkins (2005), the contents of which are incorporated herein by reference.

[0038] The term "excipient" generally refers to a carrier, diluent and / or vehicle required to formulate an effective pharmaceutical composition.

[0039] With respect to a drug or pharmacologically active agent, the term "effective amount" or "therapeutically effective amount" refers to a non-toxic amount of the drug or agent sufficient to achieve the intended effect. For the oral dosage forms of the present invention, an "effective amount" of an active substance in the composition means the amount required to achieve the intended effect when used in combination with another active substance in the composition. The determination of an effective amount varies from person to person, depending on the age and general condition of the recipient, as well as the specific active substance. The appropriate effective amount in each individual case can be determined by those skilled in the art through routine experimentation.

[0040] The terms "active ingredient," "therapeutic agent," "active substance," or "active agent" refer to a chemical entity that is effective in treating a target disorder, disease, or condition.

[0041] The term "tautomer" or "tautomeric form" refers to structural isomers with different energies that can be converted into each other through a low energy barrier. If tautomerism is possible (such as in solution), a chemical equilibrium of the tautomers can be reached. For example, proton tautomers (also known as prototropic tautomers) include interconversions performed by proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers include interconversions performed by the reorganization of some bonding electrons. Keto-enol tautomerism. Another example of tautomerism is phenol-keto tautomerism. Unless otherwise indicated, all tautomeric forms of the compounds of the present invention are within the scope of the present invention.

[0042] The compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis- and trans-isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, and racemic mixtures and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which are within the scope of the present invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All such isomers and mixtures thereof are encompassed within the scope of the present invention.

[0043] Optically active (R)- and (S)-isomers, as well as D and L isomers, can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If one enantiomer of a compound of the present invention is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, wherein the resulting diastereomeric mixture is separated and the auxiliary group is cleaved to provide the pure desired enantiomer. Alternatively, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), a diastereomeric salt is formed with an appropriate optically active acid or base, and then the diastereoisomers are resolved by conventional methods known in the art, and then the pure enantiomer is recovered. In addition, the separation of enantiomers and diastereomers is typically accomplished using chromatography, which employs a chiral stationary phase and is optionally combined with a chemical derivatization method (e.g., carbamate formation from an amine).

[0044] "Optional" or "optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not.

[0045] The compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthesis methods, and equivalent substitutions well known to those skilled in the art. Preferred embodiments include but are not limited to the examples of the present invention.

[0046] The beneficial effects of the present invention compared to the prior art include:

[0047] The compound of the present invention is an endothelin A (ETA) receptor antagonist compound. The prodrug can be completely converted into atrasentan in human liver microsomes and has a technical effect equivalent to that of atrasentan in pharmacokinetic research.

[0048] The salt of the compound represented by formula (I) provided by the present invention has improved physicochemical properties compared with the free compound represented by formula (I), and is more suitable for pharmaceutical applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0049] Figure 1 Schematic diagram of the structural formula of the salt of compound A of the present invention

[0050] Figure 2 This is the NMR spectrum of the hydrochloride obtained in Example 18 of the present invention

[0051] Figure 3 This is the NMR spectrum of the sulfate obtained in Example 19 of the present invention. DETAILED DESCRIPTION

[0052] The present invention will be described in further detail below with reference to the embodiments and drawings, but the embodiments of the present invention are not limited thereto.

[0053] The structures of the compounds were determined by nuclear magnetic resonance (NMR) or mass spectrometry (MS). -6 The unit of ppm is given. NMR measurements were performed using a Bruker AVANCE-III NMR spectrometer, using deuterated dimethyl sulfoxide (DMSO-d6) and deuterated chloroform (CDCl3) as solvents and tetramethylsilane (TMS) as the internal standard.

[0054] MS was measured using an ISQ EC mass spectrometer (manufacturer: Thermo, model: ISQ EC).

[0055] High performance liquid chromatography (HPLC) analysis was performed using a Thermo U3000 HPLC DAD instrument.

[0056] The CombiFlash rapid preparation instrument used CombiFlash Rf+LUMEN (TELEDYNE ISCO).

[0057] Thin layer chromatography silica gel plate using Yantai Yinlong HSGF 254 or GF 254 Silica gel plates, the specifications of silica gel plates used in thin layer chromatography (TLC) are 0.17mm~0.23mm, and the specifications used for thin layer chromatography separation and purification products are 0.4mm~0.5mm.

[0058] Silica gel column chromatography generally uses Rushan Shangbang silica gel 100-200 mesh silica gel as the carrier.

[0059] The present invention relates to the following reagents: DMF (N,N-dimethylformamide), KI (potassium iodide), Cs2CO3 (cesium carbonate), DCM (dichloromethane), n-hexane, EA (ethyl acetate), Py (pyridine), CF3COOH (trifluoroacetic acid), IPA (isopropyl alcohol), isopropyl ether, and acetone.

[0060] Example 1

[0061] 1-[(Ethoxycarbonyl)oxy]methyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0062]

[0063] Step A: Synthesis of 1-[(ethoxycarbonyl)oxy]methyl(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0064]

[0065] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (500 mg, 0.98 mmol), chloromethyl ethyl carbonate (270 mg, 1.96 mmol), cesium carbonate (640 mg, 1.96 mmol) and potassium iodide (325 mg, 1.96 mmol) were added to 10 mL of dry DMF at room temperature and heated to 65 °C for 2 h.

[0066] After the reaction, the reaction solution was cooled to room temperature and poured into 40 ml of ice-water solution, extracted with dichloromethane (100 ml × 3), and the organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, concentrated and evaporated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 1) to give 480 mg of colorless oily product [(ethoxycarbonyl)oxy]methyl (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 79.9%).

[0067] LC-MS: RT = 2.20 min, [M+H] + =613.42.

