CN117327077A

CN117327077A - Anilinopyrimidino heterocyclic compound, and preparation method, pharmaceutical composition and application thereof

Info

Publication number: CN117327077A
Application number: CN202311281729.6A
Authority: CN
Inventors: 韩进松; 张文慧; 吴斐华; 黄慧; 车赛; 林范洁; 周玲佳; 肖文琪; 高旭
Original assignee: China Pharmaceutical University
Current assignee: China Pharmaceutical University
Priority date: 2023-10-07
Filing date: 2023-10-07
Publication date: 2024-01-02

Abstract

The invention discloses an anilinopyrimidino heterocyclic compound, a preparation method, a pharmaceutical composition and application thereof. The compound has a structure as shown in a formula I and also comprises pharmaceutically acceptable salts thereof, can effectively inhibit WEE1 and HDAC, has an inhibition value of more than 90% optimally, and can effectively inhibit tumor cell proliferation, and the inhibition rate of more than 50%; the application is wide, the drug effect can be exerted on the molecular level and the cellular level, and the nanomolar concentration level can be optimally achieved; in addition, the preparation method is easy to expand the structure and has good universality.

Description

Anilinopyrimidino heterocyclic compound, and preparation method, pharmaceutical composition and application thereof

Technical Field

The invention relates to an anilinopyrimidino heterocyclic compound and a preparation method, a pharmaceutical composition and application thereof, in particular to an anilinopyrimidino heterocyclic compound with WEE1 and HDAC double-target inhibition activity and a preparation method, a pharmaceutical composition and application thereof.

Background

The main mechanism of action of WEE1 inhibitors is through synthetic lethal effects with the P53 gene. Because most tumors have mutation or deletion of the P53 gene, the tumors can only act depending on WEE1 kinase-mediated G2/M phase cell cycle checkpoints, and cells carrying DNA damage are blocked in the G2/M phase, so that time is provided for DNA damage repair. Inhibition of WEE1 kinase can lead cells carrying DNA damage to enter mitosis in advance, and cause mitosis disasters and apoptosis. A number of WEE1 inhibitors are currently in clinical stages, such as ZN-C3 (phase II) from Zentalis, debio-0123 (phase I/II) from Debio, IMP7068 (phase I) from Shanghai, mecang Pi pharmaceutical industry, SC0191 (phase I) developed by Shi Kang Hongren and Ming Kangde, and the like. However, the pathogenesis of tumors is complex, often involving a variety of different signaling pathways, and the effect of single-target drugs is often unsatisfactory. It has been reported that acute myeloid leukemia cells treated with WEE1 inhibitors produce an anti-proliferative effect that is reduced by WEE1 inhibitors, possibly due to CHK 1-mediated activation of compensatory pathways.

HDACs fall into four classes I, II, III, IV of 18 subtypes, with overexpression of types I, II, IV leading to normal cell cycle and metabolic behavior changes, thereby inducing tumors. Based on the fact that a plurality of HDAC inhibitors are commercially available, such as Vorinostat, romidepsin, belinostat and Panobinostat approved by the FDA in the United states, the independently developed Sidamine in China is also commercially available in China in 2015, and the drugs play a great role in the treatment of blood tumors. Studies show that HDAC participates in regulating DNA damage repair pathways, has obvious synergistic effect with WEE1 inhibitors, and can inhibit compensatory activation pathways caused by WEE1 inhibitors in various tumor cells, so that the tumor cells are re-sensitized to the WEE1 inhibitors.

Considering the problems of toxic and side effects, uneven pharmacokinetic properties and the like caused by drug-drug interaction of the combined drug, the development of WEE1/HDAC double-target drugs has very important significance and potential application value.

Disclosure of Invention

The invention aims to: the first object of the invention is to provide an anilinopyrimidino heterocyclic compound, the second object is to provide a preparation method of the compound, the third object is to provide a pharmaceutical composition containing the compound, and the fourth object is to provide an application of the compound and the pharmaceutical composition.

The technical scheme is as follows: the anilinopyrimidino heterocyclic compound disclosed by the invention has a structure shown in a formula I and further comprises pharmaceutically acceptable salts thereof:

wherein:

R ₁ selected from substituted or unsubstituted C _1-4 Alkyl, C _2-4 Alkenyl, 5-7 membered aryl, 5-7 membered heteroaryl containing 1-3N, O, S substituents selected from one or more hydrogen, halogen, C _1-6 Alkyl, C _1-6 Haloalkyl, C _1-6 Alkoxy, cyano, nitro, hydroxy, amino;

R ₂ selected from one or more of hydrogen, halogen, C _1-6 Alkyl, C _1-6 Haloalkyl, C _1-6 Alkoxy, cyano, nitro, hydroxy, amino;

R ₃ selected from hydroxy, substitutedOr unsubstituted 5-7 membered aryl, 5-7 membered heteroaryl containing 1-3N, O, S, said substituents being selected from one or more hydrogen, halogen, C _1-6 Alkyl, C _1-6 Haloalkyl, C _1-6 Alkoxy, cyano, nitro, hydroxy, amino;

x is selected fromWherein R is _a Selected from hydrogen, halogen, C _1-4 Alkoxy, 3-7 membered cycloalkoxy, C _1-4 Haloalkyl, C _1-4 Hydroxyalkyl, C _1-4 Alkoxy substituted C _1-4 Alkyl, cyano, nitro, amino, hydroxy, Q is selected from CH, N, R _b Selected from substituted or unsubstituted C _1-4 Alkyl, C _2-4 Alkenyl, 5-7 membered aryl containing 1-3N, O, S aryl heteroatoms selected from one or more of hydrogen, halogen, C _1-6 Alkyl, C _1-6 Haloalkyl, C _1-6 Alkoxy, cyano, nitro, hydroxy, amino;

y is selected from- (CH) ₂ ) _n -、-(CH ₂ ) _n O-、-(CH ₂ ) _n S-、-(CH ₂ ) _n CONH-、-(CH ₂ ) _n NHCO-、-(CH ₂ ) _n SO ₂ NH-、-(CH ₂ ) _n NHSO ₂ -、Where n=0-7.

Preferably, in the structure:

R ₁ selected from substituted or unsubstituted C _1-4 Alkyl, C _2-4 Alkenyl, phenyl, said substituents being selected from one or more of hydrogen, halogen, C _1-4 Alkyl, C _1-4 Haloalkyl, C _1-4 An alkoxy group;

R ₂ selected from one or more of hydrogen, halogen, C _1-4 An alkyl group;

R ₃ selected from hydroxy, substituted phenyl, said substituents selected from halogen, amino;

x is selected fromWherein R is _a Selected from hydrogen, halogen, C _1-4 Alkoxy, 3-5 membered cycloalkoxy, C _1-4 Haloalkyl, C _1-4 Hydroxyalkyl, C _1-4 Alkoxy substituted C _1-4 Alkyl, Q is selected from CH, N, R _b Selected from C _1-4 An alkyl group;

y is selected from- (CH) ₂ ) _n -、-(CH ₂ ) _n O-、-(CH ₂ ) _n CONH-、Where n=0-7.

Preferably, in the structure:

R ₁ selected from the group consisting of

R ₂ Selected from one or more of hydrogen, methyl, halogen;

R ₃ selected from hydroxy, o-aminophenyl, 2-amino-4-fluorophenyl;

x is selected from

Preferably, in the structure:

R ₁ selected from the group consisting of

R ₂ Selected from hydrogen;

R ₃ selected from hydroxyl groups;

x is selected from

Y is selected from- (CH) ₂ ) _n O-、-(CH ₂ ) _n CONH-、Where n=6, 7.

Specifically, the anilinopyrimidino heterocyclic compound is selected from any one of the following compounds:

preferably, the pharmaceutically acceptable salt is a salt of the anilinopyrimidino heterocyclic compound with an acid selected from any one of the following: hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, carbonic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, citric acid, malic acid, tartaric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, succinic acid, fumaric acid, salicylic acid, phenylacetic acid, mandelic acid, ferulic acid.

The preparation method of the compound is selected from any one of the following methods:

(1) When R is ₃ When the compound is hydroxyl, the methyl mercapto compound is prepared into the compound I through coupling and ammonolysis:

the compound IV is prepared by reacting the compound V with mCPBA, wherein the solvent is selected from chloroform, dichloromethane, toluene, tetrahydrofuran, 1, 4-dioxane, ethyl acetate, acetone, N-dimethylformamide or a mixed solvent of any two, and dichloromethane is preferred.

Preparing a compound II by reacting the compound IV with a compound III, wherein the base is selected from triethylamine, N-diisopropylethylamine, preferably N, N-diisopropylethylamine; the solvent is selected from chloroform, dichloromethane, toluene, tetrahydrofuran, 1, 4-dioxane, ethyl acetate, acetone, N-dimethylformamide or a mixed solvent of any two, preferably toluene.

Preparing a compound IA by reacting the compound II with hydroxylamine hydrochloride, wherein the base is selected from sodium ethoxide, potassium acetate, sodium hydroxide, potassium carbonate, sodium carbonate, preferably potassium hydroxide; the solvent is selected from methanol, ethanol, n-propanol, isopropanol, tert-butanol, n-butanol, isobutanol, preferably methanol.

(2) When R is ₃ When the compound is not hydroxyl, the methyl mercapto compound is prepared by coupling, hydrolyzing and acylating the methyl mercapto compound to obtain the compound I:

Preparing a compound IIB from the compound II through hydrolysis, wherein the base is selected from lithium hydroxide, sodium hydroxide, potassium carbonate and sodium carbonate, and sodium hydroxide is preferred; the solvent is selected from tetrahydrofuran, 1, 4-dioxane, methanol, ethanol and water, preferably tetrahydrofuran and water.

From compounds IIB and R ₃ -NH ₂ The condensing agent used for preparing the compound IB is selected from Carbonyl Diimidazole (CDI), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 1- (3-dimethylaminopropyl) -3-Ethylcarbodiimide (EDCI), 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU) or benzotriazol-1-yl-oxy-tripyrrolidinylphosphine hexafluorophosphate (PyBOP), preferably HATU; the acid binding agent is selected from N, N-diisopropylethylamine or triethylamine, preferably N, N-diisopropylethylamine; the solvent used is selected from dichloromethane, tetrahydrofuran, 1, 4-dioxane, ethyl acetate, acetone, N-dimethylformamide, preferably N, N-dimethylformamide.

Wherein R is ₁ 、R ₂ 、R ₃ X, Y are as defined above, R _c Methyl or ethyl;

and (3) salifying the corresponding acid with the compound I prepared by the method to obtain the pharmaceutically acceptable salt of the compound I.

The pharmaceutical composition comprises the anilinopyrimidino heterocyclic compound and a pharmaceutically acceptable carrier, and is prepared into common pharmaceutical preparations such as tablets, capsules, syrup, suspending agents or injection by adding common pharmaceutical auxiliary materials such as perfume, sweetener, liquid/solid filler, diluent and the like.

