Novel isoxazolopyrimidine derivative with activity of resisting new coronavirus as well as preparation method and application of novel isoxazolopyrimidine derivative

Abstract

The invention discloses a novel isoxazolopyrimidine derivative with anti-new coronavirus activity as well as a preparation method and application of the novel isoxazolopyrimidine derivative. The isoxazolopyrimidine compound provided by the invention has a good inhibition effect on SARS-CoV-2, and relates to the compound shown in the general formula (I), and pharmaceutically acceptable salts and solvent compounds thereof have a good inhibition effect on new coronavirus, and can be used for preparing antiviral drugs related to the new coronavirus for prevention, treatment or adjuvant treatment. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a novel isoxazolopyrimidine derivative with anti-new coronavirus activity, and a preparation method and application thereof. Background Art

[0002] Viruses, especially RNA viruses, occupy an important position in the top ten global health threats currently listed by the World Health Organization (WHO). More than 200 viruses are known to infect humans. The diseases that currently threaten human health include AIDS caused by human immunodeficiency virus (HIV), hepatitis caused by hepatitis C virus (HCV), global influenza caused by influenza A virus (IAV), dengue virus (DENV) and viral diseases caused by Ebola virus (EBOV) (Chem. Soc. Rev., 2021, 50, 3647-3655.). Since the approval of the first antiviral drug Idoxuridine in 1963, a total of nearly 100 antiviral drugs have been officially approved for the treatment of 9 human infectious diseases (Clin. Microbiology Rev., 2016, 29, 695-747.). However, these antiviral drugs are only used in the routine treatment of HIV, HCV and IAV. Since the outbreak of the new coronavirus, the virus has spread rapidly, further exposing the lack of antiviral drugs for the treatment of recurring or emerging viral diseases. Between 2020 and 2023, a number of chemical drugs have been approved for the treatment of SARS-CoV-2, such as Remdesivir, Baricitinib, Azvudine, and Paxlovid. These drugs mainly block the viral replication process by targeting viral proteins (especially viral enzymes) or host proteins (these proteins are involved in the life cycle of the virus), but because the virus strains mutate, this will lead to reduced drug selectivity, poor safety, and drug resistance (DrugDiscov.Today, 2024, 29, 104126.). So far, the search for drugs against SARS-CoV-2 is still ongoing, so the development of new anti-SARS-CoV-2 viral drugs has become a hot topic in the field of medicinal chemistry research. How to develop a new drug with a broad spectrum and good inhibitory effect on SARS-CoV-2 is an urgent problem to be solved by those skilled in the art. Summary of the invention

[0003] The purpose of the present invention is to provide a novel isoxazolopyrimidine derivative with anti-new coronavirus activity and a preparation method and application thereof, aiming to solve the problems raised in the above-mentioned background technology.

[0004] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0005] The present invention provides a novel isoxazolopyrimidine derivative having anti-new coronavirus activity, and a pharmaceutically acceptable salt, solvent compound or deuterated product thereof. The structural formula of the novel isoxazolopyrimidine derivative having anti-new coronavirus activity is shown in general formula I:

[0006]

[0007] The R 1 is selected from substituted or unsubstituted alkyl or cycloalkyl, substituted or unsubstituted aryl;

[0008] The R 2 is selected from substituted or unsubstituted alkyl or cycloalkyl, substituted or unsubstituted aryl;

[0009] The R 3 is selected from substituted or unsubstituted morpholinyl, substituted or unsubstituted thiomorpholinyl, substituted or unsubstituted piperidinyl, substituted or unsubstituted piperazinyl, substituted or unsubstituted pyridazinyl, substituted or unsubstituted pyrrolyl, substituted or unsubstituted imidazolyl, substituted or unsubstituted pyrazolyl, substituted or unsubstituted amine;

[0010] R 1 and R 2 The alkyl, cycloalkyl and aryl groups are optionally substituted with 1, 2, 3, 4 or 5 R; R is selected from hydrogen, deuterium, alkene, alkynyl, halogen, hydroxy, hydroxymethyl, amino, trifluoromethyl, cyano, ester, carbonyl, acyl, aminoacyl, amide, sulfonyl, aminosulfonyl, sulfonamido, alkyl, alkoxy, urea, thiourea, aryl, pyridyl, pyrimidinyl, pyridazinyl, aminopyrimidinyl, quinolyl, benzofuranyl, benzindolyl, benzindazolyl, benzothiophenyl, benzopyrazolyl, furanyl, thienyl, pyridyl, oxazolyl and pyrazolyl.

[0011] Furthermore, R 1 Selected from cyclopentyl and 4-piperidinyl.

[0012] Furthermore, R 2 Selected from 3-hydroxymethylphenyl, 4-aminophenyl, 4-ureidophenyl.

[0013] Furthermore, R 3 Selected from morpholinyl.

[0014] Furthermore, the virus includes SARS-CoV-2.

[0015] The synthesis method of isoxazolo[4,5-d]pyrimidin-7(6H)-one and its derivatives comprises the following steps:

[0016] Step 1: adding 4-aminoisoxazole-5-carboxamide compounds into a reactor;

[0017] Step 2: adding an aldehyde compound into a reactor;

[0018] Step 3: Adding the catalyst into the reactor;

[0019] Step 4: Adding solvent into the reactor;

[0020] Step 5: After the mixed solution is reacted at reflux temperature for a certain period of time, the reaction is quenched;

[0021] Step 6: adding an organic solvent for extraction, drying and concentrating the organic phase, and purifying to obtain the isoxazolopyrimidine compound represented by the general formula I.

[0022] Preferably, in step 1, the reactor is a sealed tube;

[0023] Preferably, in step 3, the catalyst is I2 and DMSO;

[0024] Preferably, in step 4, the solvent is CH3CN;

[0025] Preferably, in step 5, the reaction is quenched with an aqueous solution of sodium thiosulfate;

[0026] Preferably, in step 6, ethyl acetate is added for extraction, and the organic phase is dried and concentrated, and then purified to obtain the isoxazolopyrimidine compound represented by general formula I.

[0027] The following terms are used throughout this document and are defined as follows:

[0028] The term "pharmaceutically acceptable salt" refers to a salt of the compound of the present invention that is pharmaceutically acceptable and can retain the biological effectiveness of the free base without other side effects. Such salts include: acid addition salts formed with inorganic acids or organic acids, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; organic acids such as acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, trifluoroacetic acid, formic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, camphorsulfonic acid, glucoheptonic acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid, etc.; or salts formed when the acidic protons present on the parent compound are replaced by metal ions, such as alkali metal ions or alkaline earth metal ions, such as sodium salts, potassium salts, calcium salts and magnesium salts, etc. Or a coordination compound formed with an organic base, such as ethanolamine, diethanolamine, triethanolamine, N-methylglucamine, etc. The pharmaceutically acceptable salt of the present invention can be synthesized by conventional chemical methods from a parent compound containing an acid radical or a base. In general, the preparation method of the salt is: in water or an organic solvent or a mixture of the two, these compounds in the form of free acid or base are reacted with a stoichiometric amount of an appropriate base or acid to prepare. Generally, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred. In addition to the form of salts, the compounds provided by the present invention also exist in the form of prodrugs. The prodrugs of the compounds described herein are easily chemically changed under physiological conditions to be converted into the compounds of the present invention. In addition, drug precursors can be converted to the compounds of the present invention by chemical or biochemical methods in an in vivo environment.

[0029] The term "solvate" refers to a substance formed by combining a compound of the present invention with a pharmaceutically acceptable solvent. Pharmaceutically acceptable solvents include water, ethanol, acetic acid, etc. Solvates include stoichiometric solvents and non-stoichiometric solvents, preferably hydrates. Certain compounds of the present invention may exist in unsolvated or solvated forms, including hydrate forms. Generally speaking, solvated forms are comparable to unsolvated forms and are included within the scope of the present invention.

[0030] When referring to certain elements such as hydrogen or H, it is intended to include all isotopes of that element. For example, if the R group is defined to include hydrogen or H, it also includes deuterium and tritium, and thus, isotopically labeled compounds are within the scope of the present invention.

[0031] The term "halogen" refers to fluorine, chlorine, bromine and iodine. The term "alkyl" refers to a straight or branched saturated hydrocarbon group. In some embodiments, the alkyl group is a C1-C3 alkyl group; in other embodiments, the alkyl group is a C1-C6 alkyl group. Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl (including isopropyl and n-propyl), butyl (including n-butyl, isobutyl), pentyl (including n-pentyl, isopentyl and neopentyl), and hexyl. The alkyl group may be monovalent (such as methyl), divalent (such as methylene), polyvalent (such as methine), and may be monosubstituted (such as -CH2F) or polysubstituted (-CF3).

[0032] The term "alkoxy" refers to a group having an -O-alkyl structure, wherein the alkyl group is as defined above. Specific examples include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, tert-butoxy, isobutoxy, n-pentoxy, and the like.

[0033] The term "alkenyl" refers to a straight or branched hydrocarbon group containing one or more carbon-carbon double bonds. In some embodiments, the alkenyl is a C2-C4 alkenyl; in other embodiments, the alkenyl is a C2-C6 alkenyl; in other embodiments, the alkenyl is a C2-C8 alkenyl.

[0034] The term "ring" refers to substituted or unsubstituted cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heterocycloalkyl, heterocycloalkenyl, heterocycloalkynyl, aryl. The ring includes monocyclic, bicyclic, polycyclic. The number of atoms on the ring is defined as the number of ring members.

[0035] The term "hetero" refers to heteroatoms or heteroatoms (atomic groups containing heteroatoms), that is, atoms other than carbon (C) and hydrogen (H) and atomic groups containing these heteroatoms, such as oxygen (O), nitrogen (N), sulfur (S), etc.

[0036] The term "cycloalkyl" includes heterocycloalkyl, which refers to cyclic alkyl groups including monocyclic, bicyclic or tricyclic ring systems, wherein the bicyclic or tricyclic ring system includes spirocyclic, bridged or fused rings; the alkyl group may be monovalent, divalent or polyvalent, and may be monosubstituted or polysubstituted.

[0037] The term "cycloalkenyl" refers to a cyclic alkenyl group containing one or more unsaturated carbon-carbon double bonds, wherein the bicyclic or tricyclic ring system includes spirocyclic, bridged, and fused rings; the alkyl group may be monovalent, divalent, or polyvalent, and may be monosubstituted or polysubstituted.

[0038] The term "cycloalkynyl" refers to a cyclic alkyl group containing one or more carbon-carbon triple bonds, including monocyclic, bicyclic, and polycyclic systems, wherein the bicyclic or tricyclic system includes spirocyclic, fused, and bridged rings. The alkyl group can be monovalent, divalent, or polyvalent, and can also be monosubstituted or polysubstituted.

[0039] The term "aromatic ring" or "aryl" includes heteroaryl and refers to a polyunsaturated ring system, including a monocyclic, bicyclic, or polycyclic system, wherein at least one ring is aromatic and may be monovalent, divalent, or polyvalent, and may be mono- or poly-substituted.

[0040] The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, which may include deuterium and hydrogen variants, as long as the valence state of the particular atom is normal and the substituted compound is stable. When the substituent is oxygen (i.e., =O), it means that two hydrogen atoms are replaced. Oxygen substitution does not occur on aromatic groups. The term "optionally substituted" means that it may be substituted or not substituted, and unless otherwise specified, the type and number of the substituents may be arbitrary on the basis of chemical achievable.

[0041] Compared with the prior art, the present invention has the following beneficial effects:

[0042] The present invention discloses a compound represented by general formula (I), a pharmaceutically acceptable salt thereof, and a solvent compound thereof, which have a good inhibitory effect on the new coronavirus and can be used to prepare antiviral drugs for the prevention, treatment or auxiliary treatment of the new coronavirus. DETAILED DESCRIPTION

[0043] The present invention will be further illustrated in the following examples, which are only used to illustrate the present invention but not to limit the present invention in any way.

[0044]

[0045] Related technical routes of this patent

[0046] Example 1: Synthesis of 4-nitroisoxazole-5-carboxylic acid ethyl ester s4

[0047]

[0048] Under N2 atmosphere, S1 (30 mmol) and CDI (9.72 g, 60 mmol) were dissolved in anhydrous THF (200 mL). The reaction mixture was stirred at room temperature for 1-2 h. Nitromethane (4 mL, 90 mmol) and DBU (20 mL, 135 mmol) were added in sequence. The reaction mixture was stirred at room temperature for 36 h. After dilution with ethyl acetate (100 mL), 2N HCl aqueous solution (150 mL) was added to the mixture, the pH value was adjusted to 1-2, and then extracted with ethyl acetate (200 mL×3). The combined organic layers were extracted with water (200 mL×3) and brine (200 mL×3), respectively, and dried over anhydrous Na2SO4 to obtain compound S2 with a yield of 95-98%, which was used in the next step without further purification.

[0049] Compound S2 (29 mmol), hydroxylamine hydrochloride (2.0 g, 29 mmol) and NaHCO3 (2.4 g, 29 mmol) were dissolved in ethanol (150 mL), and the mixture was stirred at 50 ° C for 3 hours. After the reaction of the raw materials was complete, the mixture was concentrated in vacuo, diluted with water (150 mL), and extracted with ethyl acetate (100 mL×3). The combined organic layer was then washed with H2O (150 mL×3) and brine (150 mL×3), dried over anhydrous Na2SO4 and concentrated under vacuum to obtain compound S3 with a yield of 95-99%, which was used in the next step without further purification.