[0068] 1 H NMR (400MHz, DMSO) δ7.23(d,J=8.6Hz,2H),7.04(d,J=1.2Hz,1H),6.89(d,J=8.7Hz,2H),6.84–6.76(m,2H),5.98( d,J=4.6Hz,2H),5.58(q,J=6.2Hz,2H),4.12(q,J=7.1Hz,2H),3.78–3.72(m,1H),3.72(s,3H),3.49(dd,J=11.2,5. 3Hz,1H),3.26–3.20(m,3H),3.16(s,1H),3.02–2.90(m,3H),2.86–2.80(m,1H),2.71(d,J=13.8Hz,1H),1.33(dd, J=14.6,7.0Hz,2H),1.27–1.09(m,7H),0.96(dd,J=14.7,7.3Hz,2H),0.81(t,J=7.3Hz,3H),0.71(t,J=7.3Hz,3H).

[0069] Example 2

[0070] 1-[(Ethoxycarbonyl)oxy]ethyl (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0071]

[0072] Step A: Synthesis of 1-[(ethoxycarbonyl)oxy]ethyl (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0073]

[0074] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (500 mg, 0.98 mmol), 1-chloroethyl ethyl carbonate (298 mg, 1.96 mmol), cesium carbonate (640 mg, 1.96 mmol) and potassium iodide (325 mg, 1.96 mmol) were added to 10 mL of dry DMF at room temperature and heated to 65 °C for 2 h.

[0075] After the reaction, the reaction solution was cooled to room temperature and poured into 40 ml of ice-water solution, extracted with dichloromethane (100 ml × 3), and the organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, concentrated and evaporated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 1) to give 442 mg of colorless oily product 1-[(ethoxycarbonyl)oxy]methyl (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 66.5%).

[0076] LC-MS: RT = 2.21 min, [M+H] + =627.40. 1H NMR (400MHz, DMSO) δ7.27–7.17(m,2H),7.09–6.98(m,1H),6.92–6.85(m,2H),6.85–6.74(m,2H),6.60–6. 52(m,1H),5.98(d,J=5.3Hz,2H),4.11(dq,J=11.1,7.1Hz,2H),3.67-3.73(m,4H),3.57–3.44(m,1H),3.30 –3.20(m,3H),3.19–3.10(m,1H),3.01–2.85(m,3H),2.77(ddd,J=9.4,6.9,2.6Hz,1H),2.70(d,J=13.9Hz, 1H),1.39–1.22(m,6H),1.21–1.04(m,6H),1.01–0.89(m,2H),0.81(t,J=7.3Hz,3H),0.71(t,J=7.3Hz,3H)

[0077] Example 3

[0078] 1-[(Isopropyloxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0079]

[0080] Step A: Synthesis of 1-[(isopropoxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0081]

[0082] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (500 mg, 0.98 mmol), 1-chloroethyl isopropyl carbonate (325 mg, 1.96 mmol), cesium carbonate (640 mg, 1.96 mmol) and potassium iodide (325 mg, 1.96 mmol) were added to 10 mL of dry DMF at room temperature and heated to 65 °C for 2 h.

[0083] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), and the organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 1) to give 482 mg of colorless oily product 1-[(isopropoxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 76.9%).

[0084] LC-MS: RT = 2.29 min, [M+H] + =641.48. 1 H NMR(400MHz, DMSO)δ7.25(t,J=9.0Hz,2H),7.07–7.04(m,1H),6.93–6.89(m,2H),6.85–6.78(m,2H),6.59–6.54 (m,1H),5.99(d,J=6.2Hz,2H),4.79–4.69(m,1H),3.76–3.70(m,4H),3.55–3.44(m,1H),3.33–3.23(m,4H),3.21 –3.13(m,1H),3.02–2.90(m,3H),2.80–2.75(m,1H)2.71(d,J=13.9Hz,1H),1.39–1.33(m,1H),1.32(d,J=5.4Hz ,2H),1.26(d,J=5.4Hz,2H),1.22–1.14(m,9H),1.00–0.94(m,2H),0.82(t,J=7.3Hz,3H),0.72(t,J=7.3Hz,3H).

[0085] Example 4

[0086] 1-[(Methoxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0087]

[0088] Step A: Synthesis of 1-[(methoxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0089]

[0090] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (500 mg, 0.98 mmol), 1-chloroethyl methyl carbonate (270 mg, 1.96 mmol), cesium carbonate (640 mg, 1.96 mmol) and potassium iodide (325 mg, 1.96 mmol) were added to 10 mL of dry DMF at room temperature and heated to 65 °C for 2 h.

[0091] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), and the organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 1) to give 428 mg of colorless oily product 1-[(methoxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 71.3%).

[0092] LC-MS: RT = 2.20 min, [M+H] + =613.42. 1 H NMR (400MHz, DMSO) δ7.25 (dd, J=10.3, 8.7Hz, 2H), 7.08–7.04 (m, 1H), 6.91 (dd, J=8.7, 2.9 Hz,2H),6.86–6.78(m,2H),6.60–6.55(m,1H),5.99(d,J=5.1Hz,2H),3.76–3.69(m,6H),3. 56–3.44(m,1H),3.33–3.16(m,5H),3.00–2.94(m,3H),2.81–2.76(m,1H),2.71(d,J=13.7H z,1H),1.39–1.14(m,9H),1.02–0.93(m,2H),0.82(t,J=7.3Hz,3H),0.72(t,J=7.3Hz,3H).

[0093] Example 5

[0094] 1-(((2-(Methylamino)ethoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0095]

[0096] Step A: Synthesis of tert-butyl (2-(((1-chloroethoxy)carbonyl)oxy)ethyl)(methyl)carbamate

[0097]

[0098] Under ice bath, 2-(N-Boc-N-methylamino)ethanol (500 mg, 2.85 mmol) and pyridine (248 mg, 3.14 mmol) were added to 10 ml of dry dichloromethane, and 1-chloroethyl chloroformate (248 mg, 3.14 mmol) was added dropwise under ice bath. After the addition was completed, the temperature was raised to room temperature and the reaction was carried out for 1 hour.

[0099] After the reaction, the mixture was poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), and the organic layer was washed twice with 1 M / L hydrochloric acid (20 ml), washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure to give 725 mg of a colorless oily product (tert-butyl 2-(((1-chloroethoxy)carbonyl)oxy)ethyl)(methyl)carbamate (yield: 90.6%).