The anilinopyrimidino heterocyclic compound and the pharmaceutical composition thereof are applied to preparing WEE1 and HDAC double-target inhibitor drugs, and are particularly applied to preparing antitumor drugs.

The beneficial effects are that: compared with the prior art, the invention has the following remarkable advantages:

the compound can effectively inhibit WEE1 and HDAC, the inhibition rate value is optimally more than 90 percent, the proliferation of tumor cells can be effectively inhibited, and the inhibition rate is more than 50 percent; the application is wide, the drug effect can be exerted on the molecular level and the cellular level, and the nanomolar concentration level can be optimally achieved; the preparation method is easy to expand the structure and has good universality.

Detailed Description

The technical scheme of the invention is further described below by referring to examples.

Example 1: preparation of N ¹ - (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d) ]Pyrimidin-6-yl) amino) phenyl) -N ⁸ Hydroxyoctanediamide (Compound I-1)

Step 1: synthesis of intermediate 3

To a eggplant-shaped bottle was added starting material 1 (1.66 g,7.97 mmol), 2 (1.50 g,7.97 mmol), HATU (3.64 g,9.56 mmol), DIEA (1.54 g,11.95 mmol) and DCM (20 mL) dissolved and stirred at room temperature for 3 hours, TLC (DCM: meoh=35:1) monitored completion. The reaction solution was diluted with water, extracted three times with DCM, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, suction filtered, and the filtrate concentrated under reduced pressure to give 2.84g of a white solid as intermediate 3 in 94.2% yield by silica gel column chromatography. ¹ H NMR(400MHz,Chloroform-d)δ7.45(d,J＝8.9Hz,2H),7.32(d,J＝8.9Hz,2H),3.69(s,3H),2.39-2.29(m,4H),1.74(p,J＝7.3Hz,2H),1.65(p,J＝7.4Hz,2H),1.53(s,9H),1.46-1.33(m,4H).

Step 2: synthesis of intermediate 4 to a eggplant-shaped bottle were added intermediate 3 (2.80 g,7.40 mmol), TFA (19 mL) and DCM (19 mL), and the mixture was stirred at room temperature for 12 hours, and TLC (dichloromethane: methanol=20:1) monitored for completion of the reaction. The reaction solution was concentrated under reduced pressure, quenched with saturated sodium bicarbonate, extracted 3 times with ethyl acetate, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered off with suction, and the filtrate was concentrated under reduced pressure to give 1.91g of a pink solid as intermediate 4 in 92.7% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.49-9.94(m,3H),7.70(d,J＝8.9Hz,2H),7.30(d,J＝8.8Hz,2H),3.58(s,3H),2.37-2.24(m,4H),1.64-1.45(m,4H),1.35-1.23(m,4H).

Step 3: synthesis of intermediate 7

To a eggplant-shaped bottle was added starting material 5 (25.74 g,110.60 mmol), 6 (20.00 g,116.14 mmol), DIEA (73.00 mL,442.40 mmol) and THF (300 mL), and the reaction was refluxed at 75 ℃ for 12 hours, and monitored by TLC (petroleum ether: ethyl acetate=10:1) for completion. The reaction solution is concentrated under reduced pressure, diethyl ether is added for pulping, suction filtration is carried out, and the filtrate is concentrated under reduced pressure. Trifluoroacetic acid (100 mL) was added, stirred at room temperature for 1 hour, stirred at 70 ℃ for 1 hour, concentrated under reduced pressure, sodium hydroxide (6 m,200 mL) and ethanol (100 mL) were added, stirred at room temperature for 15 minutes, monitored by TLC (dichloromethane: methanol=20:1) for completion of the reaction, adjusted to acidity by adding concentrated hydrochloric acid, and concentrated under reduced pressure. The mixture of DCM and MeOH was added for dissolution, suction filtration and concentration of the filtrate under reduced pressure, and silica gel column chromatography gave 14.60g of yellow solid as intermediate 7 in 59.4% yield. ¹ H NMR(400MHz,DMSO-d ₆ )δ8.18(s,1H),5.86(ddt,J＝16.1,10.5,5.4Hz,1H),5.07-4.95(m,2H),4.32(d,J＝5.4Hz,2H),2.36(s,3H).

Step 4: synthesis of intermediate 9

To a three-necked flask were added starting material 8 (25.00 g,115.72 mmol) and diethyl ether (200 mL), replaced with nitrogen, and methyl magnesium iodide (3M diethyl ether solution, 100 mL) was added by syringe at 0deg.C, slowly warmed to room temperature, and TLC (petroleum ether: ethyl acetate=10:1) monitored for completion of the reaction. The reaction was quenched with 1M hydrochloric acid, extracted three times with EA, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered off with suction, and the filtrate was concentrated under reduced pressure to give 25.00g of a yellow oil in 100% yield. ¹ H NMR(300MHz,Chloroform-d)δ7.57(t,J＝7.7Hz,1H),7.41-7.38(m,1H),7.37(dd,J＝4.2,0.9Hz,1H),3.98(s,1H),1.55(s,6H).

Step 5: synthesis of intermediate 10

To a three-necked flask was added intermediate 7 (14.60 g,65.69 mmol), 9 (14.19 g,65.69 mmol), cuprous iodide (13.76 g,72.25 mmol), potassium carbonate (13.98 g,101.15 mmol) and 1, 4-dioxane (120 mL), replaced with argon, and trans-N, N' -dimethyl-1, 2-cyclohexanediamine (10.28 g,72.25 mmol) was added by syringe and reacted at 85℃for 16 hours, and TLC (dichloromethane: methane=20:1) was monitored for completion. The reaction solution was cooled to room temperature, quenched with ammonia water, extracted 3 times with EA, the organic layers combined and saturated sodium chloride dissolvedWashing the solution, drying with anhydrous sodium sulfate, suction filtering, concentrating the filtrate under reduced pressure, and performing silica gel column chromatography to obtain 12.30g of white solid which is intermediate 10 with a yield of 52.4%. ¹ HNMR(300MHz,Chloroform-d)δ8.96(s,1H),7.93(t,J＝7.9Hz,1H),7.78(d,J＝8.1Hz,1H),7.42(d,J＝6.8Hz,1H),5.71(ddt,J＝16.5,10.2,6.2Hz,1H),5.07(d,J＝10.2Hz,1H),4.94(d,J＝17.1Hz,1H),4.82(d,J＝6.2Hz,2H),3.84(s,1H),2.60(s,3H),1.60(s,6H).

Step 6: synthesis of intermediate 11

To a eggplant-shaped bottle was added intermediate 10 (600 mg,1.68 mmol), mCPBA (1.04 g,6.04 mmol) and toluene (10 mL), reacted at room temperature for 1 hour, TLC (petroleum ether: ethyl acetate=1:1) monitored complete reaction, intermediate 4 (218 mg,2.01 mmol), DIEA (2.00 mL,11.75 mmol) was added, reacted at room temperature for 2 hours, TLC (dichloromethane: methanol=25:1) monitored complete reaction. The reaction solution was quenched with saturated sodium bicarbonate solution, extracted with EA 3 times, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered off with suction, the filtrate was concentrated under reduced pressure, and chromatographed on silica gel to give 366mg of yellow solid as intermediate 11 in 37.1% yield. ¹ H NMR(300MHz,Chloroform-d)δ8.85(s,1H),7.92(t,J＝7.9Hz,1H),7.75(d,J＝8.0Hz,1H),7.61-7.51(m,4H),7.47(s,1H),7.38(d,J＝7.6Hz,1H),5.72(ddt,J＝16.5,10.2,6.1Hz,1H),5.06(d,J＝10.2Hz,1H),4.94(d,J＝18.3Hz,1H),4.77(d,J＝6.2Hz,2H),3.69(s,3H),2.43-2.28(m,4H),1.83-1.62(m,4H),1.45-1.37(m,4H).

Step 7: synthesis of target Compound I-1 hydroxylamine hydrochloride (710 mg,10.21 mmol), methanol (6 mL), potassium hydroxide (860 mg,15.31 mmol), were added to a eggplant-shaped bottle, reacted at 40℃for 10 minutes, suction filtered, and the filtrate was added to intermediate 11 (300 mg,0.51 mmol), reacted at room temperature for 1 hour, and TLC (dichloromethane: methanol=10:1) monitored to complete the reaction. To the reaction solution was added 1M hydrochloric acid to adjust pH to neutral, extracted 3 times with DCM, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, suction filtered, the filtrate concentrated under reduced pressure, and PTLC gave 62mg of the target compound I-1 as a yellow solid in 20.6% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.44(s,1H),10.19(s,1H),9.96(s,1H),8.87(s,1H),7.99(t,J＝7.9Hz,1H),7.75(d,J＝7.9Hz,1H),7.69-7.61(m,3H),7.56(d,J＝8.8Hz,2H),5.67(ddt,J＝16.4,10.2,5.9Hz,1H),5.39(s,1H),5.00(d,J＝10.2Hz,1H),4.82(d,J＝17.1Hz,1H),4.68(d,J＝5.9Hz,2H),2.30(t,J＝7.3Hz,2H),1.96(t,J＝7.3Hz,2H),1.64-1.40(m,10H),1.36-1.21(m,4H).

Example 2: preparation of 2- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) -N-hydroxyacetamide (I-2)

Step 1: synthesis of intermediate 13

Referring to the procedure of example 1, starting from intermediate 10 (600 mg,1.68 mmol) and 12 (277 mg,1.68 mmol), intermediate 13 (264 mg) was produced in 33.1% yield. ¹ H NMR(400MHz,Chloroform-d)δ8.86(s,1H),7.90(t,J＝7.9Hz,1H),7.77(d,J＝8.1Hz,1H),7.59(d,J＝8.6Hz,2H),7.40(d,J＝7.6Hz,1H),7.27(d,J＝8.4Hz,2H),5.71(ddt,J＝16.6,10.2,6.2Hz,1H),5.04(d,J＝10.1Hz,1H),4.93(d,J＝17.1Hz,1H),4.77(d,J＝6.2Hz,2H),3.71(s,3H),3.63(s,2H),1.60(s,6H).

Step 2: synthesis of target Compound I-2

Referring to the procedure of example 1, starting from intermediate 13 (264 mg,0.56 mmol), compound I-2 (102 mg) was produced in the yield: 38.6%. ¹ H NMR(400MHz,DMSO-d ₆ +D ₂ O)δ8.82(s,1H),8.00(t,J＝7.9Hz,1H),7.67(d,J＝8.0Hz,1H),7.62-7.53(m,3H),7.20(d,J＝8.4Hz,2H),5.61(ddt,J＝16.3,10.1,5.7Hz,1H),4.98(d,J＝11.2Hz,1H),4.77(d,J＝17.0Hz,1H),4.64(d,J＝5.8Hz,2H),3.25(s,2H),1.43(s,6H).