[0050] Under N2 atmosphere, ethyl oxalyl chloride (4.5 mL, 40 mmol) was added dropwise to a solution of compound S3 (27 mmol) and triethylamine (5.45 g, 54 mmol) in anhydrous ether (100 mL) at 0°C, and the mixture was stirred at room temperature for 24 hours. The reaction mixture was concentrated in vacuo, diluted with water (100 mL), and extracted with ethyl acetate (100 mL×3). The combined organic layer was then washed with H2O (100 mL×3) and brine (100 mL×3), dried over anhydrous Na2SO4 and concentrated in vacuo. The crude product was purified on a silica gel column to obtain compound S4 in a yield of 38-41%.

[0051] Example 2: Synthesis route of 4-aminoisoxazole-5-carboxamide 1:

[0052] (A) Synthesis route of alkyl-substituted 4-aminoisoxazole-5-carboxamides 1a-1c, 1e:

[0053]

[0054] Compound S4 (11 mmol) was dissolved in ammonia methanol solution (50 mL), and the mixture was stirred at room temperature for 3 hours. After the solvent was removed under reduced pressure, compound S5 was obtained in 95-97% yield and used in the next step without further purification.

[0055] At 0°C, compound S5 (10 mmol) was dissolved in a solvent (50 mL, ethanol / water = 2 / 1), and NH4Cl (13.25 g, 250 mmol) and zinc powder (6.5 g, 100 mmol) were added. After the mixture was stirred at room temperature for 4 hours, it was filtered to remove the insoluble matter, and the filter cake was washed with ethanol. The filtrate was concentrated in vacuo and purified on a silica gel column to obtain compounds 4-aminoisoxazole-5-carboxamide 1a-1c, 1e with a yield of 76-80%.

[0056]

[0057] The above formula is 4-amino-3-cyclopentyloxazole-5-carboxamide 1a: white solid; yield 78%; mp: 148-150°C; 1 H NMR (400MHz, CDCl3) δ6.13(s,1H),5.63(s,1H),4.18(s,2H),3.14–3.03(m,1H),2.11(d,J=8.0Hz,2H),1.96–1.82(m,4H),1.79–1.70(m,2H). 13 C NMR (100MHz, DMSO-d6) δ160.5,160.2,140.2,132.3,35.0,30.1,24.9.

[0058]

[0059] The above formula is 4-amino-3-cyclohexyloxazole-5-carboxamide 1b: white solid; yield 80%; mp: 168-170°C; 1 H NMR (400MHz, CDCl3) δ6.92–5.73(m,2H),4.27(s,2H),2.61(t,J=11.6Hz,1H),2.00(d,J=12.6Hz,2H),1.90 (d,J=7.2Hz,2H),1.76(d,J=10.4Hz,1H),1.62(d,J=12.3Hz,1H),1.41–1.33(m,2H),0.94(t,J=7.4Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ160.7,160.5,140.0,131.9,33.8,32.4,30.1,25.7.

[0060]

[0061] The above formula is 4-amino-3-(tetrahydro-2H-pyran-4-yl)isoxazole-5-carboxamide 1c: white solid; yield 79%; mp: 187-189°C; 1 H NMR(300MHz,DMSO-d6)δ7.69(s,1H),7.45(s,1H),5.19(s,2H),4.02–3.81(m,2H),3.42(td,J=1 1.5, 2.1Hz, 2H), 3.02 (tt, J=11.3, 3.7Hz, 1H), 1.85 (dd, J=12.9, 1.7Hz, 2H), 1.74–1.55 (m, 2H). 13C NMR (100MHz, DMSO-d6) δ160.4,159.6,140.2,131.9,66.9,31.1,29.9.

[0062]

[0063] The above formula is 4-amino-3-isopropylisoxazole-5-carboxamide 1e: white solid; yield 79%; mp: 144-146°C; 1 H NMR (400MHz, DMSO-d6) δ7.73(s,1H),7.48(s,1H),5.16(s,2H),3.07(dt,J=13.5,6.7Hz,1H),1.25(d,J=6.8Hz,6H). 13 C NMR (100MHz, DMSO-d6) δ161.7,160.9,140.7,132.2,25.1,20.5.

[0064] (B) Synthesis route of 3-(1-acetylpiperidin-4-yl)-4-aminoisoxazole-5-carboxamide 1d:

[0065]

[0066] Trifluoroacetic acid (5 mL) was added to a dichloromethane solution (50 mL) of compound S6 (3.69 g, 10 mmol), and the mixture was stirred at room temperature for 2 hours. After the reaction of the raw materials was complete, the mixture was concentrated in vacuo to obtain compound S7 in the form of trifluoroacetate with a yield of 100%, which was used in the next step without further purification.

[0067] Et3N (1.2 g, 12 mmol) was added to a solution of compound S7 (3.83 g, 10 mmol) in N,N-dimethylformamide (50 mL), and acetyl chloride solution (936 mg, 12 mmol) was slowly added to the above mixture at 0°C. After that, the reaction mixture was stirred at room temperature for 6 hours. After the reaction of the raw materials was complete, the reaction mixture was diluted with water (150 mL) and extracted with ethyl acetate (100 mL×3). The combined organic layer was then washed with H2O (150 mL×3) and brine (150 mL×3), dried over anhydrous Na2SO4, and the crude product was purified on a silica gel column to obtain a light yellow solid compound S8 with a yield of 65%.

[0068] Compound S8 (6.5 mmol) was dissolved in ammonia methanol solution (30 mL), and the mixture was stirred at room temperature for 3 hours. After the solvent was removed under reduced pressure, compound S9 was obtained in 99% yield and used in the next step without further purification.

[0069] At 0°C, compound S9 (6.5 mmol) was dissolved in a solvent (30 mL, ethanol / water = 2 / 1), and NH4Cl (8.6 g, 162.5 mmol) and zinc powder (4.2 g, 65 mmol) were added. After the mixture was stirred at room temperature for 4 hours, it was filtered to remove the insoluble matter, and the filter cake was washed with ethanol. The filtrate was concentrated in vacuo and purified on a silica gel column to obtain compound 3-(1-acetylpiperidin-4-yl)-4-aminoisoxazole-5-carboxamide 1d with a yield of 78%.

[0070]

[0071] The above formula is 3-(1-acetylpiperidin-4-yl)-4-aminoisoxazole-5-carboxamide 1d: white solid; yield 78%; mp: 210-212°C; 1 H NMR (400MHz, DMSO-d6) δ7.73(s,1H),7.48(s,1H),5.23(s,2H),4.41(d,J=12.5Hz,1H),3.88(d,J=13.4Hz,1H),3.13(t,J=12.5Hz,1 H),3.02(t,J=11.1Hz,1H),2.66(t,J=12.2Hz,1H),2.02(s,3H),1.93(t,J=12.4Hz,2H),1.67–1.53(m,1H),1.43(q,J=10.9Hz,1H). 13 C NMR (100MHz, DMSO-d6) δ168.1,160.4,159.4,140.2,131.9,45.8,40.8,32.0,29.6,29.0,21.4.

[0072] Example 3: Synthesis of isoxazolo[4,5-d]pyrimidin-7(6H)-one 3a-3ad:

[0073]

[0074] Compound 1 (1.0 mmol), compound 2 (1.2 mmol), I2 (254 mg, 1.0 mmol) and dimethyl sulfoxide (234 mg, 3.0 mmol) were dissolved in CH3CN (5 mL) and reacted at 80°C overnight. TLC monitored the completion of the reaction. After the reaction was completed, the mixture was diluted with an aqueous solution of sodium thiosulfate (20 mL) and extracted with ethyl acetate (20 mL×3). The organic layer was dried over anhydrous Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography to obtain isoxazolo[4,5-d]pyrimidin-7(6H)-one 3.

[0075]

[0076] The above formula is 3-cyclopentyl-5-phenylisoxazolo[4,5-d]pyrimidin-7(6H)-one 3a: white solid; yield 93%; mp: 209-211°C; 1 H NMR (400MHz, CDCl3): δ11.60(s,1H),8.18(d,J=4.2Hz,2H),7.64(s,3H),3.58(p,J=8.4Hz ,1H),2.27(d,J=7.3Hz,2H),2.17–2.08(m,2H),1.97(d,J=5.3Hz,2H),1.87–1.78(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.3,155.2,152.4,149.4,140.5,132.5,131.9,129.1,128.4,36.0,31.2,25.6.

[0077]

[0078] The above formula is 3-cyclopentyl-5-(4-methoxyphenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3b: pale yellow solid; yield 97%; mp: 202-204°C; 1 H NMR (400MHz, CDCl3) δ11.66(s,1H),8.17(d,J=8.7Hz,2H),7.12(d,J=8.7Hz,2H),3.95(s,3H),3.57(p,J=8.2H z,1H),2.31–2.23(m,2H),2.12(dd,J=12.0,7.1Hz,2H),1.99(t,J=10.0Hz,2H),1.83(dd,J=14.3,9.4Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ163.3,162.4,154.9,152.5,149.0,140.6,130.1,124.6,114.5,56.0,36.0,31.2,25.7.

[0079]

[0080] The above formula is 3-cyclopentyl-5-(p-tolyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3c: white solid; yield 94%; mp: 204-206°C; 1H NMR (400MHz, CDCl3) δ11.91(s,1H),8.09(d,J=7.9Hz,2H),7.42(d,J=7.8Hz,2H),3.57(p,J=8.0 Hz,1H),2.49(s,3H),2.26(d,J=6.6Hz,2H),2.17–2.07(m,2H),1.98(m,2H),1.88–1.77(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.3,155.2,152.4,149.3,142.0,140.5,129.7,128.3,36.0,31.2,25.6,21.5.

[0081]

[0082] The above formula is 3-cyclopentyl-5-(4-fluorophenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3d: white solid; yield 84%; mp: 209-211°C; 1 H NMR (400MHz, CDCl3) δ12.23(s,1H),8.27–8.21(m,2H),7.30(d,J=8.3Hz,2H),3.54(p,J=8.4Hz,1H ),2.23(d,J=6.0Hz,2H),2.13–2.04(m,2H),1.96(d,J=14.0Hz,2H),1.80(dd,J=13.8,7.9Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ165.8,163.3,154.2,152.3,149.3,140.4,131.0,131.0,129.0,116.3,116.1,36.0,31.1,25.6.

[0083]

[0084] The above formula is 3-cyclopentyl-5-(4-(trifluoromethyl)phenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3e: light brown solid; yield 61%; mp: 203-205°C; 1HNMR (400MHz, DMSO-d6) δ13.40(s,1H),8.25(d,J=8.1Hz,2H),7.93(d,J=8.2Hz,2H),3.47(p,J=8.1Hz, 1H), 2.14(td,J=11.6,7.5Hz,2H),2.02–1.91(m,2H),1.89–1.77(m,2H),1.72(dd,J=14.3,7.4Hz,2H). 13 CNMR(100MHz,DMSO-d6)δ163.4,154.0,152.3,149.7,140.3,136.3,132.2,131.8, 131.5,131.2,129.4,128.4,126.1,126.0,125.7,123.0,120.3,36.0,31.1,25.6.

[0085]

[0086] The above formula is 4-(3-cyclopentyl-7-oxo-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)benzonitrile 3f: pale yellow solid; yield 91%; mp: 224-226°C; 1 HNMR (400MHz, CDCl3) δ12.78(s,1H),8.42(d,J=8.2Hz,2H),7.94(d,J=8.3Hz,2H),3.59(p,J=8.2Hz,1H), 2.28(d,J=7.1Hz,2H), 2.12(dd,J=16.7,9.6Hz,2H), 2.00(d,J=14.2Hz,2H), 1.85(dd,J=14.1,8.4Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ163.4,153.8,152.4,149.8,140.3,136.7,133.1,129.2,118.7,114.2,36.0,31.2,25.6.

[0087]

[0088] The above formula is 4-(3-cyclopentyl-7-oxo-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 3g: light yellow solid; yield 94%; mp: 214-216°C; 1HNMR (400MHz, CDCl3) δ12.19 (s, 1H), 8.31 (d, J = 8.7Hz, 4H), 4.02 (s, 3H), 3.59 (p, J = 8.3Hz ,1H),2.28(d,J=7.1Hz,2H),2.18–2.08(m,2H),1.97(d,J=6.2Hz,2H),1.87–1.78(m,2H). 13 C NMR (100MHz, DMSO-d6) δ166.1,163.5,154.4,152.4,149.7,140.4,136.7,132.3,129.8,128.9,53.0,36.0,31.2,25.6.

[0089]

[0090] The above formula is 3-cyclopentyl-5-(4-nitrophenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3h: yellow solid; yield 96%; mp: 216-218°C; 1 H NMR (400MHz, DMSO-d6) δ13.52(s,1H),8.41(d,J=8.6Hz,2H),8.32(d,J=8.7Hz,2H),3.51(p,J=8. 0Hz,1H),2.21–2.13(m,2H),1.98(dd,J=12.5,7.0Hz,2H),1.89–1.80(m,2H),1.79–1.70(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.4,153.5,152.3,149.8,149.4,140.2,138.2,129.9,124.2,36.0,31.2,25.6.