[0100] Step B: Synthesis of 2,2,5-trimethyl-4,9-dioxo-3,8,10-trioxa-5-aza-11-ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0101]

[0102] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (500 mg, 0.98 mmol), tert-butyl (2-(((1-chloroethoxy)carbonyl)oxy)ethyl)(methyl)carbamate (550 mg, 1.96 mmol), cesium carbonate (640 mg, 1.96 mmol) and potassium iodide (325 mg, 1.96 mmol) were added to 10 ml of dry DMF at room temperature and the temperature was raised to 65 °C for 4 h.

[0103] After the reaction, the mixture was cooled to room temperature and poured into 40 mL of ice-water solution. Extraction was performed with dichloromethane (20 mL x 3). The combined organic phases were washed with saturated brine (50 mL), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 1) to obtain 625 mg of a colorless oily product, 2,2,5-trimethyl-4,9-dioxo-3,8,10-trioxa-5-aza-11-ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 84.4%). LC-MS: RT = 2.34 min, [M+H] + =756.49.

[0104] Step C: Synthesis of 1-(((2-(methylamino)ethoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0105]

[0106] At room temperature, 2,2,5-trimethyl-4,9-dioxo-3,8,10-trioxa-5-aza-11-ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (300 mg, 0.39 mmol) was dissolved in 10 ml of ethyl acetate solvent. 2 ml of 4 M / L dioxane hydrochloride solution was added under ice-cooling and the reaction was carried out at room temperature for 2 hours.

[0107] After the reaction, the reaction mixture was concentrated to obtain 260 mg of yellow solid 1-(((2-(methylamino)ethoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 101.6%).

[0108] LC-MS: RT = 1.82 min, [M+H] + =656.45. 1H NMR (400MHz, DMSO) δ7.60 (m, 2H), 7.21 (m, 1H), 7.01 (d, J = 8.1Hz, 2H), 6.90 (d,J=15.8Hz,2H),6.50(m,1H),6.04(s,2H),3.77(s,3H),3.17(d,J=6.4Hz ,3H),3.08–2.90(m,4H),2.52(d,J=5.6Hz,3H),2.06(d,J=4.1Hz,3H),1.9 0(s,6H),1.33(d,J=34.4Hz,4H),1.23–1.10(m,7H),0.83(t,J=7.0Hz,6H).

[0109] Example 6

[0110] 1-((((S)-2,3-dihydroxypropoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0111]

[0112] Step A: 1-Chloroethyl (((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methyl) carbonate

[0113]

[0114] Under ice bath, (S)-(+)-1,2-isopropylidene glycerol (315 mg, 2.38 mmol) and pyridine (225 mg, 2.85 mmol) were added to 10 ml of dry dichloromethane, and 1-chloroethyl chloroformate (408 mg, 2.85 mmol) was added dropwise under ice bath. After the addition was completed, the mixture was heated to room temperature and reacted for 2 hours.

[0115] After the reaction, the mixture was poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 10 / 1) to give 250 mg of colorless oily product 1-chloroethyl (((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methyl) carbonate (yield: 44.2%).

[0116] Step B: Synthesis of 1-(((((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0117]

[0118] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (100 mg, 0.20 mmol), 1-chloroethyl (((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methyl) carbonate (72 mg, 0.3 mmol), cesium carbonate (130 mg, 0.4 mmol) and potassium iodide (66.4 mg, 0.4 mmol) were added to 5 mL of dry DMF at room temperature and heated to 65 °C for 4 h.

[0119] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-cold water solution. Extraction was performed with dichloromethane (20 ml x 3). The combined organic phases were washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 3 / 1) to obtain 85 mg of colorless oily product, 1-(((((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 59.9%). LC-MS: RT = 2.21 min, [M+H] + =713.46.

[0120] Step C: Synthesis of 1-((((S)-2,3-dihydroxypropoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0121]

[0122] At room temperature, 1-(((((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (80 mg, 0.11 mmol) was dissolved in 10 ml of dichloromethane solvent, 2 ml of trifluoroacetic acid solution was added, and the reaction was carried out at room temperature for 0.5 hour.

[0123] After the reaction, the crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 2) to obtain 56 mg of a yellow solid 1-((((S)-2,3-dihydroxypropoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 74.1%).

[0124] LC-MS: RT = 2.06 min, [M+H] + =673.39. 1 H NMR (400MHz, DMSO) δ7.32–7.17(m,2H),7.12–7.04(m,1H),6.93(s,2H),6.84(s,2H),6. 55(s,1H),6.00(d,J=5.2Hz,2H),4.18–4.06(m,2H),3.90(s,1H),3.73(s,3H),3.64(s,2 H),3.55–3.42(m,2H),3.18–3.07(m,3H),3.00(s,4H),2.66(s,1H),2.44–2.35(m,2H), 2.32(s,1H),1.31(s,3H),1.18(s,3H),0.99(s,3H),0.81(t,J=7.4Hz,3H),0.74(s,3H).

[0125] Example 7

[0126] 1-[(Cyclopropyloxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0127]

[0128] Step A: Synthesis of 1-[(cyclopropyloxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0129]

[0130] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (500 mg, 0.98 mmol), 1-chloroethylcyclopropyl carbonate (240 mg, 1.96 mmol), cesium carbonate (640 mg, 1.96 mmol) and potassium iodide (325 mg, 1.96 mmol) were added to 10 mL of dry DMF at room temperature and heated to 65 °C for 2 h.

[0131] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-water solution. Extraction was performed with dichloromethane (100 ml x 3). The combined organic phases were washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 1) to obtain 412 mg of colorless oily product, 1-[(cyclopropyloxycarbonyl)oxy]ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 65.8 g). LC-MS: RT = 2.29 min, [M+H] + =639.40. 1 H NMR (400MHz, DMSO) δ7.25 (s, 2H), 7.06 (d, J = 10.4Hz, 1H), 6.90 (m, 2H), 6.83 (m, 2H), 6.57 (m, 1H), 5.98 (s, 2H), 4.09 (s, 2H), 3.73 (m, 4H), 3.5 3–3.42(m,2H),2.95(m,5H),2.71(d,J=13.2Hz,3H),1.98(s,1H),1.38–1.02(m,9H),0.99–0.87(m,2H),0.81(s,2H),0.71(d,J=7.0Hz,6H).