Example 3: preparation of 3- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) -N-hydroxypropionamide (I-3)

Step 1: synthesis of intermediate 15

To a eggplant-shaped bottle was added starting material 14 (2.70 g,16.42 mmol), tetrabutylammonium nitrate (10.00 g,32.84 mmol) and DCM (40 mL), -triflic anhydride (4.63 g,16.41 mmol) was slowly added at 30℃and the reaction was incubated for 30 min and monitored for completion by TLC (Petroleum ether: ethyl acetate=10:1). Sodium bicarbonate was added to the reaction solution, suction filtration was performed, the filtrate was concentrated under reduced pressure, and 940mg of yellow solid was obtained by silica gel column chromatography as intermediate 15, with a yield of 27.4%. ¹ H NMR(300MHz,Chloroform-d)δ8.17(d,J＝8.7Hz,2H),7.39(d,J＝8.7Hz,2H),3.69(s,3H),3.08(t,J＝7.5Hz,2H),2.70(t,J＝7.5Hz,2H).

Step 2: synthesis of intermediate 16

To a eggplant-shaped bottle was added intermediate 15 (940 mg,4.49 mmol), pd/C (100 mg), methanol (10 mL) and ethyl acetate (1 mL), and the reaction was allowed to proceed at room temperature for 12 hours, followed by TLC (Petroleum ether: ethyl acetate=10:1) to completion. The reaction solution was suction-filtered through celite, the filtrate was concentrated under reduced pressure, and 572mg of yellow solid was obtained by silica gel column chromatography as intermediate 16 in 71.0% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ6.98(d,J＝8.3Hz,2H),6.62(d,J＝8.3Hz,2H),3.66(s,3H),3.58(s,2H),2.84(t,J＝7.5Hz,2H),2.57(t,J＝7.5Hz,2H).

Step 3: synthesis of intermediate 17

Referring to the procedure of example 1, starting from intermediate 10 (600 mg,1.68 mmol) and 16 (333 mg,2.01 mmol), intermediate 17 (242 mg) was prepared in the yield: 29.5%. ¹ H NMR(400MHz,Chloroform-d)δ8.87(s,1H),7.90(d,J＝11.7Hz,1H),7.77(d,J＝12.0Hz,1H),7.55(d,J＝4.3Hz,2H),7.39(s,1H),7.28(s,1H),7.21(d,J＝12.3Hz,2H),5.77-5.66(m,1H),5.07(d,J＝10.3Hz,1H),4.96(d,J＝17.0Hz,1H),4.78(s,2H),3.70(s,3H),2.97(t,J＝9.5Hz,2H),2.66(t,J＝9.7Hz,2H),1.61(s,6H).

Step 4: synthesis of target Compound I-3

Referring to the procedure of example 1, starting from intermediate 17 (242 mg, mmol), the title compound I-3 (152 mg) was obtained in the yield: 62.8%. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.39(s,1H),10.23(s,1H),8.88(s,1H),8.72(s,1H),8.05(t,J＝7.9Hz,1H),7.76(d,J＝8.0Hz,1H),7.67-7.56(m,3H),7.16(d,J＝8.1Hz,2H),5.67(ddt,J＝16.4,10.2,5.9Hz,1H),5.35(s,1H),5.00(d,J＝8.7Hz,1H),4.83(d,J＝17.1Hz,1H),4.69(d,J＝5.8Hz,2H),2.79(t,J＝7.6Hz,2H),2.26(t,J＝7.6Hz,2H),1.47(s,6H).

Example 4: preparation of 2- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyacetamide (I-4)

Step 1: synthesis of intermediate 20

To a eggplant-shaped bottle was added intermediate 18 (5.00 g,23.90 mmol), 19 (3.66 g,23.90 mmol), cesium carbonate (9.34 g,28.68 mmol) and DMF (50 mL), stirred at room temperature for 12 hours, and TLC (dichloromethane: methanol=35:1) monitored for reaction completion. 200mL of water was added to the reaction mixture, suction filtration and infrared drying of the cake gave 6.05g of a white solid as intermediate 20 in 90.0% yield. ¹ H NMR(400MHz,Chloroform-d)δ7.28(d,J＝9.0Hz,2H),6.88(d,J＝9.0Hz,2H),4.62(s,2H),3.82(s,3H),1.53(s,9H).

Step 2: synthesis of intermediate 21

To the eggplant-shaped flask was added 20 (6.00 g,21.33 mmol), trifluoroacetic acid (20 mL) and dichloromethane (20 mL), and the mixture was stirred at room temperature for 3 hours, and TLC (dichloromethane: methanol=35:1) monitored for completion of the reaction. The reaction solution was concentrated under reduced pressure, saturated sodium bicarbonate solution was added, EA was extracted 3 times, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, suction filtered, and the filtrate was concentrated under reduced pressure to give 3.81g of a yellow liquid as intermediate 21 in 98.6% yield. ¹ H NMR(400MHz,Chloroform-d)δ6.79(d,J＝8.8Hz,2H),6.66(d,J＝8.9Hz,2H),4.58(s,2H),3.82(s,3H),3.23(s,2H).

Step 3: synthesis of intermediate 22

Referring to the procedure of example 1, starting from intermediate 10 (600 mg,1.68 mmol) and 21 (365 mg,2.01 mmol), intermediate 22 (477 mg) was produced in 57.9% yield. ¹ H NMR(300MHz,Chloroform-d)δ8.85(s,1H),7.87(t,J＝7.9Hz,1H),7.74(d,J＝7.2Hz,1H),7.52(d,J＝9.0Hz,2H),7.37(d,J＝8.4Hz,1H),6.93(d,J＝9.0Hz,2H),5.71(ddt,J＝16.5,10.1,6.1Hz,1H),5.05(d,J＝10.2Hz,1H),4.95(d,J＝17.0Hz,1H),4.76(d,J＝6.5Hz,2H),4.67(s,2H),4.04(s,1H),3.84(s,3H),1.60(s,6H).

Step 4: synthesis of target Compound I-4

Referring to the procedure of example 1, starting from intermediate 22 (463 mg,0.95 mmol), the title compound I-4 (192 mg) was obtained in the yield: 41.1%. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.94(s,1H),10.24(s,1H),9.03(s,1H),8.86(s,1H),8.05(t,J＝7.9Hz,1H),7.75(d,J＝8.1Hz,1H),7.67-7.57(m,3H),6.96(d,J＝8.7Hz,2H),5.67(ddt,J＝16.5,10.2,5.9Hz,1H),5.38(s,1H),4.99(d,J＝10.3Hz,1H),4.82(d,J＝15.6Hz,1H),4.69(d,J＝6.0Hz,2H),4.47(s,2H),1.47(s,6H).

Example 5: preparation of 3- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxypropionamide (I-5)

Step 1: synthesis of intermediate 24

To a eggplant-shaped bottle was added starting material 23 (8.05 g,52.62 mmol), 19 (7.32 g,52.62 mmol), KOH (7.38 g,131.55 mmol) and water (73 mL), and the reaction was refluxed for 12 hours at 105℃and monitored by TLC (dichloromethane: methanol=10:1) for completion. The reaction solution was cooled to room temperature, pH was adjusted to acidity by adding 6M hydrochloric acid, EA was extracted 3 times, the organic layers were combined, saturated sodium bicarbonate solution was extracted 3 times, the aqueous layer was combined, pH was adjusted to acidity by adding 6M hydrochloric acid, suction filtration was performed, and the filter cake was dried in vacuo to give 2.74g of a white solid as intermediate 24 in 24.7% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ12.49(s,1H),8.21(d,J＝9.2Hz,2H),7.16(d,J＝9.2Hz,2H),4.32(t,J＝5.9Hz,2H),2.76(t,J＝6.0Hz,2H).

Step 2: synthesis of intermediate 25

Into a three-necked flask was charged intermediate 24 (2.74 g,12.98 mmol), anhydrous methanol (30 mL), -4deg.CAcetyl chloride (0.51 g,6.49 mmol) was added by syringe and the reaction was incubated for 10 hours, monitored by TLC (dichloromethane: methanol=10:1) for completion, suction filtration and vacuum drying of the filter cake to give 2.62g of grey solid as intermediate 25 in 89.7% yield. ¹ H NMR(300MHz,Chloroform-d)δ8.22(d,J＝9.2Hz,2H),6.99(d,J＝9.3Hz,2H),4.37(t,J＝6.3Hz,2H),3.77(s,3H),2.88(t,J＝6.3Hz,2H).

Step 3: synthesis of intermediate 26

Referring to example 3, intermediate 26 (2.24 g) was prepared in 99.3% yield starting from intermediate 25 (2.6 g,11.55 mmol). ¹ HNMR(300MHz,DMSO-d ₆ )δ6.64(d,J＝8.8Hz,2H),6.49(d,J＝8.8Hz,2H),4.64(s,2H),4.04(t,J＝6.0Hz,2H),3.63(s,3H),2.71(t,J＝6.0Hz,2H).

Step 4: synthesis of target Compound I-5

The procedure of example 1 was followed, starting from intermediates 26 and 10, to give the desired compound I-5. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.57(s,1H),10.20(s,1H),8.87(s,1H),8.85(s,1H),8.05(t,J＝8.1Hz,1H),7.75(d,J＝8.0Hz,1H),7.68-7.58(m,3H),6.92(d,J＝8.8Hz,2H),5.67(ddt,J＝16.5,10.2,6.0Hz,1H),5.34(s,1H),5.00(d,J＝10.3Hz,1H),4.83(d,J＝18.6Hz,1H),4.69(d,J＝5.9Hz,2H),4.18(t,J＝6.0Hz,2H),2.44(t,J＝6.0Hz,2H),1.47(s,6H).

Example 6: preparation of 4- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxybutyramide (I-6)

The procedure of example 4 was followed, starting from methyl 4-bromobutyrate as intermediate, to give the desired compound 6. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.45(s,1H),10.25(s,1H),8.85(s,1H),8.75(s,1H),8.05(t,J＝7.9Hz,1H),7.75(d,J＝8.0Hz,1H),7.68-7.57(m,3H),6.92(d,J＝9.0Hz,2H),5.67(ddt,J＝16.4,10.2,5.9Hz,1H),5.36(s,1H),5.00(d,J＝10.3Hz,1H),4.82(d,J＝17.1Hz,1H),4.69(d,J＝6.0Hz,2H),3.95(t,J＝6.3Hz,2H),2.14(t,J＝7.4Hz,2H),1.93(p,J＝6.7Hz,2H),1.46(s,6H).

Example 7: preparation of 5- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxypentanamide (I-7)

Referring to the procedure of example 4, compound I-7 was prepared starting from methyl 5-bromopentanoate as an intermediate. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.41(s,1H),10.22(s,1H),8.85(s,1H),8.73(s,1H),8.06(t,J＝7.9Hz,1H),7.75(d,J＝8.0Hz,1H),7.68-7.55(m,3H),6.92(d,J＝9.0Hz,2H),5.67(ddt,J＝16.1,10.2,5.9Hz,1H),5.36(s,1H),5.00(d,J＝11.7Hz,1H),4.82(d,J＝18.7Hz,1H),4.69(d,J＝6.0Hz,2H),3.94(d,J＝6.0Hz,2H),1.76-1.59(m,4H),1.46(s,6H).