[0091]

[0092] The above formula is 3-cyclopentyl-5-(4-(hydroxymethyl)phenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3i: white solid; yield 95%; mp: 182-184°C; 1H NMR (400MHz, DMSO-d6) δ13.15(s,1H),8.06(d,J=8.1Hz,2H),7.50(d,J=8.0Hz,2H),5.38(t,J=5.6Hz,1H),4.61(d,J=5. 3Hz,2H),3.49(p,J=8.1Hz,1H),2.20–2.11(m,2H),2.02–1.94(m,2H),1.89–1.81(m,2H),1.75(dd,J=14.4,7.7Hz,2H). 13 CNMR(100MHz,DMSO-d6)δ163.3,155.2,152.4,149.3,146.8,140.5,130.8,128.3,126.8,62.9,36.0,31.2,25.6.

[0093]

[0094] The above formula is 3-cyclopentyl-5-(2-hydroxyphenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3j: white solid; yield 94%; mp: 213-215°C; 1 H NMR (400MHz, DMSO-d6) δ8.04(d,J=7.9Hz,1H),7.46(t,J=7.6Hz,1H),7.02(dd,J=19.3,8.0Hz,2H),3.51(d d,J=16.5,8.3Hz,1H),2.16(d,J=7.4Hz,2H),2.01–1.92(m,2H),1.82(d,J=5.8Hz,2H),1.78–1.69(m,2H). 13 C NMR (100MHz, DMSO-d6) δ162.8,157.9,155.2,151.6,149.2,139.1,133.8,129.5,119.9,117.8,116.5,35.9,31.0,25.7.

[0095]

[0096] The above formula is 3-cyclopentyl-5-(3-hydroxyphenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3k: white solid; yield 97%; mp: 219-221°C; 1H NMR (400MHz, DMSO-d6) δ13.08(s,1H),9.84(s,1H),7.51(d,J=7.5Hz,2H),7.35(t,J=7.9Hz,1H),6.99(d,J=8.7Hz,1H) ,3.48(p,J=8.1Hz,1H),2.16(d,J=7.5Hz,2H),2.03–1.94(m,2H),1.86(t,J=9.9Hz,2H),1.75(dd,J=13.3,8.5Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ163.3,158.0,155.3,152.4,149.4,140.5,133.8,130.2,119.1,115.2,36.0,31.2,25.6.

[0097]

[0098] The above formula is 3-cyclopentyl-5-(4-hydroxyphenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3l: white solid; yield 92%; mp: 214-216°C; 1 H NMR (400MHz, DMSO-d6) δ12.91(s,1H),10.21(s,1H),7.99(d,J=8.5Hz,2H),6.91(d,J=8.6Hz,2H),3.47 (p,J=8.1Hz,1H),2.15(d,J=7.3Hz,2H),2.01–1.93(m,2H),1.84(s,2H),1.74(dd,J=14.1,7.7Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ163.2,161.1,155.2,152.5,148.8,140.7,130.2,123.1,115.9,36.0,31.1,25.6.

[0099]

[0100] The above formula is 3-cyclopentyl-5-(2-ethoxyphenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3m: pale yellow solid; yield 90%; mp: 210-212°C; 1H NMR (400MHz, CDCl3) δ11.69(s,1H),8.52(d,J=7.8Hz,1H),7.55(t,J=7.8Hz,1H),7.20(t,J=7.6Hz,1H),7.11(d,J=8.3Hz,1H),4.38(q,J=6 .8Hz,2H),3.56(p,J=8.0Hz,1H),2.26(d,J=5.8Hz,2H),2.17–2.09(m,2H),1.97(d,J=6.3Hz,2H),1.86–1.77(m,2H),1.66(t,J=6.9Hz,3H). 13 C NMR (100MHz, DMSO-d6) δ163.3,156.9,154.7,151.5,149.2,140.6,133.0,131.1,122.3,121.0,113.3,64.6,36.0,31.1,25.5,14.9.

[0101]

[0102] The above formula is 3-(3-cyclopentyl-7-oxo-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 3n: light yellow solid; yield 89%; mp: 209-211°C; 1 HNMR (400MHz, CDCl3) δ12.19 (s, 1H), 8.31 (d, J = 8.7Hz, 4H), 4.02 (s, 3H), 3.59 (p, J = 8.3Hz ,1H),2.28(d,J=7.1Hz,2H),2.18–2.08(m,2H),1.97(d,J=6.2Hz,2H),1.87–1.78(m,2H). 13 C NMR (100MHz, DMSO-d6) δ166.1,163.5,154.4,152.4,149.7,140.4,136.7,132.3,129.8,128.9,53.0,35.95,31.2,25.6.

[0103]

[0104] The above formula is 3-cyclopentyl-5-(3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3o: white solid; yield 95%; mp: 191-193°C; 1H NMR (400MHz, DMSO-d6) δ13.29(s,1H),8.37(s,1H),8.17(d,J=7.4Hz,1H),7.87(d,J=7.2Hz,1H),7.59(t,J= 7.5Hz,1H),3.55–3.47(m,1H),2.15(s,2H),1.99(d,J=10.0Hz,2H),1.84(s,2H),1.74(s,2H),1.35(s,12H) 13 CNMR(100MHz,DMSO-d6)δ163.5,155.5,152.5,149.5,140.5,137.7,134.6,132.4,131.5,128.8,84.5,35.8,31.3,25.6,25.2.

[0105]

[0106] The above formula is 3-cyclopentyl-5-(pyridin-4-yl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3p: pale yellow solid; yield 48%; mp: 204-206°C; 1 H NMR (400MHz, CDCl3) δ12.63(s,1H),8.95(d,J=3.8Hz,2H),8.17(d,J=4.2Hz,2H),3.70–3.53 (m,1H),2.29(d,J=6.2Hz,2H),2.12(dd,J=12.4,7.4Hz,2H),2.00(s,2H),1.89–1.81(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.4,153.2,152.3,150.8,150.1,140.2,139.6,122.2,35.9,31.2,25.6.

[0107]

[0108] The above formula is 5-(6-aminopyridin-3-yl)-3-cyclopentylisoxazolo[4,5-d]pyrimidin-7(6H)-one 3q: pale yellow solid; yield 28%; mp: 198-200°C; 1H NMR (400MHz, DMSO-d6) δ12.85(s,1H),8.67(s,1H),8.05(d,J=8.4Hz,1H),6.69(s,2H),6.51(d,J=8.9Hz,1H ),3.48–3.41(m,1H),2.12(d,J=6.8Hz,2H),1.98–1.92(m,2H),1.82(d,J=2.5Hz,2H),1.70(d,J=4.6Hz,2H). 13 CNMR(100MHz,DMSO-d6)δ165.1,164.2,162.7,158.7,153.2,148.8,135.8,134.7,120.8,109.0,36.0,31.1,25.6.

[0109]

[0110] The above formula is 5-(6-aminopyridin-3-yl)-3-cyclopentylisoxazolo[4,5-d]pyrimidin-7(6H)-one 3r: yellow solid; yield 59%; mp: 312-314°C; 1 H NMR (400MHz, DMSO-d6) δ13.05(s,1H),8.89(s,2H),7.43(s,2H),3.47(p,J=8.1Hz,1H),2.1 5(d,J=7.7Hz,2H),2.01–1.92(m,2H),1.83(d,J=5.0Hz,2H),1.74(dd,J=14.3,7.7Hz,2H). 13 C NMR (100MHz, DMSO-d6) δ164.6,163.2,158.4,152.7,152.3,148.9,140.6,115.2,35.9,31.1,25.7.

[0111]

[0112] The above formula is 3-cyclopentyl-5-(quinolin-7-yl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3s: brown solid; yield 98%; mp: 201-203°C; 1H NMR (400MHz, DMSO-d6) δ13.51(s,1H),9.49(d,J=1.9Hz,1H),9.07(d,J=2.0Hz,1H),8.13(dd,J=7.9,4.8Hz,2H),7.94–7.89(m,1H),7.74( t,J=7.5Hz,1H),3.53(p,J=8.1Hz,1H),2.18(dt,J=11.7,5.8Hz,2H),2.00(dd,J=13.7,6.2Hz,2H),1.90–1.81(m,2H),1.79–1.70(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.5,153.6,152.4,149.7,149.5,148.7,140.4,136.7,132.1,129.7,129.3,128.2,126.8,125.6,36.0,31.3,25.7.

[0113]

[0114] The above formula is 5-(3-cyclopentyl-7-oxo-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)-1H-indole-1-carboxylic acid tert-butyl ester 3t: pale yellow solid; yield 37%; mp: 184-186°C; 1 H NMR (400MHz, CDCl3) δ10.85(s,1H),8.32(s,2H),8.01(d,J=8.7Hz,1H),7.71(s,1H),6.74(s,1H),3.56(p,J=8. 8Hz,1H),2.25(dd,J=7.8,2.3Hz,2H),2.15–2.07(m,2H),1.96(d,J=5.2Hz,2H),1.83–1.77(m,2H),1.71(s,9H). 13 CNMR (100MHz, CDCl3) δ163.5,154.6,153.3,149.4,148.4,142.1,137.2,131 .0,127.5,126.1,123.5,120.7,115.8,107.9,84.5,36.2,31.3,28.2,25.7.

[0115]

[0116] The above formula is 3-cyclopentyl-5-(1H-indazol-4-yl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3u: pale yellow solid; yield 82%; mp: 229-231°C;1 H NMR (400MHz, DMSO-d6) δ13.44(s,1H),13.26(s,1H),8.57(s,1H),7.83(dd,J=15.4,7.7Hz,2H),7.52(t,J=7.7Hz ,1H),3.63–3.51(m,1H),2.20(d,J=5.6Hz,2H),2.10–2.02(m,2H),1.85(d,J=5.1Hz,2H),1.77(d,J=5.8Hz,2H). 13 CNMR(100MHz,DMSO-d6)δ163.4,154.7,152.4,149.6,140.9,140.4,134.5,126.0,125.1,121.9,120.8,114.4,36.0,31.1,25.8.

[0117]

[0118] The above formula is 3-cyclopentyl-5-(1-methyl-1H-pyrazol-4-yl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3v: pale yellow solid; yield 90%; mp: 205-207°C; 1 HNMR(400MHz,DMSO-d6)δ12.96(s,1H),8.48(s,1H),8.19(s,1H),3.93(s,3H),3.43(p,J=8.1 Hz,1H),2.14(d,J=7.8Hz,2H),2.00–1.90(m,2H),1.85(t,J=10.0Hz,2H),1.77–1.67(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.0,152.3,150.4,148.6,140.8,138.9,132.5,116.3,39.5,36.0,31.1,25.6.

[0119]

[0120] The above formula is 3-cyclopentyl-5-(furan-2-yl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3w: white solid; yield 60%; mp: 190-192°C; 1H NMR (400MHz, CDCl3) δ11.17(s,1H),7.68(s,1H),7.47(s,1H),6.66(s,1H),3.51(p,J=8.1H z,1H),2.21(d,J=6.7Hz,2H),2.05(dd,J=13.5,6.4Hz,2H),1.92(s,2H),1.82–1.73(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.3,152.0,149.2,147.2,146.6,145.9,140.3,115.2,113.1,35.9,31.2,25.5.

[0121]

[0122] The above formula is 5-cyclohexyl-3-cyclopentylisoxazolo[4,5-d]pyrimidin-7(6H)-one 3x: white solid; yield 94%; mp: 163-165°C; 1 H NMR (400MHz, CDCl3) δ11.73(s,1H),3.48(dd,J=16.4,8.3Hz,1H),2.75(t,J=11.7Hz,1H),2.17(d,J=6.7Hz,2H),2.01(d,J=11.2 Hz,4H),1.90(d,J=9.2Hz,4H),1.77(d,J=12.4Hz,3H),1.66(d,J=8.7Hz,2H),1.45(dd,J=25.0,12.4Hz,2H),1.38–1.28(m,1H). 13 C NMR (100MHz, DMSO-d6) δ163.5,163.0,152.2,149.2,140.3,42.6,35.9,31.1,30.9,25.8,25.7,25.6.

[0123]

[0124] The above formula is 3-cyclopentyl-5-(tetrahydro-2H-pyran-4-yl)isoxazolo[4,5-d]pyrimidin-7(6H)-one 3y: white solid; yield 86%; mp: 170-172°C; 1H NMR (400MHz, CDCl3) δ12.49(s,1H),4.15(d,J=11.2Hz,2H),3.65(t,J=11.4Hz,2H),3.57–3.46(m, 1H),3.08(t,J=10.7Hz,1H),2.20(s,2H),2.05(dd,J=20.3,11.5Hz,6H),1.95(s,2H),1.79(s,2H). 13 C NMR (100MHz, CDCl3) δ163.4,160.6,153.8,148.3,141.9,67.2,40.2,36.1,31.3,30.5,25.7.