[0132] Example 8

[0133] 1-(((2-Hydroxyethoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0134]

[0135] Step A: Synthesis of 2-((tert-butyldimethylsilyl)oxy)ethyl(1-chloroethyl)carbonate

[0136]

[0137] Under ice bath, 2-tert-butyldimethylsilyloxyethanol (1000 mg, 5.67 mmol) and pyridine (739 mg, 9.36 mmol) were added to 10 ml of dry dichloromethane, and 1-chloroethyl chloroformate (900 mg, 6.24 mmol) was added dropwise under ice bath. After the addition was completed, the temperature was raised to room temperature and the reaction was carried out for 1 hour.

[0138] After the reaction, the mixture was poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 10 / 1) to give 245 mg of colorless oily product 2-((tert-butyldimethylsilyl)oxy)ethyl (1-chloroethyl) carbonate (yield: 15.4%).

[0139] Step B: Synthesis of 2,2,3,3-tetramethyl-8-oxo-4,7,9-trioxa-3-sila-10-ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolane-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0140]

[0141] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (100 mg, 0.20 mmol), 2-((tert-butyldimethylsilyl)oxy)ethyl(1-chloroethyl) carbonate (85 mg, 0.3 mmol), cesium carbonate (130 mg, 0.4 mmol) and potassium iodide (66.4 mg, 0.4 mmol) were added to 5 mL of dry DMF at room temperature and heated to 65 °C for 4 h.

[0142] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-water solution. Extraction was performed with dichloromethane (20 ml x 3). The organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 3 / 1) to obtain 62 mg of colorless oily product, 2,2,3,3-tetramethyl-8-oxo-4,7,9-trioxa-3-sila-10-ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 41.0%). LC-MS: RT = 2.75 min, [M+H] + =757.50.

[0143] Step C: Synthesis of 1-(((2-hydroxyethoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0144]

[0145] At room temperature, 2,2,3,3-tetramethyl-8-oxo-4,7,9-trioxa-3-sila-10-ethyl-(2R,3R,4SS)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (62 mg, 0.08 mmol) was dissolved in 10 ml of dichloromethane solvent. 2 ml of 4 M / L hydrochloric acid dioxane solution was added under ice-cooling and the reaction was carried out at room temperature for 2 hours.

[0146] After the reaction, the crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 1 / 2) to obtain 40 mg of a yellow solid 1-(((2-hydroxyethoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 77.9%).

[0147] LC-MS: RT = 2.07 min, [M+H] + =643.44. 1H NMR (400MHz, DMSO) δ7.71–7.38(m,2H),7.19(d,J=12.2Hz,1H),7.00(d,J=6.8Hz,2H),6.93–6.79(m,2H),6.48(s,1H),6.03(s,2H), 4.02(d,J=7.1Hz,3H),3.76(m,4H),3.57–3.48(m,3H),3.20–3.09(m,1H),3.08–2.90(m,3H),1.45–1.02(m,14H),0.91–0.68(m,8H).

[0148] Example 9

[0149] 1-(((((R)-1,4-dioxan-2-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0150]

[0151] Step A: ((R)-1,4-dioxan-2-yl)methyl (1-chloroethyl) carbonate

[0152]

[0153] Under ice bath, (R)-(1,4-dioxane-2-yl)methanol (280 mg, 2.38 mmol) and pyridine (225 mg, 2.85 mmol) were added to 10 ml of dry dichloromethane, and 1-chloroethyl chloroformate (408 mg, 2.85 mmol) was added dropwise under ice bath. After the addition was completed, the temperature was raised to room temperature and the reaction was carried out for 2 hours.

[0154] After the reaction, the mixture was poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 10 / 1) to give 212 mg of a colorless oily product ((R)-1,4-dioxan-2-yl)methyl (1-chloroethyl) carbonate (yield: 39.8%).

[0155] Step B: Synthesis of 1-(((((R)-1,4-dioxan-2-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0156]

[0157] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (100 mg, 0.20 mmol), ((R)-1,4-dioxan-2-yl)methyl(1-chloroethyl)carbonate (67.2 mg, 0.3 mmol), cesium carbonate (130 mg, 0.4 mmol) and potassium iodide (66.4 mg, 0.4 mmol) were added to 5 mL of dry DMF at room temperature and heated to 65 °C for 4 h.

[0158] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), and the organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 3 / 1) to give 80 mg of colorless oily product 1-(((((R)-1,4-dioxan-2-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 59.9%).

[0159] LC-MS: RT = 2.16 min, [M+H] + =699.42. 1H NMR (400MHz, DMSO) δ7.24(dd,J=11.6,8.6Hz,2H),7.06(d,J=11.4Hz,1H),6.91(dd,J=8.6,2.7Hz,2H),6.85–6.75(m,2H) ,6.59–6.53(m,1H),5.99(d,J=6.0Hz,2H),4.06(dd,J=9.5,4.7Hz,2H),3.75–3.56(m,8H),3.53(d,J=9.9Hz,1H),3.50–3. 41(m,2H),3.29–3.21(m,5H),2.95(dd,J=18.3,10.6Hz,3H),2.79–2.73(m,1H),2.69(d,J=14.1Hz,1H),1.33(t,J=8.5Hz, 3H),1.29–1.21(m,3H),1.17(dd,J=14.8,7.4Hz,3H),0.96(d,J=4.7Hz,2H),0.81(t,J=7.3Hz,3H),0.71(t,J=7.3Hz,3H).