Example 8: preparation of 6- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxycaproamide (I-8)

Referring to the procedure of example 4, compound I-8 was prepared starting from methyl 6-bromohexanoate. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.39(s,1H),10.22(s,1H),8.85(s,1H),8.71(s,1H),8.05(t,J＝7.8Hz,1H),7.74(d,J＝8.2Hz,1H),7.66-7.56(m,3H),6.92(d,J＝9.1Hz,2H),5.76-5.57(m,1H),5.36(s,1H),4.99(d,J＝10.3Hz,1H),4.82(d,J＝17.1Hz,1H),4.69(d,J＝5.9Hz,2H),3.94(t,J＝6.3Hz,2H),1.98(t,J＝7.2Hz,2H),1.69(p,J＝6.9Hz,2H),1.55(p,J＝7.0Hz,2H),1.46(s,6H),1.38(p,J＝7.0Hz,2H).

Example 9: preparation of 7- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyheptanamide (I-9)

Referring to the procedure of example 4, compound I-9 was prepared starting from methyl 7-bromoheptanoate. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.34(s,1H),10.22(s,1H),8.85(s,1H),8.66(s,1H),8.05(t,J＝7.7Hz,1H),7.74(d,J＝8.0Hz,1H),7.65-7.58(m,3H),6.92(d,J＝9.1Hz,2H),5.67(ddt,J＝16.2,10.2,6.0Hz,1H),5.33(s,1H),5.00(d,J＝10.3Hz,1H),4.83(d,J＝18.6Hz,1H),4.69(d,J＝6.0Hz,2H),3.94(t,J＝6.4Hz,2H),1.96(p,J＝6.7Hz,2H),1.70(p,J＝6.7Hz,2H),1.58-1.37(m,10H),1.35-1.29(m,2H).

Example 10: preparation of 8- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-10)

Referring to the procedure of example 4, compound I-10 was prepared starting from ethyl 8-bromooctanoate. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.36(s,1H),10.23(s,1H),8.85(s,1H),8.68(s,1H),8.05(t,J＝8.1Hz,1H),7.75(d,J＝7.7Hz,1H),7.67-7.55(m,3H),6.91(d,J＝8.5Hz,2H),5.67(ddt,J＝16.3,10.2,5.9Hz,1H),5.34(s,1H),4.99(d,J＝10.1Hz,1H),4.83(d,J＝15.6Hz,1H),4.69(d,J＝5.8Hz,2H),3.94(t,J＝6.4Hz,2H),1.96(t,J＝7.3Hz,2H),1.70(p,J＝6.7Hz,2H),1.58-1.22(m,14H).

Example 11: preparation of 2- (4- (4- (2-allyl-1- (6- (2-hydroxypropyl-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyacetamide (I-11)

Step 1: synthesis of intermediate 29

To a eggplant-shaped bottle was added starting material 18 (4.43 g,28.95 mmol), 28 (5 g,24.13 mmol), DIEA (7.80 g,60.32 mmol) and DMF (50 mL), stirred overnight at room temperature, monitored by TLC (dichloromethane: methanol=15:1) for complete reaction, 200mL of water was added, suction filtered, and the filter cake infrared dried to give 6.04g of yellow solid as intermediate 29 in 89.6% yield. ¹ H NMR(400MHz,Chloroform-d)δ8.14(d,J＝9.4Hz,2H),6.84(d,J＝9.4Hz,2H),3.76(s,3H),3.53-3.46(m,4H),3.32(s,2H),2.79-2.72(m,4H).

Step 2: synthesis of intermediate 30

To a eggplant-shaped bottle was added intermediate 29 (6.00 g,24.07 mmol), pd/C (300 mg), methanol (50 mL), hydrogen substitution, and reaction at room temperature for 12 hours, and TLC (dichloromethane: methanol=15:1) monitored for completion of the reaction. The reaction solution was filtered through celite, concentrated under reduced pressure, and purified by silica gel column chromatography to give 5.24g of brown solid as intermediate 30 in 87.3% yield. ¹ H NMR(300MHz,Chloroform-d)δ6.82(d,J＝8.8Hz,2H),6.65(d,J＝8.8Hz,2H),3.75(s,3H),3.29(s,2H),3.16-3.03(m,4H),2.79-2.70(m,4H).

Step 3: synthesis of target Compound I-11

The procedure of example 1 was followed, starting from intermediates 30 and 10, to give the desired compound I-11. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.48(s,1H),10.12(s,1H),8.83(s,1H),8.81(s,1H),8.05(t,J＝7.9Hz,1H),7.75(d,J＝8.1Hz,1H),7.66-7.49(m,3H),6.93(d,J＝9.0Hz,2H),5.67(ddt,J＝16.4,10.2,6.0Hz,1H),5.34(s,1H),5.00(d,J＝10.2Hz,1H),4.83(d,J＝17.0Hz,1H),4.68(d,J＝6.0Hz,2H),3.12(t,J＝4.9Hz,4H),2.95(s,2H),2.60(t,J＝4.3Hz,4H),1.47(s,6H).

Example 12: preparation of 3- (4- (4- ((2-allyl-1- (6- (2-hydroxy-prop-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxypropionamide (I-12)

The procedure of example 11 was followed, starting from methyl 3-bromopropionate, to obtain the objective compound I-12。 ¹ H NMR(400MHz,DMSO-d ₆ )δ10.43(s,1H),10.15(s,1H),8.83(s,1H),8.77(s,1H),8.05(t,J＝8.2Hz,1H),7.76(d,J＝7.9Hz,1H),7.66-7.49(m,3H),6.92(d,J＝8.7Hz,2H),5.67(ddt,J＝16.4,10.2,6.0Hz,1H),5.33(s,1H),4.99(d,J＝10.2Hz,1H),4.83(d,J＝17.1Hz,1H),4.69(d,J＝6.0Hz,2H),3.09(t,J＝5.0Hz,4H),2.58(t,J＝7.0Hz,2H),2.54-2.47(m,4H),2.18(t,J＝7.0Hz,2H),1.47(s,6H).

Example 13: preparation of 4- (4- (4- (2-allyl-1- (6- (2-hydroxypropyl-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxybutyramide (I-13)

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The procedure of example 11 was followed, starting from methyl 4-bromobutyrate, to give the title compound I-13. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.28(s,2H),8.88(s,1H),8.06(t,J＝7.8Hz,1H),7.76(d,J＝8.0Hz,1H),7.65-7.46(m,3H),6.92(d,J＝8.6Hz,2H),5.67(ddt,J＝16.4,10.1,5.9Hz,1H),5.35(s,1H),4.99(d,J＝11.7Hz,1H),4.83(d,J＝16.6Hz,1H),4.69(d,J＝6.0Hz,2H),3.19-2.99(m,4H),2.49-2.43(m,4H),2.30(t,J＝7.1Hz,2H),2.00(t,J＝7.2Hz,2H),1.69(p,J＝7.8Hz,2H),1.47(s,6H).

Example 14: preparation of 5- (4- (4- (4- (2-allyl-1- (6- (2-hydroxypropyl-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxypentanamide (I-14)

The procedure of example 11 was followed, starting from methyl 5-bromopentanoate, to give the title compound I-14. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.37(s,1H),10.15(s,1H),8.83(s,1H),8.68(s,1H),8.05(t,J＝6.3Hz,1H),7.75(d,J＝8.1Hz,1H),7.67-7.50(m,3H),6.93(d,J＝8.7Hz,2H),5.67(ddt,J＝16.5,10.2,5.9Hz,1H),5.00(d,J＝8.7Hz,1H),4.83(d,J＝18.7Hz,1H),4.68(d,J＝6.0Hz,2H),3.18-3.03(m,4H),2.58-2.52(m,4H),2.38-2.27(m,2H),1.98(t,J＝7.0Hz,2H),1.60-1.39(m,10H).

Example 15: preparation of 6- (4- (4- (2-allyl-1- (6- (2-hydroxypropyl-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxycaproamide (I-15)

The procedure of example 11 was followed, starting from methyl 6-bromohexanoate, to give the title compound I-15. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.38(s,1H),10.17(s,1H),8.83(s,1H),8.71(s,1H),8.04(t,J＝8.7Hz,1H),7.76(d,J＝7.8Hz,1H),7.67-7.46(m,3H),6.92(d,J＝8.5Hz,2H),5.67(ddt,J＝16.4,11.0,5.9Hz,1H),5.36(s,1H),4.99(d,J＝10.1Hz,2H),4.82(d,J＝17.1Hz,1H),4.69(d,J＝6.0Hz,2H),3.57-3.26(m,4H),3.19-2.97(m,4H),2.31(t,J＝8.4Hz,2H),1.96(t,J＝7.0Hz,2H),1.64-1.37(m,10H),1.25(p,J＝8.1Hz,2H).

Example 16: preparation of 7- (4- (4- (2-allyl-1- (6- (2-hydroxypropyl-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyheptanamide (I-16)

The procedure of example 11 was followed, starting from methyl 7-bromoheptanoate, to give the objective compound I-16. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.43(s,1H),10.19(s,1H),8.84(s,1H),8.05(t,J＝7.9Hz,1H),7.76(d,J＝7.9Hz,1H),7.64-7.51(m,3H),6.90(d,J＝8.6Hz,2H),5.67(ddt,J＝16.5,10.1,6.0Hz,1H),5.38(s,1H),4.99(d,J＝9.5Hz,1H),4.82(d,J＝18.7Hz,1H),4.70(d,J＝6.0Hz,2H),3.15-2.98(m,4H),2.56-2.46(m,4H),2.30(t,J＝6.6Hz,2H),1.96(t,J＝7.3Hz,2H),1.56-1.35(m,10H),1.32-1.15(m,4H).

Example 17: preparation of 8- (4- (4- (4- (2-allyl-1- (6- (2-hydroxypropyl-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-17)

The procedure of example 11 was followed, starting from methyl 8-bromooctanoate, to give the desired compound I-17. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.43(s,1H),10.19(s,1H),8.83(s,1H),8.07(t,J＝13.5Hz,1H),7.76(d,J＝7.4Hz,1H),7.67-7.47(m,3H),6.90(d,J＝8.5Hz,2H),5.67(ddt,J＝16.4,10.2,5.9Hz,1H),5.38(s,1H),4.99(d,J＝10.1Hz,1H),4.82(d,J＝18.0Hz,1H),4.70(d,J＝5.7Hz,2H),3.13-2.98(m,4H),2.50-2.40(m,4H),2.27(t,J＝7.3Hz,2H),1.95(t,J＝7.3Hz,2H),1.56-1.34(m,10H),1.31-1.15(m,6H).