[0125]

[0126] The above formula is 3-cyclopentyl-5-isopropylisoxazolo[4,5-d]pyrimidin-7(6H)-one 3z: white solid; yield 86%; mp: 157-159°C; 1 H NMR (400MHz, CDCl3) δ11.76(s,1H),3.47(p,J=8.1Hz,1H),3.08(dt,J=13.8,6.9Hz,1H),2.17(dd ,J=6.3,4.8Hz,2H),2.05–1.98(m,2H),1.95–1.87(m,2H),1.80–1.72(m,2H),1.41–1.36(m,6H). 13 C NMR (100MHz, DMSO-d6) δ164.4,163.1,152.2,149.2,140.3,35.9,33.3,31.1,25.6,21.0.

[0127]

[0128] The above formula is 3-(3-cyclohexyl-7-oxo-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 3aa: light yellow solid; yield 95%; mp: 228-230°C; 1 HNMR (400MHz, CDCl3) δ12.34(s,1H),8.88(s,1H),8.46(d,J=7.9Hz,1H),8.29(d,J=7.8Hz,1H),7.70(t,J=7.8Hz ,1H),4.05(s,3H),3.27–3.17(m,1H),2.18(d,J=13.0Hz,2H),1.95–1.81(m,5H),1.66(s,1H),1.51–1.37(m,2H).13 C NMR (100MHz, DMSO-d6) δ165.6,163.1,154.1,151.9,149.0,139.8,132.8,132.7,131.8,130.1,129.3,128.8,52.5,34.7,30.1,25.5,25.3.

[0129]

[0130] The above formula is 3-(7-oxo-3-(tetrahydro-2H-pyran-4-yl)-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 3ab: pale yellow solid; yield 90%; mp: 224-226°C; 1 H NMR (400MHz, DMSO-d6) δ13.43(s,1H),8.63(s,1H),8.32(d,J=7.6Hz,1H),8.16(d,J=7.7Hz,1H),7.72(t,J=7.8 Hz,1H),3.97(d,J=11.0Hz,2H),3.92(s,3H),3.56(t,J=10.9Hz,2H),3.43(t,J=10.2Hz,1H),2.05–1.94(m,4H). 13 CNMR(100MHz,DMSO-d6)δ166.1,162.5,154.7,152.3,149.7,140.2,133.2,132.3,130.6,129.8,129.3,66.9,53.0,32.5,30.3.

[0131]

[0132] The above formula is 3-(3-(1-acetylpiperidin-4-yl)-7-oxo-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 3ac: pale yellow solid; yield 69%; mp: 189-191°C; 1H NMR(400MHz,DMSO-d6)δ13.44(s,1H),8.65(s,1H),8.32(d,J=7.3Hz,1H),8 .16(d,J=7.4Hz,1H),7.72(t,J=7.8Hz,1H),4.40(d,J=12.7Hz,1H),3.92(s, 4H),3.51–3.40(m,1H),3.28(d,J=12.3Hz,1H),2.86(t,J=11.9Hz,1H),2.1 3(t,J=14.9Hz,2H),2.05(s,3H),1.95–1.85(m,1H),1.75(t,J=13.7Hz,1H). 13 CNMR(100MHz,DMSO-d6)δ168.6,166.1,162.4,154.7,152.3,149.7,140.2,1 33.1,132.3,130.6,129.8,129.3,53.0,45.8,40.9,33.3,30.1,29.5,21.8.

[0133]

[0134] The above formula is 3-(3-isopropyl-7-oxo-6,7-dihydroisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 3ad: white solid; yield 99%; mp: 180-182°C; 1 HNMR (400MHz, CDCl3) δ12.33(s,1H),8.89(s,1H),8.47(d,J=7.5Hz,1H),8.29(d,J= 7.4Hz,1H),7.70(t,J=7.7Hz,1H),4.05(s,3H),3.50(s,1H),1.56(d,J=6.8Hz,6H). 13 CNMR(100MHz,DMSO-d6)δ166.1,164.4,154.4,152.3,149.6,140.2,133.1,133.0,132.3,130.5,129.7,129.2,52.9,26.3,20.7.

[0135] Example 4: Synthesis route of morpholine ring substituted isoxazolopyrimidine compounds 4a-4u:

[0136]

[0137] POCl3 (383 mg, 5.0 mmol) was added to a solution of compound 3 (0.5 mmol) in CH3CN (10 mL), and the temperature was raised to 80°C for 5 hours. After the reaction of the raw materials was complete, the reaction solution was cooled to room temperature, and the mixture was concentrated under reduced pressure. The residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 10 / 1, v / v) to obtain compound S10 with a yield of 75-90%.

[0138] Morpholine (47 mg, 0.54 mmol) and Et3N (68 mg, 0.68 mmol) were added to a stirred solution of compound S10 (0.45 mmol) in dichloromethane (10 mL) and reacted at room temperature for 1 hour. After the reaction of the raw materials was complete, the reaction mixture was diluted with water (10 mL), extracted with dichloromethane (10 mL × 3), dried over anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 20 / 1, v / v) to obtain compound 4 in a yield of 67-87%.

[0139]

[0140] The above formula is 3-cyclopentyl-7-morpholine-5-phenylisoxazolo[4,5-d]pyrimidine 4a: white solid; yield 83%; mp: 224-226°C; 1 H NMR (400MHz, CDCl3) δ8.42(d,J=5.1Hz,2H),7.45(s,3H),4.16(s,4H),3.89(d,J=4.0Hz,4H),3.58(p,J =8.0Hz,1H),2.23(d,J=6.9Hz,2H),2.13(td,J=14.7,7.9Hz,2H),2.02–1.90(m,2H),1.85–1.72(m,2H). 13 C NMR (100MHz, CDCl3) δ163.0,160.2,148.2,145.9,143.7,138.0,130.0,128.3,128.2,66.9,45.6,36.2,31.2,25.8.

[0141]

[0142] The above formula is 3-cyclopentyl-5-(4-methoxyphenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidine 4b: white solid; yield 85%; mp: 121-123°C; 1H NMR(400MHz, CDCl3)δ8.40(d,J=8.8Hz,2H),7.01(d,J=8.8Hz,2H),4.20–4.16(m,4H),3.93–3.90(m,7H),3 .60(p,J=8.2Hz,1H),2.31–2.22(m,2H),2.20–2.12(m,2H),2.00(dt,J=9.0,5.7Hz,2H),1.88–1.78(m,2H). 13 C NMR (100MHz, CDCl3) δ162.9,161.3,160.1,148.1,145.8,143.4,130.8,129.7,113.6,66.9,55.4,45.7,36.2,31.2,25.8.

[0143]

[0144] The above formula is 3-cyclopentyl-5-(4-methoxyphenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidine 4c: white solid; yield 86%; mp: 131-133°C; 1 H NMR (400MHz, CDCl3) δ8.30(d,J=8.2Hz,2H),7.24(s,2H),4.17–4.13(m,4H),3.90–3.86(m,4H),3.57(p,J=8. 2Hz,1H),2.41(s,3H),2.27–2.19(m,2H),2.17–2.09(m,2H),2.00–1.91(m,2H),1.79(tt,J=7.9,3.8Hz,2H). 13 C NMR (100MHz, CDCl3) δ162.9,160.3,148.1,145.9,143.6,140.1,135.4,129.1,128.1,66.9,45.6,36.2,31.2,25.8,21.5.

[0145]

[0146] The above formula is 3-cyclopentyl-5-(4-fluorophenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidine 4d: white solid; yield 63%; mp: 125-127°C; 1H NMR (400MHz, CDCl3) δ8.44–8.39(m,2H),7.13(t,J=8.7Hz,2H),4.17–4.13(m,4H),3.91–3.87(m,4H),3.57(p,J=8 .2Hz,1H),2.22(dt,J=11.7,5.9Hz,2H),2.17–2.07(m,2H),2.01–1.92(m,2H),1.80(ddd,J=10.5,8.0,3.2Hz,2H). 13 C NMR (100MHz, CDCl3) δ165.5,163.0,162.9,159.3,148.2,145.8,143.6,134.2,130.2,130.1,115.3,115.1,66.8,45.5,36.2,31.2,25.8.

[0147]

[0148] The above formula is 3-cyclopentyl-7-morpholine-5-(4-(trifluoromethyl)phenyl)isoxazolo[4,5-d]pyrimidine 4e: white solid; yield 67%; mp: 151-153°C; 1 H NMR (400MHz, CDCl3) δ8.53(d,J=8.1Hz,2H),7.71(d,J=8.3Hz,2H),4.19–4.15(m,4H),3.92–3.89(m,4H),3. 59(p,J=8.2Hz,1H),2.31–2.21(m,2H),2.18–2.09(m,2H),2.01–1.93(m,2H),1.82(qt,J=10.2,5.2Hz,2H). 13 C NMR (100MHz, CDCl3) δ163.0,158.8,148.2,145.8,143.8,141.3,132.1,131.7,131 .4,131.1,128.4,125.6,125.3,125.2,122.9,120.2,66.8,45.7,36.2,31.2,25.8.

[0149]

[0150] The above formula is 4-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)benzonitrile 4f: white solid; yield 82%; mp: 209-211°C; 1H NMR (400MHz, CDCl3) δ8.53(d,J=8.3Hz,2H),7.75(d,J=8.4Hz,2H),4.19–4.15(m,4H),3.91–3.88(m,4H),3.57(p ,J=8.3Hz,1H),2.23(dt,J=11.7,6.0Hz,2H),2.16–2.07(m,2H),2.00–1.92(m,2H),1.81(tt,J=8.1,4.0Hz,2H). 13 CNMR (100MHz, CDCl3) δ163.0,158.3,148.2,145.7,143.9,142.1,132.2,128.7,119.0,113.2,66.8,45.9,36.18,31.2,25.8.

[0151]

[0152] The above formula is 4-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 4g: white solid; yield 81%; mp: 219-221°C; 1 H NMR(400MHz, CDCl3)δ8.48(d,J=8.5Hz,2H),8.12(d,J=8.5Hz,2H),4.19–4.14(m,4H),3.96(s,3H),3.92–3.8 8(m,4H),3.59(p,J=8.2Hz,1H),2.29–2.21(m,2H),2.18–2.09(m,2H),2.01–1.92(m,2H),1.86–1.75(m,2H). 13 C NMR (100MHz, CDCl3) δ167.1,163.0,159.2,148.2,145.8,143.8,142.2,131.2,129.6,128.1,66.9,52.3,45.6,36.2,31.2,25.8.

[0153]

[0154] The above formula is 2-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenol 4h: white solid; yield 82%; mp: 179-181°C; 1H NMR (400MHz, CDCl3) δ13.38(s,1H),8.36(d,J=7.8Hz,1H),7.36(t,J=7.5Hz,1H),7.02(d,J=8.0Hz,1H),6.93(t,J=7.7Hz,1H),4.19(s,4H ),3.90(s,4H),3.59–3.48(m,1H),2.24(dd,J=14.6,6.8Hz,2H),2.07(dd,J=12.4,7.3Hz,2H),1.93(s,2H),1.81(dd,J=12.1,6.5Hz,2H). 13 C NMR (100MHz, CDCl3) δ161.7,161.0,160.2,147.9,143.5,142.2,132.4,129.4,119.0,118.8,117.7,66.8,45.4,36.2,30.9,25.8.

[0155]

[0156] The above formula is 3-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenol 4i: white solid; yield 81%; mp: 221-223°C; 1 H NMR (400MHz, CDCl3) δ8.00(d,J=7.5Hz,1H),7.91(s,1H),7.33(t,J=7.8Hz,1H),6.93(d,J=7.8Hz,1H),5.12( s,1H),4.15(s,4H),3.88(s,4H),3.57(t,J=8.1Hz,1H),2.22(s,2H),2.12(s,2H),1.95(s,2H),1.78(s,2H). 13 C NMR (100MHz, CDCl3) δ163.0,160.2,158.4,148.7,145.8,144.3,139.8,130.3,119.7,118.2,115.6,66.9,46.2,36.5,31.6,26.2.

[0157]

[0158] The above formula is 4-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenol 4j: white solid; yield 82%; mp: 220–222°C; 1H NMR (400MHz, CDCl3) δ7.99(d,J=7.6Hz,1H),7.91(s,1H),7.33(t,J=7.8Hz,1H),6.92(d,J=7.5Hz,1H),5.35(s,1H) ,4.15(s,4H),3.88(s,4H),3.62–3.52(m,1H),2.22(d,J=6.5Hz,2H),2.16–2.08(m,2H),1.94(s,2H),1.77(s,2H). 13 C NMR (100MHz, CDCl3) δ162.7,160.0,158.1,148.5,145.5,144.0,139.5,130.0,119.5,117.9,115.4,66.7,45.9,36.3,31.3,26.0.

[0159]

[0160] The above formula is 3-cyclopentyl-5-(2-ethoxyphenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidine 4k: yellow solid; yield 85%; mp: 80-82°C; 1 H NMR(400MHz, CDCl3)δ7.76(dd,J=7.6,1.7Hz,1H),7.39–7.33(m,1H),7.02(dd,J=13.0,7.9Hz,2H),4.15–4.11(m,4H),4.10–4.06(m,2H),3.86– 3.83(m,4H),3.57(p,J=8.1Hz,1H),2.24–2.17(m,2H),2.15–2.07(m,2H ),1.93–1.84(m,2H),1.75(tt,J=8.0,3.8Hz,2H),1.35(d,J=7.0Hz,3H). 13 C NMR (100MHz, CDCl3) δ162.8,161.2,157.3,147.9,145.6,143.1,131.9,130.4,129.1,120.5,113.4,66.9,64.3,45.7,36.3,31.2,25.5,15.0.