[0160] Example 10

[0161] 1-((((oxetan-3-yl)oxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0162]

[0163] Step A: 1-Chloroethyloxetan-3-yl carbonate

[0164]

[0165] Under ice bath, oxetane-3-ol (176 mg, 2.38 mmol) and pyridine (225 mg, 2.85 mmol) were added to 10 ml of dry dichloromethane, and 1-chloroethyl chloroformate (408 mg, 2.85 mmol) was added dropwise under ice bath. After the addition was completed, the mixture was warmed to room temperature and reacted for 2 hours.

[0166] After the reaction, the mixture was poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, concentrated under reduced pressure and evaporated to dryness. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 10 / 1) to give 200 mg of colorless oily product 1-chloroethyloxetane-3-yl carbonate (yield: 46.7%).

[0167] Step B: Synthesis of 1-((((oxetan-3-yl)oxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0168]

[0169] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (100 mg, 0.2 mmol), 1-chloroethyloxetane-3-yl carbonate (54 mg, 0.3 mmol), cesium carbonate (130 mg, 0.4 mmol) and potassium iodide (66.4 mg, 0.4 mmol) were added to 5 mL of dry DMF at room temperature and heated to 65 °C for 4 h.

[0170] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), and the organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 3 / 1) to give 76 mg of colorless oily product 1-((((oxetane-3-yl)oxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 58.0%).

[0171] LC-MS: RT = 2.18 min, [M+H] + =655.39. 1H NMR (400MHz, DMSO) δ7.24(t,J=8.4Hz,2H),7.05(d,J=12.1Hz,1H),6.94–6.86(m,2H),6.85–6.71(m,2H),6.61–6.50(m ,1H),5.99(d,J=5.7Hz,2H),5.41–5.26(m,1H),4.82–4.63(m,2H),4.53–4.32(m,2H),3.80–3.60(m,4H),3.57–3.41(m ,1H),3.27–3.11(m,4H),2.98(dd,J=14.1,6.6Hz,3H),2.82–2.74(m,1H),2.68(s,1H),1.32(t,J=7.1Hz,3H),1.26(dd ,J=18.2,7.3Hz,3H),1.17(dd,J=14.8,7.1Hz,3H),0.96(d,J=6.6Hz,2H),0.81(t,J=7.3Hz,3H),0.71(t,J=7.3Hz,3H).

[0172] Example 11

[0173] 1-(((((S)-1,4-dioxan-2-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0174]

[0175] Step A: ((S)-1,4-dioxan-2-yl)methyl (1-chloroethyl) carbonate

[0176]

[0177] Under ice bath, (S)-(1,4-dioxane-2-yl)methanol (280 mg, 2.38 mmol) and pyridine (225 mg, 2.85 mmol) were added to 10 ml of dry dichloromethane, and 1-chloroethyl chloroformate (408 mg, 2.85 mmol) was added dropwise under ice bath. After the addition was completed, the temperature was raised to room temperature and the reaction was carried out for 2 hours.

[0178] After the reaction, the mixture was poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 10 / 1) to give 219 mg of a colorless oily product ((S)-1,4-dioxan-2-yl)methyl (1-chloroethyl) carbonate (yield: 41.0%).

[0179] Step B: Synthesis of 1-(((((S)-1,4-dioxan-2-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate

[0180]

[0181] Atrasentan (2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylic acid (100 mg, 0.20 mmol), ((S)-1,4-dioxan-2-yl)methyl(1-chloroethyl)carbonate (44.8 mg, 0.3 mmol), cesium carbonate (130 mg, 0.4 mmol) and potassium iodide (66.4 mg, 0.4 mmol) were added to 5 mL of dry DMF at room temperature and heated to 65 °C for 4 h.

[0182] After the reaction, the mixture was cooled to room temperature and poured into 40 ml of ice-water solution, extracted with dichloromethane (20 ml × 3), and the organic phases were combined, washed with saturated brine (50 ml), dried over anhydrous magnesium sulfate, filtered, and concentrated to dryness under reduced pressure. The crude compound was purified by silica gel column chromatography (eluent: n-hexane / ethyl acetate = 3 / 1) to give 92 mg of colorless oily product 1-(((((S)-1,4-dioxan-2-yl)methoxy)carbonyl)oxy)ethyl-(2R,3R,4S)-4-(benzo[d][1,3]dioxolan-5-yl)-1-[2-(dibutylamino)-2-oxoethyl]-2-(4-methoxyphenyl)pyrrolidine-3-carboxylate (yield: 65.8%).

[0183] LC-MS: RT = 2.14 min, [M+H] + =699.43. 1H NMR (400MHz, DMSO) δ7.24(dd,J=11.6,8.7Hz,2H),7.06(d,J=10.7Hz,1H),6.91(dd,J=8.6,2.7Hz,2H),6.85–6.70( m,2H),6.59–6.53(m,1H),5.99(d,J=6.2Hz,2H),4.09–3.97(m,2H),3.75–3.65(m,6H),3.56(dd,J=22.9,9.5Hz,3H ),3.47–3.38(m,2H),3.23(dd,J=31.0,20.1Hz,5H),2.96(dt,J=16.3,8.0Hz,3H),2.84–2.62(m,2H),1.33(t,J=8. 2Hz,3H),1.29–1.20(m,3H),1.19–1.07(m,3H),0.96(d,J=7.4Hz,2H),0.81(t,J=7.3Hz,3H),0.71(t,J=7.3Hz,3H).

[0184] Example 12

[0185] Microsomal studies of compounds

[0186] (1) Experimental materials

[0187] Human liver microsomes were purchased from Red Liver Disease Research (Shanghai) Co., Ltd.

[0188] Reagents: DMSO (dimethyl sulfoxide), acetonitrile, formic acid, and propranolol (internal standard) are all commercially available.

[0189] Instrument: Thermo Fisher LC-MS (U300UPLC, TSQ QUANTUMN ULTRA triple quadrupole mass spectrometer).