Example 18: preparation of 8- (4- (2-allyl-1- (3- (2-hydroxypropyl-2-yl) phenyl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-18)

Step 1: referring to the procedure of example 4, intermediate 37 was prepared starting from intermediates 35 and 19. ¹ H NMR(300MHz,DMSO-d ₆ )δ6.67(d,J＝8.9Hz,2H),6.58(d,J＝8.9Hz,2H),4.04(q,J＝7.1Hz,2H),3.80(t,J＝6.4Hz,2H),2.27(t,J＝7.4Hz,2H),1.63(p,J＝6.6Hz,1H),1.52(p,J＝7.5Hz,1H),1.43-1.22(m,6H),1.17(t,J＝7.1Hz,3H).

Step 2: referring to the method of example 1, intermediate 34 was prepared starting from intermediate 32. ¹ H NMR(400MHz,Chloroform-d)δ8.92(s,1H),7.58(t,J＝1.9Hz,1H),7.54(t,J＝1.6Hz,1H),7.50(t,J＝7.7Hz,1H),7.30(dd,J＝2.2,1.3Hz,1H),5.77-5.66(m,1H),5.14(dq,J＝10.1,1.1Hz,1H),4.99(dq,J＝17.0,1.3Hz,1H),4.46(dt,J＝6.0,1.4Hz,2H),2.52(s,3H),1.64(s,6H).

Step 3: with reference to the method of example 1,starting from intermediates 34 and 37, the target compound I-18 was prepared. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.37(s,1H),10.14(s,1H),8.83(s,1H),8.70(s,1H),7.74-7.42(m,5H),7.31(s,1H),6.85(d,J＝8.9Hz,2H),5.68(ddt,J＝16.1,10.7,5.6Hz,1H),5.23(s,1H),5.08(d,J＝11.9Hz,1H),4.91(d,J＝17.1Hz,1H),4.23(s,2H),3.90(t,J＝6.4Hz,2H),1.95(t,J＝7.3Hz,2H),1.67(p,J＝6.9Hz,2H),1.56-1.17(m,14H).

Example 19: preparation of 8- (4- (4- (2-allyl-1- (3- (2-hydroxypropyl-2-yl) phenyl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-19)

Referring to the method of example 11, intermediate 40 was prepared starting from intermediates 35 and 28. ¹ H NMR(300MHz,Chloroform-d)δ6.83(d,J＝8.8Hz,2H),6.66(d,J＝8.7Hz,2H),4.14(q,J＝7.1Hz,2H),3.46(s,2H),3.10(t,J＝5.0Hz,4H),2.64(t,J＝5.0Hz,4H),2.41(t,J＝8.1Hz,2H),2.30(t,J＝7.5Hz,1H),1.72-1.48(m,4H),1.41-1.21(m,12H).

The procedure of example 1 was followed, starting from intermediates 34 and 40, to give the desired compound I-19. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.35(s,1H),10.10(s,1H),8.81(s,1H),8.71(s,1H),7.68-7.42(m,5H),7.35-7.25(m,1H),6.86(d,J＝9.1Hz,2H),5.68(ddt,J＝16.2,10.7,5.6Hz,1H),5.22(s,1H),5.08(dd,J＝10.3,1.5Hz,1H),4.91(dd,J＝17.1,1.6Hz,1H),4.41-4.17(m,2H),3.06(t,J＝5.0Hz,4H),2.52(t,J＝5.0Hz,4H),2.33(t,J＝6.2Hz,2H),1.94(t,J＝7.3Hz,2H),1.55-1.18(m,16H).

Example 20: preparation of N-hydroxy-8- (4- (1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -2-isopropyl-3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) octanamide (I-20)

Step 1: synthesis of intermediate 42

To a eggplant-shaped bottle were added starting material 5 (10.00 g,42.98 mmol) and ethanol (200 mL), hydrazine hydrate (5.24 mL,86.96 mmol) was added at 0deg.C, the reaction was incubated for 1 hour, and TLC (Petroleum ether: ethyl acetate=5:1) monitored for completion. The reaction solution was concentrated under reduced pressure, and was slurried with methyl tert-butyl ether, suction filtered, and the filtrate was concentrated under reduced pressure to give 5.19g of a white solid as intermediate 42 in a yield of 52.9%. ¹ H NMR(300MHz,DMSO-d ₆ )δ9.03(s,1H),8.50(s,1H),4.80(s,2H),4.28(q,J＝7.1Hz,2H),2.53(s,3H),1.30(t,J＝7.1Hz,3H).

Step 2: synthesis of intermediate 43

To a eggplant-shaped flask was added intermediate 42 (5.00 g,21.92 mmol), acetone (100 mL), and the mixture was reacted at 70℃for 12 hours, concentrated under reduced pressure, dissolved in methanol (100 mL), sodium cyanoborohydride (2.07 g,32.89 mmol) and concentrated hydrochloric acid (2 mL) were added at 0℃and reacted at room temperature for 12 hours, and TLC (Petroleum ether: ethyl acetate=3:1) was monitored to complete the reaction. The reaction was quenched by adding saturated sodium bicarbonate solution, extracted 3 times with ethyl acetate, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure, and purified by silica gel column chromatography to give 4.15g of a white solid as intermediate 43 in 70.0% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ9.20(d,J＝6.2Hz,1H),8.53(s,1H),5.37-5.27(m,1H),4.29(q,J＝7.1Hz,2H),3.35(s,1H),3.28-3.15(m,1H),2.52(s,3H),1.30(t,J＝7.1Hz,3H),1.02(d,J＝6.2Hz,6H).

Step 3: synthesis of intermediate 44

To the eggplant-shaped flask was added intermediate 43 (4.10 g,15.17 mmol), sodium hydroxide solution (5 m,50 mL), methanol (25 mL), stirred at room temperature for 12 hours, and TLC (dichloromethane: methanol=3:1) monitored for reaction completion. Concentrated hydrochloric acid is added into the reaction solution to adjust the pH to be acidic, the mixture is concentrated under reduced pressure, the DCM and the MeOH mixed solvent are added for dissolution, the suction filtration and the concentration of the filtrate under reduced pressure are carried out, and 1.73g of white solid which is intermediate 44 is obtained through silica gel column chromatography, and the yield is 50.8%. ¹ H NMR(300MHz,DMSO-d ₆ )δ8.22(s,1H),4.57(hept,J＝6.6Hz,1H),2.39(s,3H),1.22(d,J＝6.7Hz,6H).

Step 4: synthesis of target Compound I-20

Referring to the procedure of example 1, compound I-20 was prepared starting from intermediate 44. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.42(s,1H),10.18(s,1H),8.85-8.58(m,2H),8.07(t,J＝6.9Hz,1H),7.75-7.57(m,4H),6.89(d,J＝8.4Hz,2H),5.37(s,1H),4.28-4.07(m,1H),3.91(t,J＝6.6Hz,2H),1.96(t,J＝6.9Hz,2H),1.67(p,J＝6.9Hz,2H),1.56-1.07(m,22H).

Example 21: preparation of N-hydroxy-8- (4- (1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -2-isopropyl-3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) octanamide (I-21)

Referring to the procedure of example 1, starting from intermediates 45 and 40, compound I-21 was prepared. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.37(s,1H),10.14(s,1H),8.85-8.63(m,2H),8.08(t,J＝7.8Hz,1H),7.72(d,J＝8.0Hz,1H),7.65(d,J＝7.7Hz,1H),7.56(d,J＝8.1Hz,2H),6.89(d,J＝8.8Hz,2H),5.35(s,1H),4.22-4.07(m,1H),3.08(t,J＝4.9Hz,4H),2.48(t,J＝4.9Hz,4H),2.30(t,J＝7.3Hz,2H),1.94(t,J＝7.3Hz,2H),1.56-1.12(m,22H).

Example 22: preparation of 8- (4- (2-allyl-1- (6- (2-methoxypropane-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-22)

Step 1: synthesis of intermediate 48

To a eggplant-shaped bottle were added intermediate 9 (5.50 g,25.45 mmol), methyl iodide (10.84 g,76.36 mmol) and anhydrous tetrahydrofuran (100 mL), sodium hydride (2.93 g,76.36 mmol) was added, and the reaction was stirred at room temperature for 12 hours, and monitored by TLC (Petroleum ether: ethyl acetate=5:1) for completion. Quenching with water, extracting with EA for 3 times, mixing the organic layers, washing with saturated sodium chloride solution, drying with anhydrous sodium sulfate, filtering, and filtering Concentrated under reduced pressure to give 5.42g of a yellow oil as intermediate 48 in 92.6% yield. ¹ H NMR(400MHz,Chloroform-d)δ7.61-7.49(m,2H),7.36(dd,J＝6.8,1.9Hz,1H),3.20(s,3H),1.55(s,6H).

Step 2: synthesis of target Compound I-22

The procedure of example 1 was followed, starting from intermediates 48 and 37, to give the desired compound I-22. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.38(s,1H),10.23(s,1H),8.86(s,1H),8.70(s,1H),8.08(t,J＝7.9Hz,1H),7.81(d,J＝7.9Hz,1H),7.62(d,J＝8.4Hz,2H),7.46(d,J＝7.5Hz,1H),6.92(d,J＝8.5Hz,2H),5.67(ddt,J＝16.4,10.1,5.9Hz,1H),4.99(dd,J＝10.3,1.5Hz,1H),4.82(dd,J＝17.0,1.5Hz,1H),4.71(d,J＝5.9Hz,2H),3.93(t,J＝6.4Hz,2H),3.09(s,3H),1.96(t,J＝7.3Hz,2H),1.68(p,J＝6.9Hz,2H),1.57-1.17(m,14H).

Example 23: preparation of 8- (4- (4- (2-allyl-1- (6- (2-methoxypropane-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-23)

The procedure of example 1 was followed starting from intermediates 49 and 40 to give the title compound I-23. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.38(s,1H),10.20(s,1H),8.84(s,1H),8.08(t,J＝7.9Hz,1H),7.82(d,J＝8.0Hz,1H),7.59(d,J＝8.4Hz,2H),7.46(d,J＝7.6Hz,1H),6.92(d,J＝8.7Hz,2H),5.67(ddt,J＝16.4,10.2,6.0Hz,1H),4.99(d,J＝10.2Hz,1H),4.82(d,J＝18.7Hz,1H),4.71(d,J＝6.0Hz,2H),3.19-3.09(m,4H),2.66-2.52(m,4H),2.33(t,J＝7.4Hz,2H),1.95(t,J＝7.3Hz,2H),1.60-1.36(m,10H),1.26(s,6H).

Example 24: preparation of N-hydroxy-8- (4- (1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2-phenyl-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) octanamide (I-24)

Step 1: synthesis of intermediate 51

To a eggplant-shaped bottle was added starting material 5 (5.00 g,21.49 mmol), phenylhydrazine (2.32 g,21.49 mmol), triethylamine (8.70 g,85.96 mmol) and tetrahydrofuran (100 mL), stirred overnight at room temperature, TLC (Petroleum ether: ethyl acetate=3:1) monitored the reaction was complete, diluted with water, EA extracted 3 times, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, suction filtered, the filtrate concentrated under reduced pressure, and a white solid 2.85g was obtained as intermediate 51 by silica gel column chromatography in 43.6% yield. ¹ H NMR(400MHz,Chloroform-d)δ9.65(s,1H),8.70(s,1H),7.28-7.21(m,2H),6.96-6.87(m,3H),6.35(s,1H),4.41(q,J＝7.1Hz,2H),2.29(s,3H),1.43(t,J＝7.1Hz,3H).