[0161]

[0162] The above formula is 3-cyclopentyl-7-morpholine-5-(3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)isoxazolo[4,5-d]pyrimidine 4l: white solid; yield 83%; mp: 129-131°C;1 H NMR (400MHz, CDCl3) δ8.84(s,1H),8.55(d,J=7.4Hz,1H),7.93(d,J=7.1Hz,1H),7.51(t,J=7.4Hz,1H),4.21 (s,4H),3.93(s,4H),3.71–3.59(m,1H),2.28(s,2H),2.17(s,2H),2.01(s,2H),1.83(s,2H),1.42(s,12H). 13 C NMR (100MHz, CDCl3) δ163.2,160.4,148.2,145.9,143.6,137.4,136.5,134.3,131.2,127.8,83.9,66.9,45.7,36.1,31.4,25.8,25.0.

[0163]

[0164] The above formula is 3-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)benzoic acid methyl ester 4m: white solid; yield 84%; mp: 138-140°C; 1 H NMR (400MHz, CDCl3) δ9.09(s,1H),8.66(d,J=7.6Hz,1H),8.15(d,J=7.6Hz,1H),7.57(t,J=7.7Hz,1H),4.21(s,4H),4.01(s, 3H),3.93(s,4H),3.63(p,J=8.1Hz,1H),2.28(d,J=6.8Hz,2H),2.20–2.12(m,2H),2.00(s,2H),1.85(dd,J=14.2,8.5Hz,2H). 13 C NMR (100MHz, CDCl3) δ167.2,163.0,159.3,148.2,145.8,143.8,138.5,132.6,130.9,130.4,129.4,128.4,66.9,52.3,45.7,36.1,31.25,25.8.

[0165]

[0166] The above formula is 3-cyclopentyl-7-morpholin-5-(pyridin-4-yl)isoxazolo[4,5-d]pyrimidine 4n: white solid; yield 82%; mp: 135-137°C; 1H NMR (400MHz, CDCl3) δ8.73(d,J=5.1Hz,2H),8.26(d,J=5.8Hz,2H),4.20–4.15(m,4H),3.92–3.88(m,4H),3.58(t ,J=8.2Hz,1H),2.24(dd,J=12.5,4.6Hz,2H),2.17–2.07(m,2H),1.97(dd,J=15.4,7.0Hz,2H),1.85–1.77(m,2H). 13 CNMR (100MHz, CDCl3) δ163.0,158.0,150.2,148.2,145.7,145.2,144.1,122.1,66.8,45.6,36.2,31.2,25.8.

[0167]

[0168] The above formula is 5-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)pyrimidin-2-amine 4o: white solid; yield 82%; mp: 227-229°C; 1 H NMR (400MHz, CDCl3) δ9.24(s,2H),5.33(s,2H),4.13(s,4H),3.88(s,4H),3.60–3.49(m ,1H),2.25–2.17(m,2H),2.11(dd,J=13.4,5.7Hz,2H),1.94(s,2H),1.82–1.74(m,2H). 13 C NMR (100MHz, CDCl3) δ163.4,162.8,158.6,157.2,148.2,145.4,143.6,122.1,66.8,45.2,36.1,31.2,25.8.

[0169]

[0170] The above formula is 3-cyclopentyl-7-morpholin-5-(quinolin-7-yl)isoxazolo[4,5-d]pyrimidine 4p: white solid; yield 81%; mp: 170-172°C; 1H NMR (400MHz, CDCl3) δ9.94(s,1H),9.10(s,1H),8.16(d,J=8.4Hz,1H),7.98(d,J=7.8Hz,1H),7.77(t,J=7.2Hz,1H),7.60(t,J=7 .1Hz,1H),4.22(s,4H),3.92(s,4H),3.68–3.56(m,1H),2.26(d,J=2.6Hz,2H),2.18(d,J=7.2Hz,2H),2.00(s,2H),1.82(s,2H). 13 C NMR (100MHz, CDCl3) δ162.9,158.2,150.7,148.6,148.1,145.6,143.7,135 .2,130.5,130.1,129.2,128.7,127.5,126.7,66.8,45.9,36.1,31.2,25.8.

[0171]

[0172] The above formula is 3-cyclopentyl-5-(1H-indazol-5-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidine 4q: white solid; yield 82%; mp: 238-240°C; 1 H NMR (400MHz, CDCl3) δ10.26(s,1H),9.05(s,1H),8.30(d,J=7.2Hz,1H),7.65(d,J=8.0Hz,1H),7.54(t,J=7.7Hz,1H ),4.24(s,4H),3.95(s,4H),3.72–3.61(m,1H),2.28(dd,J=19.6,9.4Hz,4H),2.02(s,2H),1.88(d,J=18.4Hz,2H). 13 C NMR (100MHz, CDCl3) δ162.9,160.3,148.2,145.7,143.6,141.0,136.8,131.5,126.6,122.5,121.6,111.7,66.9,45.7,36.4,31.1,25.7.

[0173]

[0174] The above formula is 3-cyclopentyl-5-(1-methyl-1H-pyrazol-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidine 4r: white solid; yield 83%; mp: 134-136°C; 1H NMR (400MHz, CDCl3) δ8.12(s,1H),8.04(s,1H),4.13(s,4H),3.99(s,3H),3.89(s,4H),3.56 (p,J=7.9Hz,1H),2.23(d,J=6.9Hz,2H),2.14–2.06(m,2H),1.95(s,2H),1.84–1.76(m,2H). 13 C NMR (100MHz, CDCl3) δ162.6,156.8,148.2,145.5,143.1,139.6,130.8,123.6,66.8,45.5,39.2,36.1,31.2,25.7.

[0175]

[0176] The above formula is 3-cyclopentyl-5-(furan-2-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidine 4s: white solid; yield 80%; mp: 93-95°C; 1 H NMR (400MHz, CDCl3) δ7.60(s,1H),7.19(s,1H),6.53(s,1H),4.13(s,4H),3.87(s,4H),3.60(p,J=8.1H z,1H),2.24(d,J=7.4Hz,2H),2.04(dd,J=12.2,6.9Hz,2H),1.91(s,2H),1.78(dd,J=13.8,8.0Hz,2H). 13 C NMR (100MHz, CDCl3) δ163.0,153.7,152.5,148.2,145.4,144.4,143.2,112.4,111.8,66.8,45.4,35.9,31.4,25.5.

[0177]

[0178] The above formula is 5-cyclohexyl-3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidine 4t: white solid; yield 87%; mp: 80-82°C; 1H NMR (400MHz, CDCl3) δ4.06 (s, 4H), 3.84 (s, 4H), 3.51 (m, 1H), 2.77 (t, J = 11.5Hz, 1H), 2.17 (s, 2H), 2.06 –1.95(m,4H),1.92–1.80(m,4H),1.74(s,3H),1.65–1.56(m,2H),1.45–1.35(m,2H),1.33–1.26(m,1H). 13 C NMR (100MHz, CDCl3) δ170.2,162.6,148.1,145.3,143.3,66.8,47.1,45.2,36.1,32.3,31.2,26.3,26.2,25.5.

[0179]

[0180] The above formula is 3-cyclopentyl-7-morpholin-5-(tetrahydro-2H-pyran-4-yl)isoxazolo[4,5-d]pyrimidine 4u: white solid; yield 90%; mp: 80-82°C; 1 H NMR (400MHz, CDCl3) δ4.07 (s, 6H), 3.84 (s, 4H), 3.59–3.47 (m, 3H), 3.01 (t, J = 11.1Hz ,1H),2.16(s,2H),2.09–1.97(m,4H),1.91(d,J=11.5Hz,4H),1.73(d,J=14.9Hz,2H). 13 C NMR (100MHz, CDCl3) δ168.1,162.5,148.2,145.3,143.4,67.9,66.8,45.4,43.8,36.1,31.8,31.1,25.5.

[0181] Example 5: Synthesis route of 1-(4-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)-3-methylurea 4w:

[0182]

[0183] After adding POCl3 (383 mg, 5.0 mmol) to a CH3CN (10 mL) solution of compound 3h (163 mg, 0.5 mmol), the temperature was raised to 80 ° C for 5 hours. After the reaction of the raw materials was complete, the reaction solution was cooled to room temperature, the mixture was concentrated under reduced pressure, and the residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 10 / 1, v / v). Then, morpholine (47 mg, 0.54 mmol) and Et3N (68 mg, 0.68 mmol) were added to a dichloromethane (10 mL) solution of the purified product (155 mg, 0.45 mmol) and reacted at room temperature for 1 hour. After the reaction of the raw materials was complete, the reaction mixture was diluted with water (10 mL), extracted with dichloromethane (10 mL × 3), dried with anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 20 / 1, v / v) to obtain compound S11 with a yield of 87%.

[0184] At 0°C, compound S11 (198 mg, 0.5 mmol) was dissolved in a solvent (10 mL, ethanol / water = 2 / 1), and NH4Cl (663 mg, 12.5 mmol) and zinc powder (325 mg, 5 mmol) were added. After the mixture was stirred at room temperature for 4 hours, it was filtered to remove the insoluble matter, and the filter cake was washed with ethanol. The filtrate was vacuum concentrated and purified on a silica gel column to obtain compound 4v with a yield of 78%.

[0185] To a dichloromethane solution (2 mL) of compound 4v (37 mg, 0.1 mmol) was added BTC (29.6 mg, 0.1 mmol) and Et3N (30.3 mg, 0.3 mmol) at 0°C, and the mixture was reacted for 15 minutes to obtain compound S12. Methylamine hydrochloride (34 mg, 0.5 mmol) was then added to the reaction solution at room temperature, and the mixture was reacted overnight. The reaction mixture was diluted with water (10 mL), extracted with EA (10 mL×3), dried over anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (ethyl acetate) to obtain white solid compound 4w in a yield of 93%.

[0186]

[0187] The above formula is 4-(3-cyclopentyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)aniline 4v: light yellow solid; yield 78%; mp: 190-192°C; 1H NMR(400MHz, CDCl3) δ8.28(d,J=8.3Hz,2H),6.77(d,J=8.3Hz,2H),4.17(s,4H),3.91(s,4H), 3.69–3.54(m,1H),2.26(d,J=6.3Hz,2H),2.22–2.12(m,2H),1.99(s,2H),1.88–1.78(m,2H). 13 C NMR (100MHz, DMSO-d6) δ162.4,160.6,151.4,148.2,145.4,143.1,129.7,125.2,113.6,66.4,45.6,36.0,31.1,25.7.

[0188]

[0189] The above formula is 1-(4-(3-cyclopentyl-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)-3-methylurea 4w: white solid; yield 93%; mp: 266-268°C; 1 H NMR (400MHz, CDCl3) δ8.77(s,1H),8.23(d,J=8.7Hz,2H),7.51(d,J=8.7Hz,2H),6.08(d,J=4.6Hz,1H),4.07–4.02(m,4H),3.82–3.77(m,4H),3 .51(p,J=8.1Hz,1H),2.66(d,J=4.6Hz,3H),2.15(dt,J=11.8,6.0Hz,2H),2.01(dd,J=12.5,7.3Hz,2H),1.91–1.83(m,2H),1.79–1.69(m,2H). 13 C NMR (100MHz, CDCl3) δ162.9,160.3,156.6,148.7,145.8,144.0,143.6,130.9,129.5,129.4,117.9,66.9,46.3,36.5,31.5,27.2,26.2.

[0190] Example 6: Synthesis route of (3-(isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5a-5j:

[0191]

[0192] After adding POCl3 (383 mg, 5.0 mmol) to a CH3CN (10 mL) solution of compounds 3n, 3aa-3ad (0.5 mmol), the temperature was raised to 80°C and the reaction was carried out for 5 hours. After the reaction of the raw materials was complete, the reaction solution was cooled to room temperature, the mixture was concentrated under reduced pressure, and the residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 10 / 1, v / v). Then, morpholine (47 mg, 0.54 mmol) and Et3N (68 mg, 0.68 mmol) were added to a dichloromethane (10 mL) solution of the purified product (0.45 mmol) and the reaction was carried out at room temperature for 1 hour. After the reaction of the raw materials was complete, the reaction mixture was diluted with water (10 mL), extracted with dichloromethane (10 mL×3), dried with anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 20 / 1, v / v) to obtain compound S13 with a yield of 85%.

[0193] To a solution of S13 (0.2 mmol) in anhydrous tetrahydrofuran (5 mL) was added LiAlH4 (0.2 mL, 0.2 mmol, 1.0 mol / L THF solution), and the mixture was reacted at 0 ° C for 30 minutes. The mixture was then reacted at room temperature for 1 hour. The reaction mixture was quenched with an aqueous NaOH solution (3 mL) and filtered to remove insoluble matter. The combined organic layer was dried over anhydrous Na2SO4 and concentrated under reduced pressure to give a white solid compound in a yield of 64-78%.