[0190] (2) Experimental methods

[0191] Accurately weigh a certain amount of compound and dissolve it in DMSO to prepare a 10 mM stock solution. Dilute the stock solution to a 100 μM working solution with diluent (ACN:H₂O = 1:1). Then, dilute the solution to a 3 μM dosing solution with 0.1 M potassium phosphate buffer for later use. Add 75 μL of liver microsomes to 925 μL of 0.1 M potassium phosphate buffer, mix well, and prepare a 1.5 mg / mL liver microsomal suspension. Preincubate at 37°C for 10 min. To prepare the zero-point sample, take 15 μL of the liver microsomal suspension and add 6 mM NADPH solution. Immediately add 150 μL of propranolol acetonitrile solution for precipitation. Then add 15 μL of the dosing solution and mix well. To prepare the 20-min and 60-min samples, take 15 μL of the dosing solution and add 15 μL of the liver microsomal suspension and 15 μL of the 6 mM NADPH solution. Mix well and incubate at 37°C for 20 and 60 min, respectively. All samples were prepared in duplicate. At the relevant time points, 150 μL of propranolol acetonitrile solution was added to terminate the reaction. All samples were centrifuged at 4000 rpm for 5 minutes. 100 μL of the supernatant was added to 100 μL of ultrapure water and mixed before LC-MS / MS analysis. LC-MS / MS analysis conditions were as follows:

[0192] Chromatographic column: Waters ACQUITY™ PREMIER HSS T3, 50*2.1 mm, 1.8 μm.

[0193] Mobile phase: Water (0.1% formic acid)-acetonitrile Gradient elution according to the table below

[0194]

[0195]

[0196] (3) Data processing

[0197] Taking the initial 0 point as 100%, the relative remaining content of the drug at each time point was calculated, and taking the atrasentan at each time point as 100%, the relative amount of the example compound converted to atrasentan was calculated. The results are shown in Tables 1 and 2.

[0198] Table 1. Changes of prototypes of various drugs in microsomes

[0199]

[0200]

[0201] Table 2. Amount of Example Compounds Converted into Atrasentan in Microsomes

[0202]

[0203] The results showed that the compound of Example 3 could be rapidly metabolized in human microsomes and could be completely converted into atrasentan.

[0204] Example 13

[0205] Pharmacokinetic study of the compound in rats

[0206] (1) Experimental materials

[0207] SD rats: male, 200-300 g, purchased from Beijing Weitonglihua Experimental Animal Technology Co., Ltd.

[0208] Reagents: DMSO (dimethyl sulfoxide), PEG-400 (polyethylene glycol 400), normal saline, heparin, acetonitrile, formic acid, and propranolol (internal standard) are all commercially available.

[0209] Instrument: Thermo Fisher LC-MS / MS (U300UPLC, TSQ QUANTUMN ULTRA triple quadrupole mass spectrometer).

[0210] (2) Experimental methods

[0211] The compound was weighed and dissolved in DMSO-PEG-400-saline (5:60:35, v / v / v). After oral administration to rats, 200 μL of venous blood was collected at 15 minutes, 30 minutes, 1 hour, 2 hours, 5 hours, 7 hours, and 24 hours after administration into heparinized EP tubes with sodium fluoride. The blood was centrifuged at 12,000 rpm for 2 minutes, and plasma was frozen at -80°C for analysis. A precisely weighed amount of the test compound was dissolved in DMSO to 2 mg / mL to prepare a stock solution. An appropriate amount of the stock solution was accurately aspirated and diluted with acetonitrile to prepare a series of standard solutions. 20 μL of each standard solution was accurately aspirated and added to 180 μL of blank plasma. The solution was vortexed to mix thoroughly. Plasma samples were prepared at concentrations equivalent to 1, 3, 5, 10, 30, 100, 300, 1000, and 3000 ng / mL. Duplicate samples were analyzed at each concentration to construct a standard curve. Take 30 μL of plasma, add 200 μL of acetonitrile solution of internal standard propranolol (50 ng / mL), vortex mix, add 100 μL of purified water, vortex mix again, centrifuge at 4000 rpm for 5 minutes, and collect the supernatant for LC-MS / MS analysis. LC-MS / MS detection conditions are as follows:

[0212] Chromatographic column: Waters ACQUITY™ PREMIER HSS T3, 50*2.1 mm, 1.8 μm.

[0213] Mobile phase: Water (0.1% formic acid)-acetonitrile Gradient elution according to the table below

[0214]

[0215] (3) Data processing

[0216] After LC-MS / MS detection of blood drug concentration, WinNonlin 6.1 software was used to calculate the pharmacokinetic parameters using the non-compartmental model method. The results are shown in Table 3

[0217] Table 3 Pharmacokinetic parameters of atrasentan in SD rats after oral administration of atrasentan, Example 3 and Example 4

[0218]

[0219] Note: The dosage is converted to atrasentan

[0220] The exposure levels of the compounds of Examples 3 and 4 in rats were higher than that of atrasentan at the same dose, and their absorption after oral administration was better than that of atrasentan.

[0221] Example 14

[0222] Pharmacokinetic study of the compound in rats

[0223] (1) Experimental materials

[0224] SD rats: male, 200-300 g, purchased from Beijing Weitonglihua Experimental Animal Technology Co., Ltd.

[0225] Reagents: DMSO (dimethyl sulfoxide), PEG-400 (polyethylene glycol 400), normal saline, heparin, acetonitrile, formic acid, and propranolol (internal standard) are all commercially available.

[0226] Instrument: Thermo Fisher LC-MS / MS (U300UPLC, TSQ QUANTUMN ULTRA triple quadrupole mass spectrometer).

[0227] (2) Experimental methods

[0228] The compound was weighed and dissolved in DMSO-PEG-400-saline (5:60:35, v / v / v). After oral administration to rats, 200 μL of venous blood was collected at 15 minutes, 30 minutes, 1 hour, 2 hours, 5 hours, 7 hours, and 24 hours after administration into heparinized EP tubes with sodium fluoride. The blood was centrifuged at 12,000 rpm for 2 minutes, and plasma was frozen at -80°C for analysis. A precisely weighed amount of the test compound was dissolved in DMSO to 2 mg / mL to prepare a stock solution. An appropriate amount of the stock solution was accurately aspirated and diluted with acetonitrile to prepare a series of standard solutions. 20 μL of each standard solution was accurately aspirated and added to 180 μL of blank plasma. The solution was vortexed to mix thoroughly. Plasma samples were prepared at concentrations equivalent to 1, 3, 5, 10, 30, 100, 300, 1000, and 3000 ng / mL. Duplicate samples were analyzed at each concentration to construct a standard curve. Take 30 μL of plasma, add 200 μL of acetonitrile solution of internal standard propranolol (50 ng / mL), vortex mix, add 100 μL of purified water, vortex mix again, centrifuge at 4000 rpm for 5 minutes, and collect the supernatant for LC-MS / MS analysis. LC-MS / MS detection conditions are as follows:

[0229] Chromatographic column: Waters ACQUITY™ PREMIER HSS T3, 50*2.1 mm, 1.8 μm.