Step 2: synthesis of target Compound I-24

Referring to the procedure of example 20, compound I-24 was prepared starting from intermediate 51. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.44-10.29(m,2H),8.93(s,1H),8.69(s,1H),8.02(t,J＝7.8Hz,1H),7.88(s,1H),7.67(s,1H),7.45(d,J＝7.8Hz,1H),7.35(d,J＝4.3Hz,4H),7.22-7.14(m,1H),6.95(d,J＝8.5Hz,2H),5.08(s,1H),3.96(t,J＝6.4Hz,2H),1.96(t,J＝7.2Hz,2H),1.71(p,J＝7.2Hz,2H),1.51(p,J＝7.3Hz,2H),1.44-1.20(m,6H),0.98(s,6H).

Example 25: preparation of N-hydroxy-8- (4- (1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2-phenyl-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) octanamide (I-25)

The procedure of example 1 was followed, starting from intermediates 53 and 40, to give the desired compound I-25.LC/MS: m/z=680.37 [ m+h ]] ⁺ .

Example 26: preparation of 8- (4- (2-allyl-3-oxo-1- (pyridin-2-yl) -2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-26)

The procedure of example 1 was followed, starting from intermediates 56 and 7, to give the desired compound I-26. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.38(s,1H),10.24(s,1H),8.85(s,1H),8.70(s,1H),8.54(d,J＝4.6Hz,1H),8.09(s,1H),7.88(d,J＝8.1Hz,1H),7.61(s,2H),7.43-7.35(m,1H),6.91(d,J＝8.3Hz,2H),5.68(ddt,J＝16.4,10.2,5.9Hz,1H),5.01(d,J＝10.2Hz,1H),4.87(d,J＝18.8Hz,1H),4.60(s,2H),3.93(t,J＝6.5Hz,2H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.7Hz,2H),1.50(p,J＝7.3Hz,2H),1.43-1.20(m,6H).

Example 27: preparation of 8- (4- (4- (2-allyl-3-oxo-1- (pyridin-2-yl) -2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-27)

The procedure of example 1 was followed, starting from intermediates 57 and 40, to give the desired compound I-27. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.38(s,1H),10.20(s,1H),8.84(s,1H),8.54(d,J＝6.8Hz,1H),8.12-8.07(m,2H),7.89(d,J＝8.1Hz,2H),7.67-7.55(m,2H),7.43-7.35(m,1H),6.92(d,J＝6.1Hz,2H),5.75-5.61(m,1H),5.02(d,J＝10.2Hz,2H),4.87(d,J＝17.1Hz,1H),4.61(s,2H),3.11(s,4H),2.53(s,4H),2.33(t,J＝7.3Hz,2H),1.94(t,J＝7.3Hz,2H),1.55-1.40(m,4H),1.31-1.20(m,6H).

Example 28: preparation of 8- (4- (2-allyl-3-oxo-1- (6- (trifluoromethyl) pyridin-2-yl) -2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-28)

The procedure of example 1 was followed, starting from intermediates 60 and 7, to give the desired compound I-28.LC/MS: m/z=586.24 [ m+h ]] ⁺ .

Example 29: preparation of 8- (4- (4- (2-allyl-3-oxo-1- (6- (trifluoromethyl) pyridin-2-yl) -2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-29)

The procedure of example 1 was followed, starting from intermediates 61 and 40, to give the desired compound I-29.LC/MS: m/z=654.30 [ m+h] ⁺ .

Example 30: preparation of 8- (4- (2-allyl-1- (6-methoxypyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-30)

Referring to the procedure of example 1, starting from intermediates 64 and 7, compound I-30 was prepared. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.36(s,1H),10.23(s,1H),8.85(s,1H),8.68(s,1H),7.96(t,J＝8.0Hz,1H),7.62(s,2H),7.44(d,J＝6.8Hz,1H),6.90(d,J＝8.7Hz,2H),6.80(d,J＝8.1Hz,1H),5.70(ddt,J＝16.4,10.2,5.9Hz,1H),5.04(d,J＝11.9Hz,1H),4.93(d,J＝18.7Hz,1H),4.66(d,J＝5.7Hz,2H),3.93(t,J＝6.5Hz,2H),3.89(s,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.7Hz,2H),1.50(p,J＝7.2Hz,2H),1.42-1.20(m,8H).

Example 31: preparation of 8- (4- (4- (2-allyl-1- (6-methoxypyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-31)

According to the method of example 1, compound I-31 was obtained starting from the intermediate. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.36(s,1H),10.19(s,1H),8.83(s,1H),8.70(s,1H),7.98(s,1H),7.59(s,2H),7.45(d,J＝7.7Hz,1H),6.91(d,J＝8.8Hz,2H),6.80(d,J＝8.1Hz,1H),5.69(ddt,J＝16.4,10.3,5.9Hz,1H),5.04(dd,J＝10.3,1.5Hz,1H),4.93(dd,J＝17.2,1.6Hz,1H),4.66(d,J＝6.0Hz,2H),3.89(s,3H),3.09(s,4H),2.53(s,4H),2.31(t,J＝7.5Hz,2H),1.94(t,J＝7.3Hz,2H),1.54-1.38(m,4H),1.35-1.19(m,6H).

Example 32: preparation of 8- (4- (2-allyl-1- (6-ethoxypyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-32)

Step 1: synthesis of intermediate 69

Absolute ethanol (5.8 g,126.64 mmol) and tetrahydrofuran (50 mL) were added to a eggplant-shaped bottle, sodium hydride was added at 0 ℃, a tetrahydrofuran solution (50 mL) of the starting material 68 (10 g,42.21 mmol) was added dropwise, the reaction was monitored by TLC (petroleum ether: ethyl acetate=10:1) for 12 hours at room temperature, the reaction was complete, water quenching was added, EA was extracted 3 times, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, suction filtered, and the filtrate was concentrated under reduced pressure, and purified by silica gel column chromatography to give 7.11g of a transparent oil as intermediate 69 in 83.4% yield. ¹ H NMR(300MHz,Chloroform-d)δ7.42(t,J＝6.0Hz,1H),7.04(dd,J＝7.5,0.7Hz,1H),6.67(dd,J＝8.2,0.7Hz,1H),4.36(q,J＝7.1Hz,2H),1.39(t,J＝7.1Hz,3H).

Step 2: synthesis of target Compound I-32

Compound I-32 was prepared by the method of reference example 30 starting from intermediates 69 and 7. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.35(s,1H),10.22(s,1H),8.84(s,1H),8.68(s,1H),7.96(t,J＝8.0Hz,1H),7.63(d,J＝8.5Hz,2H),7.41(d,J＝7.7Hz,1H),6.90(d,J＝8.7Hz,2H),6.78(d,J＝8.1Hz,1H),5.69(ddt,J＝16.4,10.2,5.8Hz,1H),5.04(dd,J＝10.3,1.5Hz,1H),4.92(dd,J＝17.1,1.7Hz,1H),4.63(d,J＝5.8Hz,2H),4.30(q,J＝7.0Hz,2H),3.93(t,J＝6.5Hz,2H),1.95(t,J＝7.4Hz,2H),1.69(p,J＝6.7Hz,2H),1.50(p,J＝7.3Hz,2H),1.44-1.18(m,9H).

Example 33: preparation of 8- (4- (4- (2-allyl-1- (6-ethoxypyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-33)

Referring to the procedure of example 1, starting from intermediates 70 and 40, compound I-33 was prepared. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.36(s,1H),10.18(s,1H),8.83(s,1H),8.69(s,1H),7.96(t,J＝8.0Hz,1H),7.60(d,J＝8.8Hz,2H),7.42(d,J＝7.7Hz,1H),6.91(d,J＝8.7Hz,2H),6.78(d,J＝8.1Hz,1H),5.69(ddt,J＝16.4,10.3,5.8Hz,1H),5.04(dd,J＝10.3,1.5Hz,1H),4.92(dd,J＝17.2,1.6Hz,1H),4.63(d,J＝5.9Hz,2H),4.31(q,J＝7.0Hz,2H),3.10(t,J＝4.9Hz,4H),2.55(t,J＝5.1Hz,4H),2.35(t,J＝7.3Hz,2H),1.94(t,J＝7.3Hz,2H),1.57-1.15(m,9H).

Example 34: preparation of 8- (4- (2-allyl-1- (6-cyclopropylpyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenoxy) -N-hydroxyoctanoamide (I-34)

Referring to the procedure of example 32, starting from intermediates 68 and 74, compound I-34 was prepared. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.35(s,1H),10.24(s,1H),8.85(s,1H),8.68(s,1H),7.99(t,J＝8.2Hz,1H),7.68-7.46(m,3H),6.92(d,J＝8.7Hz,2H),6.83(d,J＝8.1Hz,1H),5.69(ddt,J＝16.5,10.2,6.0Hz,1H),5.03(dd,J＝10.3,1.5Hz,1H),4.91(dd,J＝17.1,1.5Hz,1H),4.74(d,J＝5.9Hz,2H),4.21(tt,J＝6.3,3.0Hz,1H),3.94(t,J＝6.5Hz,2H),1.95(t,J＝7.3Hz,2H),1.70(p,J＝6.7Hz,2H),1.50(p,J＝7.4Hz,2H),1.44-1.19(m,6H),0.88-0.68(m,4H).

Example 35: preparation of 8- (4- (4- (2-allyl-1- (6-cyclopropylpyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3,4-d ] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (I-35)

Referring to the procedure of example 1, starting from intermediates 76 and 40, compound I-35 was prepared. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.36(s,1H),10.20(s,1H),8.83(s,1H),8.69(s,1H),7.99(s,1H),7.67-7.44(m,3H),6.92(d,J＝8.7Hz,2H),6.83(d,J＝8.1Hz,1H),5.69(ddt,J＝16.5,10.2,6.0Hz,1H),5.03(dd,J＝10.3,1.5Hz,1H),4.91(dd,J＝17.2,1.6Hz,1H),4.74(d,J＝6.0Hz,2H),4.22(tt,J＝6.3,3.1Hz,1H),3.11(s,4H),2.53(s,4H),2.34(t,J＝7.4Hz,2H),1.94(t,J＝7.3Hz,2H),1.54-1.38(m,4H),1.36-1.12(m,6H),0.85-0.67(m,4H).