[0194]

[0195] The above formula is (3-(3-cyclopentyl-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5a: light yellow solid; yield 77%; mp: 134-136°C; 1 H NMR (400MHz, CDCl3) δ8.39 (s, 1H), 8.35 (s, 1H), 7.46 (d, J = 3.6Hz, 2H), 4.79 (s, 2H), 4.15 (s, 4H), 3.88 (s, 4H) ,3.58(p,J=7.8Hz,1H),2.24(d,J=7.1Hz,2H),2.16–2.07(m,2H),1.97(d,J=13.6Hz,2H),1.83–1.77(m,2H). 13 C NMR (100MHz, CDCl3) δ163.0,160.0,148.1,145.8,143.6,141.0,138.3,128.8,128.6,127.6,126.8,66.9,65.5,45.6,36.2,31.2,25.7.

[0196]

[0197] The above formula is (3-(3-cyclopentyl-7-((2S,6R)-2,6-dimethylmorpholino)isoxazolo[4,5-d]pyrimidin-5-yl)phenylmethanol 5b: white solid; yield 74%; mp: 130-132°C; 1 H NMR (400MHz, CDCl3) δ8.38(s,1H),8.34(t,J=4.5Hz,1H),7.46(t,J=6.2Hz,2H),4.88(d,J=10.4Hz,2H),4.80(s,2H),3.77(ddd,J=10.4,6.3,2. 4Hz,2H),3.59(p,J=8.1Hz,1H),2.92(s,2H),2.31–2.20(m,2H),2.16–2 .07(m,2H),1.99–1.91(m,2H),1.82–1.77(m,2H),1.33(d,J=6.2Hz,6H). 13 CNMR (100MHz, CDCl3) δ163.0,160.1,147.9,145.7,143.6,141.0,138.4,128.8,128.7,127.7,126.8,71.9,65.5,50.3,36.2,31.2,25.7,18.9.

[0198]

[0199] The above formula is (R)-(3-(3-cyclopentyl-7-(3-methylmorpholino)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5c: white solid; yield 73%; mp: 128-130°C; 1 H NMR (400MHz, CDCl3) δ8.39 (s, 1H), 8.35 (s, 1H), 7.46 (s, 2H), 5.02 (d, J = 4.4H z,1H),4.79(s,2H),4.70(d,J=12.8Hz,1H),4.10(d,J=10.6Hz,1H),3.92–3. 81(m,2H),3.69(t,J=11.8Hz,1H),3.59(dd,J=16.5,8.9Hz,2H),2.23(s,2H) ,2.12(s,2H),1.95(s,2H),1.84(s,1H),1.79(s,2H),1.48(d,J=6.6Hz,3H). 13CNMR (100MHz, CDCl3) δ163.0,160.1,148.1,145.7,143.7,141.0,138.4,128.7, 128.6,127.6,126.8,71.1,67.1,65.5,48.6,40.0,36.2,31.2,31.2,25.7,15.0.

[0200]

[0201] The above formula is (3-(3-cyclopentyl-7-((2-morpholinoethyl)amino)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5d: colorless oil; yield 64%; 1 H NMR(400MHz, CDCl3)δ8.43(s,1H),8.38(s,1H),7.46(s,2H),6.09(s,1H),4.79(s,2H),3.86(s,2H),3.76(s,4H), 3.58(dd,J=16.1,8.0Hz,1H),2.72(s,2H),2.55(s,4H),2.23(s,2H),2.16–2.09(m,2H),1.96(s,3H),1.79(s,2H). 13 C NMR (100MHz, CDCl3) δ163.2,160.5,147.9,143.5,141.1,138.4,128.7,128.6,127.6,126.8,66.9,65.4,56.9,53.4,36.3,31.3,25.7.

[0202]

[0203] The above formula is (3-(3-cyclopentyl-7-thiomorpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5e: white solid; yield 75%; mp: 159-161°C; 1 H NMR(400MHz, CDCl3)δ8.38(s,1H),8.36–8.31(m,1H),7.46(dd,J=4.2,1.1Hz,2H),4.79(s,2H),4.46–4.42(m,4H),3.58(p ,J=8.2Hz,1H),2.82–2.78(m,4H),2.23(dt,J=11.5,5.9Hz,2H),2.16–2.08(m,2H),1.99–1.91(m,2H),1.84–1.78(m,2H). 13C NMR (100MHz, CDCl3) δ163.0,160.1,147.7,145.9,143.6,141.0,138.3,128.7,128.6,127.6,126.7,65.4,48.4,36.2,31.2,27.4,25.7.

[0204]

[0205] The above formula is (3-(3-cyclopentyl-7-(4-methylpiperazin-1-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5f: white solid; yield 78%; mp: 170-172°C; 1 HNMR(400MHz, CDCl3)δ8.40(s,1H),8.35(dd,J=6.3,2.7Hz,1H),7.46(d,J=5.1Hz,2H),4.79(s,2H),4.18–4.13(m,4H),3.57(p,J=8.2Hz ,1H),2.60–2.56(m,4H),2.36(s,3H),2.23(dd,J=12.6,4.7Hz,2H),2.11(dd,J=12.7,7.6Hz,2H),1.98–1.90(m,2H),1.83–1.74(m,2H). 13 C NMR (100MHz, CDCl3) δ162.9,160.0,148.0,145.6,143.7,141.1,138.5,128.7,128.6,127.6,126.8,65.5,54.9,46.1,44.7,36.2,31.2,25.7.

[0206]

[0207] The above formula is (3-(3-cyclopentyl-7-(piperidin-1-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5g: white solid; yield 73%; mp: 155-157°C; 1 H NMR (400MHz, CDCl3) δ8.40(s,1H),8.38–8.35(m,1H),7.45(d,J=5.1Hz,2H),4.79(s,2H),4.11(d,J=5.5Hz,4H),3.59(dd,J=16 .4,8.2Hz,1H),2.24(dd,J=12.7,4.8Hz,2H),2.12(dd,J=12.7,7.6Hz,2H),1.95(dd,J=8.8,5.9Hz,2H),1.77(t,J=7.4Hz,8H).13 CNMR (100MHz, CDCl3) δ162.9,160.1,148.0,145.3,143.9,141.0,138.6,128.5,128.5,127.5,126.7,65.4,46.4,36.2,31.2,26.1,25.7,24.8.

[0208]

[0209] The above formula is (3-(3-cyclohexyl-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5h: light yellow solid; yield 74%; mp: 140-142°C; 1 H NMR(400MHz, CDCl3)δ8.39(s,1H),8.37–8.33(m,1H),7.47(d,J=4.8Hz,2H),4.80(s,2H),4.18–4.14(m,4H),3.9 2–3.87(m,4H),3.24(ddd,J=11.6,7.6,3.3Hz,1H),2.19(d,J=11.8Hz,2H),1.94–1.79(m,6H),1.52–1.42(m,2H). 13 CNMR (100MHz, CDCl3) δ163.3,160.2,148.3,145.8,143.7,141.1,138.5,128.9,128.8,127.8,126.9,67.0,65.7,45.8,35.6,30.7,26.2,26.1.

[0210]

[0211] The above formula is (3-(7-morpholin-3-(tetrahydro-2H-pyran-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5i: white solid; yield 72%; mp: 152-154°C; 1 HNMR (400MHz, CDCl3) δ8.39 (s, 1H), 8.35 (s, 1H), 7.48 (s, 2H), 4.81 (d, J = 5.2Hz, 2H), 4.16 (s, 4H), 4.1 2(s,2H),3.89(s,4H),3.67(t,J=11.3Hz,2H),3.49(s,1H),2.27–2.13(m,4H),1.85(t,J=5.7Hz,1H). 13C NMR (100MHz, CDCl3) δ161.6,160.3,148.1,145.4,143.8,141.1,138.1,128.9,128.7,127.7,126.8,67.7,66.8,65.4,45.7,32.8,30.1.

[0212]

[0213] The above formula is (3-(3-isopropyl-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5j: white solid; yield 72%; mp: 143-145°C; 1 H NMR (400MHz, CDCl3) δ8.40 (s, 1H), 8.36 (s, 1H), 7.47 (s, 2H), 4.80 (d, J = 4.2Hz, 2H), 4.1 6(s,4H),3.89(s,4H),3.52(dt,J=13.4,6.6Hz,1H),1.80(s,1H),1.57(d,J=6.8Hz,6H). 13 C NMR (100MHz, CDCl3) δ164.0,160.1,148.2,145.6,143.6,141.0,138.3,128.8,128.6,127.6,126.8,66.9,65.5,45.6,26.5,20.5.

[0214] Example 7: Synthesis route of acetyl chloride, ethyl and hydrogen substituted piperidinyl isoxazolo[4,5-d]pyrimidin-5-ylphenylmethanol 6a-6c:

[0215]

[0216] After adding POCl3 (383 mg, 5.0 mmol) to a CH3CN (10 mL) solution of compound 3ac (198 mg, 0.5 mmol), the temperature was raised to 80 ° C for 5 hours. After the reaction of the raw materials was complete, the reaction solution was cooled to room temperature, the mixture was concentrated under reduced pressure, and the residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 10 / 1, v / v). Then, morpholine (47 mg, 0.54 mmol) and Et3N (68 mg, 0.68 mmol) were added to a dichloromethane (10 mL) solution of the purified product (186 mg, 0.45 mmol) and reacted at room temperature for 1 hour. After the reaction of the raw materials was complete, the reaction mixture was diluted with water (10 mL), extracted with dichloromethane (10 mL × 3), dried with anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 20 / 1, v / v) to obtain compound S14 with a yield of 83%.

[0217] To a stirred solution of S14 (2.33 g, 5 mmol) in anhydrous tetrahydrofuran (5 mL) was added LiAlH4 (5 mL, 5 mmol, 1.0 mol / L THF solution) at 0 ° C, and the mixture was reacted for 6 hours. Afterwards, the reaction mixture was quenched with an aqueous NaOH solution (10 mL) and filtered to remove insoluble matter. The combined organic layer was dried over anhydrous Na2SO4, the mixture was concentrated under reduced pressure, and the mixture was purified by flash column chromatography (dichloromethane / methanol=10 / 1, v / v) to obtain white solid compound 6a at a yield of 15%, white solid compound 6b at a yield of 30%, and white solid compound 6c at a yield of 50%.

[0218]

[0219] The above formula is 1-(4-(5-(3-(hydroxymethyl)phenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-3-yl)piperidin-1-yl)ethan-1-one 6a: white solid; yield 15%; mp: 194-196°C; 1 H NMR (400MHz, CDCl3) δ8.39(s,1H),8.32(s,1H),7.47(s,2H),4.80(s,2H),4.65(d,J=13.1Hz,1H),4.16(s,4H),3.96(d,J=13.3Hz,1H),3.8 9(s,4H),3.48(s,1H),3.34(t,J=12.4Hz,1H),2.94(t,J=12.0Hz,1H),2.25(t,J=12.0Hz,2H),2.16(s,3H),2.06(dd,J=20.7,12.8Hz,2H). 13C NMR (100MHz, CDCl3) δ169.2,161.3,160.4,148.1,145.3,143.8,141.2,138.1,129.0, 128.7,127.6,126.8,66.8,65.4,60.5,46.3,41.4,33.6,29.8,29.2,21.6,21.1,14.3.

[0220]

[0221] The above formula is (3-(3-(1-ethylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6b: white solid; yield 30%; mp: 196-198°C; 1 HNMR(400MHz,DMSO-d6)δ10.92(s,1H),8.35(d,J=42.1Hz,2H),7.44(s,2H),5.37(s,1H),4.59(s,2H),4.08 (s,4H),3.82(s,4H),3.60(d,J=7.4Hz,3H),3.13(s,4H),2.39(s,4H),2.02(s,1H),1.27(d,J=31.3Hz,7H). 13 C NMR (100MHz, CDCl3) δ162.0,160.4,148.2,145.6,143.7,141.5,138.2,12 8.8,128.6,127.5,126.8,66.9,65.2,53.0,52.7,45.6,33.8,29.4,12.0.

[0222]

[0223] The above formula is (3-(7-morpholin-3-(piperidin-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6c: white solid; yield 50%; mp: 225-227°C; 1 H NMR (400MHz, DMSO-d6) δ9.28(s,1H),8.38(s,1H),8.33–8.28(m,1H),7.45(d,J=4.5Hz,2H),5.37(t,J=5.7Hz,1H),4.60(d,J= 5.3Hz,2H),4.08(s,4H),3.82(d,J=3.6Hz,4H),3.57(dtd,J=11.8,7.9,4.1Hz,2H),3.15(t,J=10.2Hz,2H),2.36–2.24(m,4H). 13CNMR(100MHz,DMSO-d6)δ160.5,159.6,147.9,144.5,143.6,142.8,137.1,128.6,128.2,126.5,126.2,66.0,62.9,45.2,42.5,30.7,25.8.

[0224] Example 8: Synthesis of piperidinylisoxazolo[4,5-d]pyrimidin-5-ylphenylmethanol 6d-6h containing different substituents:

[0225]

[0226] To a dichloromethane solution (5 mL) of compound 6c (79 mg, 0.2 mmol) was added acyl chloride or bromide or anhydride (0.24 mmol) and Et3N (41 mg, 0.4 mmol) at room temperature, and the mixture was reacted for 3 hours. After the reaction of the raw materials was complete, the reaction mixture was diluted with water (10 mL), extracted with DCM (10 mL × 3), dried over anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (petroleum ether / ethyl acetate = 1 / 1, v / v) to obtain the compound.