[0230] Mobile phase: Water (0.1% formic acid)-acetonitrile Gradient elution according to the table below

[0231]

[0232] (3) Data processing

[0233] After LC-MS / MS detection of blood drug concentration, WinNonlin 6.1 software was used to calculate the pharmacokinetic parameters using the non-compartmental model method. The results are shown in Table 4

[0234] Table 4 Pharmacokinetic parameters of atrasentan and Example 7 after oral administration to SD rats

[0235]

[0236] The exposure of the compound of Example 7 in rats was significantly lower than that of atrasentan at the same dosage, and its absorption after oral administration was lower than that of atrasentan.

[0237] Based on Examples 12, 13 and 14, in general, the compound of Example 3 can be rapidly converted into atrasentan both in vivo and in vitro, and the exposure is higher than that of atrasentan under oral equimolar doses.

[0238] Example 15 Salt Screening Example

[0239] Compound A was dissolved in different solvents such as isopropanol, acetone, and ethyl acetate, and acids (17 acids in total, as shown in the table below) were added at the same molar ratio. After a certain reaction time, the solution became clear, and solids were precipitated by adding antisolvents and lowering the temperature.

[0240] Table 5 Screening of 17 acid ligands

[0241]

[0242] Compound A is an oily substance that can only precipitate as a solid after forming salts with a few acids. As can be seen from the table above, compound A is quite difficult to form salts with, as only a few acids—sulfuric acid, phosphoric acid, oxalic acid, and hydrochloric acid—can form salts.

[0243] Example 16 Preparation of Oxalate

[0244] The free base of compound A (3.5 g) was dissolved in isopropanol (10 ml), and oxalic acid dihydrate (689 mg) was added. The reaction was stirred at room temperature for 1 h and concentrated to obtain an oil. Isopropyl ether (50 ml) was added under ice bath and stirred to precipitate a solid. The mixture was stirred under ice bath for 1 day and filtered to obtain an oxalate solid.

[0245] Compound A free base (2.4 g) was reacted with oxalic acid dihydrate (46 mg) in ethyl acetate (1 w / w) to form a salt, which was added dropwise to isopropyl ether (15 w / w) at -5°C, stirred for 1 day, filtered, and dried to obtain a solid.

[0246] Compound A free base (2.2 g) was reacted with oxalic acid (70 mg) in isopropanol (2 ml), concentrated, and isopropyl acetate (0.5 ml) was added. Isopropyl ether (15 ml) was added at -15°C, stirred for 4 h, and filtered to obtain a solid.

[0247] NMR data of oxalate:

[0248]

[0249] Attribution:

[0250]

[0251] Analysis:

[0252] δ = 0.93 is the methyl hydrogen on carbon atoms 34 and 37, with 6 protons, which is two sets of triplet peaks;

[0253] δ = 1.28 is the methyl hydrogen on the carbon atom No. 45, with 3 protons, and is a set of doublets;

[0254] δ = 1.34 is the methyl hydrogen on C atom No. 46, and the methylene hydrogen on C atoms No. 33 and 36, with 7 protons, which are one triplet peak and two doublet peaks respectively;

[0255] δ = 1.52 is the methylene hydrogen on carbon atoms 32 and 35, with 4 protons, which is two sets of triplet peaks;

[0256] δ = 1.59 is the methyl hydrogen on the 39th carbon atom, with 3 protons, and is a set of doublets;

[0257] δ = 3.24, 3.29, 3.31, 3.35, 3.41, 3.59, 3.68 are respectively the methylene hydrogens on carbon atoms 30, 31, 26, 11, 9, and 10, with 12 protons, which are three sets of multiple peaks;

[0258] δ = 3.78 is the methyl hydrogen on the 25th carbon atom, with 3 protons, which is a group of single peaks;

[0259] δ = 4.39 is the methine hydrogen on the 7th carbon atom, with a proton number of 1, which is two sets of doublet peaks;

[0260] δ = 5.08 is the methine hydrogen on the 44th carbon atom, with a proton number of 1, which is a group of multiple peaks;

[0261] δ = 5.93 is the methylene hydrogen on the 20th carbon atom, with 2 protons, forming a triplet peak;

[0262] δ=6.63, 6.72, 6.82 are respectively the methine hydrogen on carbon atoms 18, 15, and 14, with 3 protons, which are three sets of doublet peaks;

[0263] δ = 6.91 is the methine hydrogen on the C atoms of benzene rings 4 and 6, with 2 protons, and is two sets of triplet peaks;

[0264] δ = 7.20 is the methine hydrogen on the C atoms of benzene rings 1 and 3, with 2 protons, and is two sets of triplet peaks;

[0265] δ = 7.51 is the methine hydrogen on C atom No. 38, with a proton number of 1, which is a group of multiple peaks;

[0266] The peak with δ=11.05 is the carboxyl hydrogen on the C atom of oxalic acid, which has 2 protons and is a group of single peaks.

[0267] Example 17 Preparation of Phosphate

[0268] The free base of compound A (4.1 g) was dissolved in isopropanol (12 ml), phosphoric acid (800 mg) was added and the mixture was reacted for 1 h. Isopropyl ether (80 ml) was added under ice bath and stirred for 4 h to obtain a phosphate solid.