Example 36: preparation of 8- (4- (4- (6- (2, 6-dichlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (Compound II-1)

Step 1: synthesis of intermediate 78

To a eggplant-shaped bottle was added starting material 5 (10 g,42.98 mmol), methylamine hydrochloride (6.67 g,98.85 mmol), DIEA (18.39 g,141.83 mmol) and tetrahydrofuran (100 mL), and the reaction was allowed to proceed at 70℃for 12 hours, followed by completion by TLC (Petroleum ether: ethyl acetate=5:1). Suction filtration and concentration of the filtrate under reduced pressure gave 9.77g of a white solid as intermediate 78 in 100% yield. ¹ H NMR(300MHz,Chloroform-d)δ8.63(s,1H),8.22(s,1H),4.33(q,J＝7.0Hz,2H),3.10(d,J＝4.4Hz,3H),2.57(s,3H),1.38(t,J＝7.0Hz,3H).

Step 2: synthesis of intermediate 79

To the eggplant-shaped flask was added intermediate 78 (9.77 g,42.98 mmol), sodium hydroxide (2.06 g,51.58 mmol), tetrahydrofuran (100 mL) and water (100 mL), and the reaction was stirred at 50℃for 12 hours, and monitored by TLC (dichloromethane: methanol=10:1) for completion. Adding concentrated hydrochloric acid to adjust pH to acidity, concentrating under reduced pressure to remove tetrahydrofuran, Suction filtration and vacuum drying of the filter cake gave 8.56g of white solid as intermediate 79 in 100% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ8.96(s,1H),8.51(s,1H),3.05(d,J＝4.8Hz,3H),2.57(s,3H).

Step 3: synthesis of intermediate 80

To the eggplant-shaped flask was added intermediate 79 (8.56 g,42.98 mmol), DMF (5 d) and thionyl chloride (100 mL), and reacted at 90℃for 12 hours. The reaction solution was concentrated under reduced pressure, slurried with toluene, suction filtered, and the filtrate was concentrated under reduced pressure and transferred to a three-necked flask. To a three-necked flask was added 2, 6-dichloroaniline (6.96 g,42.98 mmol), pyridine (4.25 g,53.73 mmol) and dichloromethane (100 mL), and the mixture was stirred at room temperature for 12 hours, and TLC (Petroleum ether: ethyl acetate=5:1) monitored for completion of the reaction. The reaction solution was concentrated under reduced pressure, and 6.67g of a yellow solid was obtained by silica gel column chromatography as intermediate 80 in 45.2% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.33(s,1H),8.74(s,1H),8.62(s,1H),7.60(d,J＝7.9Hz,2H),7.41(t,J＝8.1Hz,1H),3.57(s,3H),2.95(d,J＝3.6Hz,3H).

Step 4: synthesis of intermediate 81

To the eggplant-shaped flask was added intermediate 80 (6.67 g,19.43 mmol), cesium carbonate (25.32 g,77.71 mmol), dibromomethane (10.13 g,58.28 mmol) and acetonitrile (100 mL), and the reaction was allowed to proceed to 80℃for 7 days, followed by TLC (Petroleum ether: ethyl acetate=5:1) to monitor completion of the reaction. The reaction solution was concentrated under reduced pressure, diluted with water, extracted with EA 3 times, the organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, suction-filtered, and the filtrate was concentrated under reduced pressure, and subjected to silica gel column chromatography to give 2.31g of a yellow solid as intermediate 81 in 33.4% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ8.83(s,1H),7.47(s,1H),7.10(t,J＝8.1Hz,1H),5.21(s,2H),3.10(s,3H),2.55(s,3H).

Step 5: synthesis of target Compound II-1

The procedure of example 1 was followed, starting from intermediates 81 and 40, to give the target compound II-1. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.36(s,1H),9.74(s,1H),8.45(s,1H),7.90(s,1H),7.70-7.57(m,4H),7.48(t,J＝7.6Hz,1H),6.91(d,J＝8.8Hz,2H),4.95(s,2H),3.12(s,4H),3.09(s,3H),2.60(s,4H),2.41(t,J＝7.6Hz,2H),1.94(t,J＝7.3Hz,2H),1.54-1.41(m,4H),1.29-1.21(m,6H).

Example 37: preparation of 8- (4- (6- (2, 6-dichlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-2)

The procedure of example 1 was followed, starting from intermediates 81 and 37, to give the target compound II-2. ¹ H NMR(300MHz,DMSO-d6)δ10.36(s,1H),9.77(s,1H),8.69(s,1H),8.46(s,1H),7.71-7.59(m,4H),7.54-7.43(m,1H),6.89(d,J＝9.1Hz,2H),4.96(s,2H),3.93(t,J＝6.5Hz,2H),3.09(s,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.6Hz,2H),1.53-1.15(m,8H).

Example 38: preparation of 8- (4- (6- (2-chlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-3)

In the same manner as in example 36, except for using intermediate 79 and 2-chloroaniline as raw materials, compound II-3 was produced. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.39(s,1H),9.74(s,1H),8.69(s,1H),8.45(s,1H),7.66(d,J＝8.5Hz,2H),7.62(dd,J＝7.2,2.1Hz,1H),7.52(dd,J＝7.4,2.2Hz,1H),7.44(pd,J＝7.3,1.7Hz,4H),6.89(d,J＝9.0Hz,2H),5.06(d,J＝9.0Hz,1H),4.89(d,J＝9.2Hz,1H),3.92(t,J＝6.5Hz,2H),3.18(d,J＝5.1Hz,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.7Hz,2H),1.50(p,J＝7.3Hz,2H),1.44-1.36(m,2H),1.35-1.20(m,4H).

Example 39: preparation of 8- (4- (4- (6- (2-chlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (II-4)

The procedure of example 37 was followed, starting from intermediates 85 and 40, to give the desired compound II-4. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.35(s,1H),9.68(s,1H),8.68(s,1H),8.44(s,1H),7.67-7.58(m,3H),7.55-7.38(m,3H),6.90(d,J＝9.1Hz,2H),4.98(d,J＝42.8Hz,2H),3.07(s,3H),3.07(t,J＝3.7Hz,4H),2.48(t,J＝3.7Hz,1H),2.30(t,J＝7.3Hz,2H),1.94(t,J＝7.3Hz,2H),1.57-1.37(m,4H),1.25(d,J＝8.4Hz,6H).

Example 40: preparation of 8- (4- (6- (2-fluorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-5)

The procedure of example 36 was followed, starting from intermediate 5, 2-fluoroaniline and 37, to give the target compound II-5. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.40(s,1H),9.75(s,1H),8.69(s,1H),8.46(s,1H),7.66(d,J＝8.5Hz,2H),7.53-7.23(m,4H),6.89(d,J＝9.0Hz,2H),5.04(s,2H),3.93(t,J＝6.4Hz,2H),3.09(s,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.5Hz,2H),1.50(p,J＝7.4Hz,2H),1.43-1.18(m,6H).

Example 41: preparation of 8- (4- (6- (2-fluorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-6)

The procedure of example 36 was followed, starting from intermediate 5 and 2-fluoroaniline and 40, to give the target compound II-6. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.34(s,1H),9.69(s,1H),8.68(s,1H),8.45(s,1H),7.62(d,J＝8.5Hz,2H),7.51-7.24(m,4H),6.90(d,J＝8.7Hz,2H),5.03(s,2H),3.15-2.99(m,J＝7.3Hz,7H),2.48(t,J＝5.0Hz,4H),2.30(t,J＝7.4Hz,2H),1.94(t,J＝7.4Hz,2H),1.54-1.37(m,4H),1.36-1.25(m,6H).

Example 42: preparation of N-hydroxy-8- (4- (8-methyl-5-oxo-6- (o-tolyl) -5,6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) octanamide (II-7)

The procedure of example 36 was followed, starting from intermediate 5, 2-methylaniline and 37, to give the title compound II-7.LC/MS: m/z=519.28 [ m+h ]] ⁺ . ¹ H NMR(400MHz,DMSO-d ₆ )δ10.40(s,1H),9.70(s,1H),8.68(s,1H),8.44(s,1H),7.66(d,J＝8.5Hz,2H),7.41-7.21(m,4H),6.89(d,J＝9.0Hz,2H),5.09(d,J＝9.2Hz,1H),4.82(d,J＝9.4Hz,1H),3.92(t,J＝6.5Hz,2H),3.08(s,3H),2.19(s,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.7Hz,2H),1.50(p,J＝7.3Hz,2H),1.44-1.36(m,2H),1.35-1.19(m,4H).

Example 43: preparation of N-hydroxy-8- (4- ((8-methyl-5-oxo-6- (o-tolyl) -5,6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) octanamide (II-8)

The procedure of example 36 was followed, starting from intermediate 5, 2-methylaniline and 40, to give the title compound II-8. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.34(s,1H),9.63(s,1H),8.68(s,1H),8.43(s,1H),7.62(d,J＝8.3Hz,3H),7.37-7.23(m,4H),6.90(d,J＝9.1Hz,2H),5.09(d,J＝9.4Hz,1H),4.81(d,J＝9.3Hz,1H),3.13-2.99(m,7H),2.49(t,J＝5.0Hz4H),2.30(t,J＝7.4Hz,2H),2.19(s,3H),1.94(t,J＝7.3Hz,2H),1.54-1.39(m,4H),1.32-1.21(m,6H).

Example 44: preparation of 8- (4- (6- (3-chlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-9)

In the same manner as in example 36, intermediate 5,3-chloroaniline and 37 are used as raw materials to prepare the target compound II-9. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.36(s,1H),9.75(s,2H),8.68(s,1H),8.48(s,1H),7.66(d,J＝8.4Hz,2H),7.50(t,J＝2.0Hz,1H),7.46(t,J＝8.0Hz,1H),7.39-7.31(m,2H),6.89(d,J＝9.1Hz,2H),5.12(s,2H),3.92(t,J＝6.5Hz,2H),3.11(s,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.6Hz,2H),1.50(p,J＝7.4Hz,2H),1.32(m,6H).

Example 45: preparation of 8- (4- (4- (6- (3-chlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (II-10)

The procedure of example 36 was followed, starting from intermediate 5, 3-chloroaniline and 40, to give the target compound II-10. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.34(s,1H),9.71(s,1H),8.66(s,1H),8.47(s,1H),7.62(d,J＝6.6Hz,2H),7.49(t,J＝1.9Hz,1H),7.46(t,J＝8.0Hz,1H),7.39-7.30(m,2H),6.90(d,J＝9.0Hz,2H),5.12(s,2H),3.10(s,3H),3.08(t,J＝4.8Hz,4H),2.49(t,J＝4.8Hz,4H),2.31(t,J＝7.4Hz,2H),1.94(t,J＝7.3Hz,2H),1.54-1.38(m,4H),1.34-1.17(m,6H).

Example 46: preparation of 8- (4- (6- (4-chlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-11)

The procedure of example 36 was followed, starting from intermediate 5, 4-chloroaniline and 37, to give the target compound II-11. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.40(s,1H),9.75(s,1H),8.68(s,1H),8.48(s,1H),7.66(d,J＝8.7Hz,2H),7.49(d,J＝8.8Hz,2H),7.41(d,J＝8.9Hz,2H),6.89(d,J＝9.1Hz,2H),5.10(s,2H),3.92(t,J＝6.5Hz,2H),3.10(s,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝7.4Hz,2H),1.50(p,J＝7.4Hz,2H),1.41-1.18(m,6H).