[0227]

[0228] The above formula is cyclopropyl (4-(5-(3-(hydroxymethyl)phenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-3-yl)piperidin-1-yl)methanone 6d: white solid; yield 95%; mp: 195-197°C; 1 H NMR (400MHz, CDCl3) δ8.38 (s, 1H), 8.36–8.31 (m, 1H), 7.47 (d, J = 5.2Hz, 2H), 4.80 (s, 2H) ,4.66(d,J=12.5Hz,1H),4.36(d,J=12.5Hz,1H),4.18–4.14(m,4H),3.91–3.88(m,4H),3. 56–3.48(m,1H),3.41(t,J=10.0Hz,1H),2.97(t,J=12.1Hz,1H),2.32–2.22(m,2H),2.11– 1.99(m,2H),1.86–1.79(m,1H),1.03(dd,J=7.3,3.1Hz,2H),0.79(dd,J=7.9,2.8Hz,2H). 13C NMR (100MHz, CDCl3) δ172.1,161.3,160.4,148.1,145.3,143.8,141.3,138.0,1 28.9,128.6,127.5,126.8,66.8,65.3,45.5,42.2,33.8,30.0,29.2,11.18,7.4.

[0229]

[0230] The above formula is tert-butyl 4-(5-(3-(hydroxymethyl)phenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-3-yl)piperidine-1-carboxylate 6e: white solid; yield 95%; mp: 92-94°C; 1 H NMR (400MHz, CDCl3) δ8.37(s,1H),8.33(d,J=6.3Hz,1H),7.46(d,J=6.2Hz,2H),4.79(s,2H),4.22(d,J=4.9Hz,2H),4.17–4.13(m,4H), 3.90–3.87(m,4H),3.38(ddd,J=11.3,7.7,3.8Hz,1H),3.01(t,J=10.3Hz,2H),2.18(d,J=11.4Hz,2H),2.08–2.02(m,2H),1.50(s,9H). 13 C NMR (100MHz, CDCl3) δ161.7,160.4,154.9,148.1,145.4,143.8,141.3,138.1 ,128.9,128.7,127.6,126.8,79.8,66.9,65.4,45.8,43.4,33.8,29.4,28.6.

[0231]

[0232] The above formula is (4-(5-(3-(hydroxymethyl)phenyl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-3-yl)piperidin-1-yl)phenyl)methanone 6f: white solid; yield 94%; mp: 106-108°C; 1 H NMR (400MHz, CDCl3) δ8.40(s,1H),8.34(t,J=4.4Hz,1H),7.45(dd,J=15.7,3.8Hz,7H),4.79(s,2H),4. 19–4.15(m,4H),3.92–3.88(m,4H),3.58–3.50(m,1H),3.22(d,J=44.8Hz,2H),2.16(t,J=67.9Hz,6H).13 CNMR (100MHz, CDCl3) δ170.7,161.3,160.5,148.2,145.4,144.0,141.3,138.1,136.2 ,129.8,129.1,128.8,128.7,127.7,127.0,126.9,66.9,65.5,47.7,33.8,30.0,29.5.

[0233]

[0234] The above formula is (3-(3-(1-(methylsulfonyl)piperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6g: white solid; yield 95%; mp: 222-224°C; 1 H NMR (400MHz, CDCl3) δ8.39(s,1H),8.33(dd,J=6.3,2.6Hz,1H),7.47(d,J=5.1Hz,2H),4.80(s,2H),4.19 –4.15(m,4H),3.92–3.88(m,6H),3.44–3.34(m,1H),3.07–2.99(m,2H),2.86(s,3H),2.35–2.26(m,4H). 13 C NMR (100MHz, CDCl3) δ161.0,160.6,148.2,145.4,144.0,141.2,138.1,129.1,128.8,127.7,126.9,66.9,65.5,45.7,35.2,32.9,29.0.

[0235]

[0236] The above formula is (3-(3-(1-benzylpiperidin-4-yl)-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6h: white solid; yield 96%; mp: 87-89°C; 1 H NMR (400MHz, CDCl3) δ8.40(s,1H),8.33(t,J=4.4Hz,1H),7.45(d,J=5.0Hz,2H),7.36(dd,J=11.1,7.1Hz,4H),7.29(s,1H),4.79( s,2H),4.16–4.12(m,4H),3.90–3.86(m,4H),3.61(s,2H),3.27–3.18(m,1H),3.04(d,J=8.4Hz,2H),2.22(dd,J=14.7,5.3Hz,6H).13 C NMR (100MHz, CDCl3) δ162.1,160.3,148.1,145.6,143.7,141.3,138.2,137.8,129.5 ,128.8,128.6,128.4,127.6,127.3,126.8,66.9,65.3,63.3,53.3,45.7,33.7,29.5.

[0237] Example 9: Synthesis of (3-(7-morpholine-3-(1-(pyridin-3-ylmethyl)piperidin-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6i:

[0238]

[0239] 4-(Bromomethyl)pyridine hydrobromide (61 mg, 0.24 mmol) and Et3N (41 mg, 0.4 mmol) were added to a solution of compound 6c (79 mg, 0.2 mmol) in N,N-dimethylformamide (5 mL) at 60°C, and the mixture was reacted overnight. The reaction mixture was diluted with water (10 mL), extracted with EA (10 mL×3), dried over anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (dichloromethane / methanol=30 / 1, v / v) to give compound 6i as a white solid with a yield of 99%.

[0240]

[0241] The above formula is (3-(7-morpholine-3-(1-(pyridin-3-ylmethyl)piperidin-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6i: white solid; yield 99%; mp: 158-160°C; 1 H NMR(400MHz, CDCl3)δ8.66(s,1H),8.53–8.50(m,1H),8.46(s,1H),8.33(dd, J=6.2,2.7Hz,1H),7.72(d,J=7.7Hz,1H),7.46(d,J=5.1Hz,2H),7.31–7.27( m,1H),4.81(s,2H),4.17–4.14(m,4H),3.91–3.87(m,4H),3.61(s,2H),3.25 (s,1H),3.01(d,J=5.9Hz,2H),2.30(d,J=8.5Hz,4H),2.16(d,J=6.6Hz,2H). 13C NMR (100MHz, CDCl3) δ162.1,160.4,150.5,148.5,148.2,145.7,143.8,141.5,138.3,13 6.9,134.1,129.0,128.6,127.6,126.9,123.5,66.9,65.3,60.5,53.4,45.9,33.6,29.6.

[0242] Example 10: Synthesis of (3-(3-(1-(2-methoxyethyl)piperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6j:

[0243]

[0244] 1-Bromo-2-methoxyethane (33 mg, 0.24 mmol) and DIPEA (52 mg, 0.4 mmol) were added to an ethanol solution (5 mL) of compound 6c (79 mg, 0.2 mmol) at 80°C, and the mixture was reacted overnight. The mixture was concentrated under reduced pressure, and the residue was purified by flash column chromatography (petroleum ether / ethyl acetate=10 / 1, v / v) to obtain a light yellow solid compound 6j with a yield of 89%.

[0245]

[0246] The above formula is (3-(3-(1-(2-methoxyethyl)piperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6j: light yellow solid; yield 89%; mp: 200-202°C; 1 H NMR (400MHz, CDCl3) δ8.43(s,1H),8.32(td,J=4.7,1.6Hz,1H),7.44(d,J=4.9Hz,2H),4.79(s,2H),4.17–4.13(m,4H),3.90–3.87(m,4H),3. 59(t,J=5.6Hz,2H),3.38(s,3H),3.28–3.19(m,1H),3.14(d,J=8.8Hz,2H),2.70(t,J=5.5Hz,2H),2.33(dd,J=16.7,9.4Hz,4H),2.23(s,2H). 13C NMR (100MHz, CDCl3) δ161.9,160.4,148.2,145.6,143.8,141.5,138.2,128. 8,128.6,127.6,126.8,70.2,66.9,65.3,59.0,58.1,53.8,45.6,33.5,29.2.

[0247] Example 11: Synthesis of (3-(3-(1-cyclopentylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6k:

[0248]

[0249] To a solution of compound 6c (79 mg, 0.2 mmol) in N,N-dimethylformamide (5 mL) was added bromocyclopentane (36 mg, 0.24 mmol) and Et3N (41 mg, 0.4 mmol) at 80°C, and the mixture was reacted overnight. The reaction mixture was diluted with water (10 mL), extracted with EA (10 mL×3), dried over anhydrous Na2SO4, and concentrated in vacuo. The residue was purified by flash column chromatography (dichloromethane / methanol=20 / 1, v / v) to give compound 6k as a white solid with a yield of 62%.

[0250]

[0251] The above formula is (3-(3-(1-cyclopentylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6k: white solid; yield 62%; mp: 187-189°C; 1 HNMR (400MHz, CDCl3) δ8.44(s,1H),8.30(t,J=3.6Hz,1H),7.44(d,J=4.6Hz,2H),4.79(s,2H),4.18–4.14(m,4H),3.90–3.86 (m,4H),3.21(s,2H),2.62(dd,J=15.5,7.8Hz,1H),2.27–2.21(m,7H),1.92(d,J=8.5Hz,2H),1.73(s,2H),1.60–1.50(m,4H). 13 CNMR (100MHz, CDCl3) δ161.3,160.5,148.1,145.5,143.8,141.7,138.1,128. 9,128.5,127.4,127.0,68.0,66.9,65.1,52.0,45.7,33.1,29.7,28.4,24.1.

[0252] Example 12: Synthesis route of 4-(3-(1-benzylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)aniline 7a:

[0253]

[0254] Compound S6 (369 mg, 1.0 mmol) was dissolved in 5 mL of ammonia methanol solution and stirred at room temperature for 3 h. The solvent was evaporated under reduced pressure to obtain a white product S15 with a yield of 99%.

[0255] Compound S15 (340 mg, 1.0 mmol) was dissolved in dichloromethane (5 mL), 1.5 mL TFA was added and stirred at room temperature for 2 h. The solvent was evaporated under reduced pressure to obtain trifluoroacetate salt of S16 as pale yellow needle-like crystals with a yield of 100%.

[0256] The trifluoroacetate of compound S16 (354 mg, 1.0 mol) was suspended in acetonitrile and 2 eq. of triethylamine was added and stirred at room temperature for 2 h for dissociation. The residue was evaporated to dryness under reduced pressure to remove the residual triethylamine. 5 mL of anhydrous acetonitrile and DIPEA (194 mg, 1.5 mmol) were added to the residue and the temperature was lowered to 0°C. Then, a solution of benzyl bromide (204 mg, 1.2 mmol) in anhydrous acetonitrile was slowly added dropwise at 0°C. After the addition was complete, the reaction was stirred at room temperature. After TLC monitoring, the reaction was completed, and the solution was concentrated under reduced pressure. A saturated sodium carbonate solution was added to the resulting solution and extracted with chloroform. Then, the light yellow solid S17 was obtained with a yield of 81% by column chromatography purification.

[0257] Compound S17 (264 mg, 0.8 mmol) was suspended in 3 mL of a mixed solvent (ethanol: water = 2:1) and cooled to 0°C. Then, ammonium chloride (1.06 g, 20 mmol) was added, and zinc powder (512 mg, 8 mmol) was slowly added in batches, and the mixture was reacted at room temperature for 4 h. The insoluble matter was removed by suction filtration, the filter cake was washed, the filtrate was evaporated to dryness under reduced pressure, and the light yellow solid S18 was obtained by column chromatography purification with a yield of 78%. 1 HNMR (300MHz, CDCl3) δ7.40–7.26(m,5H),5.81(d,J=158.2Hz,2H),4.29(s,2H),3.55(s,2H),3.00(d,J =11.9Hz,2H),2.64(dd,J=15.6,7.9Hz,1H),2.12(dd,J=16.4,9.0Hz,2H),1.95(dt,J=6.8,3.6Hz,4H).

[0258] Compound S18 (180 mg, 0.6 mmol) was dissolved in 2 mL of anhydrous THF, triphosgene (90 mg, 0.3 mmol) was added, and the mixture was heated to reflux for 1 h. The reaction mixture was cooled to room temperature, and the white precipitate was collected by suction filtration and washed with ethyl acetate to obtain the hydrochloride of the target compound S19 with a yield of 90%.

[0259] The mixture of the hydrochloride of compound S19 (163 mg, 0.5 mmol) and 5 mL of phosphorus oxychloride was heated and refluxed for 24 h, then concentrated under reduced pressure, and the residue was poured into crushed ice to quench. Ethyl acetate was added to the aqueous phase and the pH was adjusted to 8-9 with 6N NaOH aqueous solution, then extracted, the organic phase was washed with water until neutral, washed three times with saturated sodium chloride, dried with anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The residue was dissolved in 8 mL of dichloromethane, cooled in an ice-salt bath, and then TEA (101 mg, 1 mmol) was added. A DCM solution of morpholine (52 mg, 0.6 mmol) was added dropwise to the above solution and stirred for 10 min. After TLC monitoring of the reaction was complete, water was added to dilute the reaction solution, DCM was extracted, the organic phase was washed with water and saturated sodium chloride, and the solvent was evaporated and column chromatography was performed to obtain a light yellow solid S20 with a yield of 60%. 1 H NMR (300MHz, CDCl3) δ7.39–7.27(m,5H),7.24(dd,J=5.9,2.0Hz,1H),4.20–3.97(m,4H),3.83(dd,J=6.1,3.6Hz,4H ),3.55(s,2H),3.17–3.07(m,1H),3.00(d,J=11.7Hz,2H),2.14(dtd,J=13.7,10.8,3.2Hz,6H),2.03–1.97(m,1H).