[0269] Phosphate NMR data:

[0270]

[0271] Attribution:

[0272]

[0273] Analysis:

[0274] δ = 0.73, 0.83 are the methyl hydrogens on carbon atoms 34 and 37, with 6 protons, which are two sets of triplet peaks;

[0275] δ = 1.25 is the methyl hydrogen on C atoms 45, 46, and 39, the methylene hydrogen on C atoms 33 and 36, and the methylene hydrogen on C atoms 32 and 35. The number of protons is 14, which is a multiplet.

[0276] δ = 2.78, 2.99, 3.19, 3.31, 3.50 are respectively the methylene hydrogens on carbon atoms 30, 31, 26, 11, 9, and 10, with 12 protons, which are three sets of multiple peaks;

[0277] δ = 4.75 is the methine hydrogen on the 7th carbon atom, with a proton number of 1, which is two sets of doublet peaks;

[0278] δ = 6.00 is the methylene hydrogen on the 20th carbon atom, with 2 protons, forming a triplet peak;

[0279] δ=6.57, 6.84 are respectively the methine hydrogen on carbon atoms 18, 15, and 14, with 3 protons, forming three sets of doublets;

[0280] δ = 6.92 is the methine hydrogen on the C atoms of benzene rings 4 and 6, with 2 protons, and is two sets of triplet peaks;

[0281] δ=7.06, 7.27 are the methine hydrogen on the C atoms of benzene ring No. 1 and No. 3, and the methine hydrogen on the C atom No. 38. The number of protons is 3, which is two sets of triplet peaks.

[0282] Elemental analysis: P element content (%) = 6.46, relative standard deviation (%) = 0.2.

[0283] Detection instrument: full spectrum direct reading plasma atomic force spectrometer, iCAP6500Duo (Thermo, USA);

[0284] Detection conditions: Incident power: 1150W Plasma gas flow rate: 14L / min Atomizer flow rate: 0.5L / min.

[0285] Example 18 Preparation of hydrochloride

[0286] Weigh Compound A free base (202.5 mg) into a 5 mL vial, add MeOH (1.0 mL) and an equimolar ratio of methanolic hydrochloric acid. Suspend and stir at -20°C for 1 day. Remove the methanol by rotary evaporation. The sample becomes a gelatinous substance, which remains after vacuum drying. Add isopropyl ether (2.0 mL), suspend and stir at -20°C for 1 day, and a solid precipitates. Centrifuge and vacuum dry to obtain a solid.

[0287] NMR spectra such as Figure 2 shown.

[0288] Example 19 Preparation of sulfate

[0289] Compound A free base (203.1 mg) was weighed into a 5 mL vial. IPA (1.0 mL) and an equimolar ratio of sulfuric acid were added. The mixture was suspended and stirred at -20°C for 1 day. The IPA was then removed by vacuum drying, resulting in a gel-like sample. Isopropyl ether (2.0 mL) was added, and the mixture was suspended and stirred at -20°C for 1 day. A solid precipitated, which was then centrifuged and dried under vacuum to obtain a solid.

[0290] NMR spectra such as Figure 3 shown.

[0291] Example 20 Stability Study

[0292] Sulfate, phosphate, hydrochloride and oxalate were placed under 25°C / 60% RH and 40°C / 75% RH conditions for 1 week, respectively, and the chemical stability of the samples was tested by HPLC.

[0293]

[0294] As can be seen from the above table, the oxalate and phosphate obtained by the present invention have good stability and are better than hydrochloride and sulfate.

[0295] Example 21 Hygroscopicity Study

[0296] Sulfate, phosphate, hydrochloride, and oxalate were evaluated for their hygroscopicity using a dynamic moisture sorption instrument (DVS). The test measured the percentage change in mass of the samples at a constant temperature of 25°C as humidity varied. The DVS test results are as follows:

[0297]

[0298] As can be seen from the above table, the oxalate and phosphate obtained by the present invention have lower hygroscopicity and are better than hydrochloride and sulfate.

[0299] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered as equivalent replacement methods and are included in the scope of protection of the present invention.

Claims

1. A salt of a compound represented by formula (I), characterized in that M is an inorganic acid or an organic acid, wherein n:x=1:1-2; The inorganic acid is selected from sulfuric acid, hydrochloric acid, and phosphoric acid; and the organic acid is selected from oxalic acid.

2. The salt of the compound represented by formula (I) according to claim 1, characterized in that The inorganic acid is selected from phosphoric acid.

3. The salt of the compound represented by formula (I) according to claim 1, characterized in that The inorganic acid is selected from phosphoric acid, wherein n=3, x=3-6.

4. The salt of the compound represented by formula (I) according to claim 1, characterized in that The inorganic acid is selected from phosphoric acid, wherein n=3, x=3; or n=3, x=4; or n=3, x=5; or n=3, x=6.

5. The salt of the compound represented by formula (I) according to claim 1, characterized in that The organic acid is selected from oxalic acid.

6. The salt of the compound represented by formula (I) according to claim 1, characterized in that The organic acid is selected from oxalic acid, wherein n=1, x=1.

7. A method for preparing a salt of a compound according to any one of claims 1 to 6, characterized in that: The compound A Prepared by mixing with acid.

8. The preparation method according to claim 7, characterized in that The molar ratio of the compound to the acid molecule is 1:1-2.

9. The preparation method according to claim 7, comprising the steps of: 1) Weigh an appropriate amount of free base and dissolve it in a benign solvent; 2) Weighing an appropriate amount of ionic acid and adding it to step 1) to react and form a salt; 3) back-titrifying the reaction solution of step 2) into a poor solvent and stirring to precipitate; 4) Rapidly centrifuge or allow to stand to obtain the salt of the compound.

10. A pharmaceutical composition comprising a salt of the compound represented by formula (I) according to any one of claims 1 to 6, and one or more pharmaceutically acceptable carriers.

11. Use of a salt of the compound represented by formula (I) according to any one of claims 1 to 6 in the preparation of a medicament for treating and / or preventing endothelin A receptor antagonist-related diseases.

12. The use according to claim 11, characterized in that Such diseases include chronic kidney disease, IgA, FSGS, Alport's disease and hypertension.

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