[0260] Compound S20 (124 mg, 0.3 mmol), p-aminophenylboronic acid pinacol ester (99 mg, 0.45 mmol), Pd(Pcy3)2Cl2 (11 mg, 0.015 mmol) and CsF (137 mg, 0.9 mmol) were placed in a three-necked flask, the air was evacuated under reduced pressure and then filled with N2, and the process was repeated three times; then, 3 mL of solvent (NMP / water = 9 / 1) was injected into the three-necked flask. After the reaction mixture was reacted at 100 ° C for 48 h under N2 gas conditions, water was added to dilute and extracted with ethyl acetate, the organic phase was washed with water and saturated sodium chloride three times each, the solvent was removed under reduced pressure, and the residue was purified by column chromatography to obtain a yellow solid 7a with a yield of 32%.

[0261]

[0262] The above formula is 4-(3-(1-benzylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)aniline 7a: yellow solid; yield 32%; mp: 118-119°C. 1 H NMR (300MHz, CDCl3) δ8.26–8.21(m,2H),7.41–7.26(m,5H),6.75–6.70(m,2H),4.14–4.11(m, 4H),3.88–3.85(m,6H),3.63(s,2H),3.30–3.13(m,1H),3.07–3.03(m,2H),2.42–2.09(m,6H).

[0263] Example 13: Synthesis of benzylpiperidinylmorpholineisoxazolo[4,5-d]pyrimidine-5-urea 7b and 7c containing different substituents:

[0264]

[0265] Compound 7a (94 mg, 0.2 mmol) was dissolved in 2 mL of anhydrous DCM and triethylamine (61 mg, 0.6 mmol) was added. The mixture was cooled in an ice-water bath, triphosgene (30 mg, 0.1 mmol) was added, and the mixture was stirred for 15 min in an ice-water bath, and then 5 eq. of amine and 6 eq. of triethylamine were added, and the reaction was continued overnight. After the reaction was completed, water was added to the reaction solution and extracted with DCM, and the organic phase was washed with water and saturated sodium chloride three times, and the solvent was evaporated under reduced pressure. The residue was purified by column chromatography or preparative thin layer chromatography to obtain the corresponding urea derivatives 7b and 7c in a yield of 30-50%.

[0266]

[0267] The above formula is 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)-3-methylurea 7b: yellow solid; yield 50%; mp: 147-149°C; 1 H NMR (300MHz, DMSO-d6) δ8.79(s,1H),8.23(d,J=8.7Hz,2H),7.52(d,J=8.7Hz,2H),7.35–7.25(m,5H),6.08(q,J=4.8Hz,1H),4.05–4.03(m,4 H),3.81–3.78(m,4H),3.54(s,2H),3.17–3.09(m,1H),2.93(d,J=11.4Hz,2H),2.66(d,J=4.5Hz,3H),2.22–2.10(m,4H),2.04–1.92(m,2H).

[0268]

[0269] The above formula is 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)-3-(4-(4-methylpiperazin-1-yl)phenyl 7c: yellow solid; yield 30%; mp: 144-145°C; 1 HNMR(300MHz,DMSO-d6)δ9.00(s,1H),8.54(s,1H),8.30–8.26(m,2H),8.16(d,J=3.0Hz,1 H),7.71(dd,J=9.0,2.7Hz,1H),7.58–7.53(m,2H),7.36–7.25(m,5H),6.83(d,J=9.0Hz,1H ),4.06–4.04(m,4H),3.81–3.79(m,4H),3.56(s,2H),3.43–3.40(m,4H),3.20–3.12(m,1H) ,2.97–2.93(m,2H),2.48–2.45(m,4H),2.26(s,3H),2.26–2.12(m,4H),2.05–1.91(m,2H).

[0270] Example 14: Synthetic routes of piperidinylmorpholineisoxazolo[4,5-d]pyrimidine-5-urea 7d and 7e containing different substituents:

[0271]

[0272] Compound 7b (527 mg, 1.0 mmol) was dissolved in 10 mL of 1,2-dichloroethane and ACE-Cl (1.1 g, 8 mmol) was added and stirred at room temperature overnight. After the reaction was completed, the solvent was evaporated under reduced pressure, methanol was added and refluxed for 2 h. Methanol was evaporated under reduced pressure to obtain a crude product (551 mg) of debenzylated hydrochloride. 10 mL of DCM and triethylamine (354 mg, 3.5 mmol) were added to the crude product and cooled to 0 ° C in an ice-water bath. A DCM solution of (Boc)2O (240 mg, 1.1 mmol) was added dropwise to the above reaction solution and stirred for 3 h. Water and DCM were added for extraction, and the organic phase was washed with water and saturated sodium chloride three times. The organic phase was concentrated under reduced pressure and purified by column chromatography to obtain white solid S21 with a yield of 76%.

[0273] Compound S21 (385 mg, 0.7 mmol) was dissolved in 5 mL of DCM, and 3 mL of trifluoroacetic acid was added. The mixture was stirred at room temperature for 2 h, and the solvent was evaporated under reduced pressure to obtain the ditrifluoroacetate salt of yellow solid S22 with a yield of 99%.

[0274] The ditrifluoroacetate of compound S22 (133 mg, 0.2 mmol) was suspended in anhydrous dichloromethane, and 2.0 eq. of triethylamine was added, and the mixture was cooled to 0° C. in an ice bath. Then, 1.2 eq. of anhydride or bromide in anhydrous dichloromethane solution was added dropwise to the reaction solution, and the reaction was continued to be stirred overnight or monitored by TLC. After the reaction was completed, saturated sodium bicarbonate was added, and dichloromethane was extracted. The organic phase was washed three times with water and saturated sodium chloride, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by column chromatography to obtain the corresponding piperidine derivatives 7d and 7e in a yield of 63-75%.

[0275]

[0276] The above formula is 1-methyl-3-(4-(7-morpholin-3-(1-(pyridin-3-ylmethyl)piperidin-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)urea 7d: yellow solid; yield 63%; mp: 146-148°C; 1 H NMR(300MHz,DMSO-d6)δ8.79(s,1H),8.58–8.53(m,1H),8.52–8.46(m,1H), 8.28–8.18(m,2H),7.84–7.7(m,1H),7.57–7.46(m,2H),7.43–7.33(m,1H),6 .09(q,J=4.8Hz,1H),4.19–3.91(m,4H),3.90–3.70(m,4H),3.59(s,2H),3.2 3–3.05(m,1H),3.04–2.80(m,2H),2.67(d,J=4.2Hz,3H),2.22–1.92(m,6H).

[0277]

[0278] The above formula is 1-methyl-3-(4-(7-morpholin-3-(1-(2,2,2-trifluoroacetyl)piperidin-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)urea 7e: white solid; yield 75%; mp: 203-205°C; 1H NMR (300MHz, DMSO-d6) δ8.77(s,1H),8.24–8.21(m,2H),7.53–7.50(m,2H),6.07(q,J=4.8Hz,1H),4.39–4.32(m,1H),4.06–3.9 8(m,5H),3.81–3.78(m,4H),3.64–3.52(m,2H),3.30–3.22(m,1H),2.66(d,J=4.5Hz,3H),2.36–2.27(m,2H),2.08–1.86(m,2H).

[0279] Example 15: Determination of the anti-new coronavirus activity of compounds

[0280] Experimental method: 1×10 4 Vero-E6 cells (ATCC CCL81) were inoculated into 96-well plates (10% FBSDMEM) one day in advance. After the cells adhered, drug and virus inoculation began. Use maintenance solution (2% FBSDMEM) to dilute the drug with a mother solution concentration of 10mM to a final concentration of 20μM. After dilution, discard the cell culture medium in the 96-well culture plate and add 100μL of diluted drug to each well. At the same time, use maintenance solution to dilute the new coronavirus 10-fold, for a total of 8 concentration gradients. Subsequently, 100μL of the gradient diluted virus solution was added to the cell wells containing the drug, with 3 replicates for each dilution. The final drug concentration in the cells was 10μM. After culturing in a cell culture incubator (37°C, 5% CO2) for 72 hours, the cytopathic effect was observed, and the half tissue infection dose (TCID 50 / mL). In the experiment, the drug-free experimental group was used as a negative control, and remdesivir was used as a positive control.

[0281] The specific experimental results are as follows:

[0282] Positive control drug remdesivir IC 50 =3.97μM.

[0283] The following compounds have inhibitory activity against Beta, Delta and Omicron strains of Sara-CoV-2 in Vero-E6 cells (TCID 50 / mL≤1.5μM).

[0284]

[0285] (3-(3-Cyclopentyl-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 5a

[0286]

[0287] (3-(3-(1-Benzylpiperidin-4-yl)-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)methanol 6h

[0288]

[0289] 4-(3-(1-Benzylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)aniline 7a

[0290]

[0291] 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinoisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)-3-methylurea 7b

[0292]

[0293] 1-(4-(3-(1-benzylpiperidin-4-yl)-7-morpholinisoxazolo[4,5-d]pyrimidin-5-yl)phenyl)-3-(4-(4-methylpiperazin-1-yl)phenyl 7c

[0294]

[0295] 1-Methyl-3-(4-(7-morpholin-3-(1-(pyridin-3-ylmethyl)piperidin-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)urea 7d

[0296]

[0297] 1-Methyl-3-(4-(7-morpholino-3-(1-(2,2,2-trifluoroacetyl)piperidin-4-yl)isoxazolo[4,5-d]pyrimidin-5-yl)phenyl)urea 7e

[0298] The above description is only used to illustrate the technical solutions and features of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can easily think of various equivalent modifications or replacements within the technical scope disclosed by the present invention, and these modifications or replacements should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention shall be based on the protection scope of the claims.

Claims

1. A novel isoxazolopyrimidine derivative having anti-new coronavirus activity, and a pharmaceutically acceptable salt, solvate or deuterated product thereof, wherein the structural formula of the novel isoxazolopyrimidine derivative having anti-new coronavirus activity is shown in general formula I: When R 1 When R is selected from substituted or unsubstituted alkyl or cycloalkyl, substituted or unsubstituted aryl; 2 is selected from substituted or unsubstituted alkyl or cycloalkyl, substituted or unsubstituted aryl; said R 3 is selected from a substituted or unsubstituted morpholine ring, a substituted or unsubstituted thiomorpholine ring, a substituted or unsubstituted piperidine ring, a substituted or unsubstituted piperazine ring, a substituted or unsubstituted pyridazine ring, a substituted or unsubstituted pyrrole ring, a substituted or unsubstituted imidazole ring, a substituted or unsubstituted pyrazole ring, and a substituted or unsubstituted amine group; R 1 and R 2 The alkyl, cycloalkyl and aryl groups are optionally substituted with 1, 2, 3, 4 or 5 R; R is selected from the group consisting of hydrogen, deuterium, alkene, alkynyl, halogen, hydroxy, hydroxymethyl, amino, trifluoromethyl, cyano, ester, carbonyl, acyl, aminoacyl, amide, sulfonyl, aminosulfonyl, sulfonamido, alkyl, alkoxy, urea, thiourea, aryl, pyridyl, pyrimidinyl, pyridazinyl, aminopyrimidinyl, quinolyl, benzofuranyl, benzindolyl, benzindazolyl, benzothiophenyl, benzopyrazolyl, furanyl, thienyl, pyridyl, oxazolyl and pyrazolyl; R 1 Selected from cyclopentyl, 4-piperidinyl; R 2 Selected from 3-hydroxymethylphenyl, 4-aminophenyl, 4-ureidophenyl; R 3 is selected from morpholinyl; The viruses include SARS-CoV-2.

2. The method for preparing a novel isoxazolopyrimidine derivative having anti-new coronavirus activity according to claim 1, characterized in that: The steps include: Step 1: adding 4-aminoisoxazole-5-carboxamide compounds into a reactor; Step 2: adding an aldehyde compound into a reactor; Step 3: Adding the catalyst into the reactor; Step 4: Adding solvent into the reactor; Step 5: After the mixed solution is reacted at reflux temperature for a certain period of time, the reaction is quenched; Step 6: adding an organic solvent for extraction, drying and concentrating the organic phase, and purifying to obtain the isoxazolopyrimidine compound represented by the general formula I.

3. A method for preparing a novel isoxazolopyrimidine derivative having anti-new coronavirus activity according to claim 2, characterized in that: In step 1, the reactor is sealed; In step 3, the catalyst is I2 and DMSO; In step 4, the solvent is CH3CN; In step 5, the reaction is quenched with an aqueous solution of sodium thiosulfate; In step 6, ethyl acetate is added for extraction, and the organic phase is dried, concentrated, and then purified to obtain the isoxazolopyrimidine compound represented by the general formula I.

Images