Example 47: preparation of 8- (4- (4- (6- (4-chlorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (II-12)

The procedure of example 36 was followed, starting from intermediate 5, 4-chloroaniline and 40, to give the target compound II-12. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.35(s,1H),9.69(s,1H),8.69(s,1H),8.46(s,1H),7.61(d,J＝8.0Hz,2H),7.49(d,J＝8.7Hz,2H),7.40(d,J＝8.7Hz,2H),6.90(d,J＝8.9Hz,2H),5.09(s,2H),3.10(s,3H),3.07(t,J＝4.2Hz,4H),2.49(t,J＝4.2Hz,4H),2.30(t,J＝7.3Hz,2H),1.94(t,J＝7.2Hz,2H),1.59-1.37(m,4H),1.16-1.16(m,6H).

Example 48: preparation of 8- (4- (6- (2-chloro-6-fluorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-13)

The procedure of example 36 was followed, starting from intermediate 5, 2-chloro-6-fluoroaniline and 37, to give the title compound II-13. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.38(s,1H),9.79(s,1H),8.68(s,1H),8.47(s,1H),7.66(d,J＝8.5Hz,2H),7.55-7.47(m,2H),7.42(ddd,J＝9.6,6.2,3.6Hz,1H),6.89(d,J＝9.0Hz,2H),4.97(q,J＝9.6Hz,2H),3.93(t,J＝6.5Hz,2H),3.09(s,3H),1.95(t,J＝7.3Hz,2H),1.69(p,J＝6.6Hz,2H),1.50(p,J＝7.4Hz,2H),1.42-1.21(m,6H).

Example 49: preparation of 8- (4- (4- (6- (2-chloro-6-fluorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4,5-d ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (II-14)

The procedure of example 36 was followed, starting from intermediate 5, 2-chloro-6-fluoroaniline and 40, to give target compound II-14. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.35(s,1H),9.74(s,1H),8.68(s,1H),8.46(s,1H),7.62(d,J＝7.8Hz,2H),7.54-7.49(m,2H),7.49-7.37(m,1H),6.90(d,J＝9.0Hz,2H),4.96(q,J＝9.6Hz,2H),3.16-3.02(m,7H),2.51-2.45(m,4H),2.31(t,J＝7.4Hz,2H),1.94(t,J＝7.3Hz,2H),1.56-1.39(m,4H),1.34-1.18(m,6H).

Example 50: preparation of 8- (4- (6- (2, 6-dichlorophenyl) -5-oxo-5, 6,8, 9-tetrahydroimidazo [1,2-a ] pyrimido [5,4-e ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-15)

Step 1: synthesis of intermediate 88

Triphosgene (9.16 g,30.87 mmol) and DCM (50 mL) were added to the eggplant-shaped bottle, intermediate 87 (5 g,30.87 mmol) was slowly added, triethylamine (9.01 mL,6.48 mmol) was added dropwise at-4℃and reacted at room temperature for 12 hours after the addition, TLC (Petroleum ether: ethyl acetate=10:1) monitored complete reaction, and concentrated under reduced pressure to give 5.80g of yellow solid as intermediate 88 in 99% yield. The next step was directly carried out without purification.

Step 2: synthesis of intermediate 90

To a eggplant-shaped bottle was added a solution of intermediate 88 (5.80 g,30.86 mmol) in diethyl ether (90 mL), a solution of intermediate 89 (15.27 g,95.28 mmol) in diethyl ether (90 mL) was added dropwise at 0 ℃ and reacted for 4 hours at room temperature, TLC (dichloromethane: methanol=20:1) monitored complete reaction, suction filtration was performed, and the filter cake was chromatographed on silica gel to give 8.86g of white solid as intermediate 90 in 82.4% yield. ¹ H NMR(300MHz,DMSO-d ₆ )δ8.09(s,1H),7.48(d,J＝8.1Hz,2H),7.26(dd,J＝8.6,7.6Hz,1H),6.80(t,J＝5.3Hz,1H),6.38(t,J＝5.7Hz,1H),3.21-2.96(m,4H),1.39(s,9H).

Step 3: synthesis of intermediate 91

To a eggplant-shaped flask was added intermediate 90 (8.86 g,25.36 mmol), DCM (100 mL) and a 1, 4-dioxane solution of hydrogen chloride (4M, 100 mL), and reacted at room temperature for 12 hours TLC (dichloromethane: methanol=20:1) monitored the reaction was complete. The reaction solution was suction-filtered, and the filter cake was vacuum-dried to obtain 7.22g of a white solid as intermediate 91 in 100% yield. The next step was directly carried out without purification. ¹ H NMR(300MHz,DMSO-d ₆ )δ8.43(s,1H),8.09(s,2H),7.49(d,J＝8.1Hz,2H),7.27(dd,J＝8.6,7.6Hz,1H),6.85(d,J＝5.1Hz,1H),3.31(q,J＝6.5Hz,2H),2.86(t,J＝6.7Hz,2H).

Step 4: synthesis of intermediate 92

To a eggplant-shaped bottle was added intermediate 91 (7.22 g,25.36 mmol), 5 (8.85 g,38.04 mmol), DIEA (13.11 g,101.43 mmol) and acetonitrile (125 mL), nitrogen was replaced, and the reaction was continued at 80 ℃ for 3 hours, monitored by TLC (dichloromethane: methanol=20:1) to complete the reaction, filtered off with suction, and dried with infrared to give 9.77g of a white solid as intermediate 92 in 86.7% yield. ¹ H NMR(400MHz,DMSO-d ₆ )δ8.55(s,1H),8.39(t,J＝5.8Hz,1H),8.05(s,1H),7.47(d,J＝8.1Hz,2H),7.28-7.22(m,1H),6.48(t,J＝5.8Hz,1H),4.27(q,J＝7.1Hz,2H),2.48(s,3H),1.30(t,J＝7.1Hz,3H).

Step 5: synthesis of intermediate 93

To the eggplant-shaped flask were added intermediate 92 (9.77 g,21.99 mmol) and phosphorus oxychloride (100 mL), and the reaction was monitored by TLC (petroleum ether: ethyl acetate=1:1) at 110 ℃ for 12 hours to complete the reaction. The reaction solution was concentrated under reduced pressure, saturated sodium bicarbonate solution was added at 0deg.C, and the cake was slurried with ethyl acetate, suction filtered, and infrared-dried to give 5.86g of a white solid as intermediate 93 in a yield of 70.1%. ¹ H NMR(400MHz,DMSO-d ₆ )δ8.77(s,1H),7.70(d,J＝8.0Hz,2H),7.57(dd,J＝8.8,7.5Hz,1H),4.18(t,J＝8.7Hz,2H),3.83(t,J＝8.7Hz,2H),2.59(s,3H).

Step 6: synthesis of target Compound II-15

The procedure of example 1 was followed, starting from intermediates 93 and 37, to give the target compound II-15. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.34(s,1H),10.19(s,1H),8.72-8.59(m,2H),7.79-7.48(m,5H),6.92(d,J＝9.0Hz,2H),4.17(t,J＝8.2Hz,2H),3.94(t,J＝6.5Hz,2H),3.82(t,J＝8.8Hz,2H),1.95(t,J＝7.3Hz,2H),1.70(p,J＝6.5Hz,2H),1.50(p,J＝7.4Hz,2H),1.33(m,6H).

Example 51: preparation of 8- (4- (4- (6- (2, 6-dichlorophenyl) -5-oxo-5, 6,8, 9-tetrahydroimidazo [1,2-a ] pyrimido [5,4-e ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (II-16)

The procedure of example 50 was followed, starting from intermediates 40 and 93, to give the desired compound II-16. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.34(s,1H),10.30(s,1H),8.65(s,1H),7.89(s,1H),7.74-7.62(m,3H),7.59-7.47(m,2H),6.92(d,J＝8.6Hz,2H),4.17(t,J＝7.0Hz,2H),3.81(t,J＝8.8Hz,2H),3.11(t,J＝5.0Hz,4H),2.60-2.50(m,4H),2.33(t,J＝7.4Hz,2H),1.94(t,J＝7.3Hz,2H),1.54-1.39(m,4H),1.35-1.15(m,6H).

Example 52: preparation of 8- (4- (4- (6- (o-tolyl) -8-methyl-5-oxo-5, 6-dihydropyridin [4,3-d ] pyrimidin-2-yl) amino) phenyl) piperazin-1-yl) -N-hydroxyoctanoamide (II-17)

Step 1: synthesis of intermediate 98

To the eggplant-shaped flask were added starting material 95 (95.00 g,658.94 mmol) and 96 (94.21 g,790.73 mmol), reacted at 80℃for 10 minutes followed by intermediate 97 (59.40 g,658.94 mmol), reacted at 110℃for 15 hours, and TLC monitored the reaction to be complete (petroleum ether: ethyl acetate=10:1). The reaction solution was cooled to room temperature, diluted with ethyl acetate, washed with saturated sodium bicarbonate solution, saturated sodium chloride solution, dried over anhydrous sodium sulfate, suction-filtered, and the filtrate was concentrated under reduced pressure to give 93.04g of a white solid as intermediate 98 in a yield of 62.4% by silica gel column chromatography. ¹ H NMR(300MHz,Chloroform-d)δ8.94(s,1H),4.39(q,J＝7.1Hz,2H),3.15(q,J＝7.5Hz,2H),2.62(s,3H),1.41(t,J＝7.1Hz,3H),1.31(t,J＝7.5Hz,3H).

Step 2: synthesis of intermediate 99 to a eggplant-shaped bottle was added intermediate 98 (93.04 g,411.15 mmol) and lithium hydroxide (19.69 g,822.31 mmol)Tetrahydrofuran (600 mL) and water (600 mL), and the reaction was monitored by TLC (dichloromethane: methanol=10:1) at room temperature for 3 hours. The reaction solution was concentrated under reduced pressure, pH was adjusted to acidity by adding 1M hydrochloric acid, suction filtration was performed, and the filter cake was dried under vacuum to obtain 44.86g of a white solid as an intermediate 99 in a yield of 100%. ¹ H NMR(300MHz,DMSO-d ₆ )δ8.90(s,1H),3.08(q,J＝7.4Hz,2H),2.56(s,3H),1.21(t,J＝7.4Hz,3H).

Step 3: synthesis of intermediate 100

To a eggplant-shaped bottle was added intermediate 99 (5.00 g,25.22 mmol), o-methylaniline (2.70 g,25.22 mmol), HATU (11.51 g,30.27 mmol), DIEA (4.89 g,37.83 mmol) and DMF (100 mL), and the reaction was allowed to proceed for 3 hours at room temperature, followed by TLC (Petroleum ether: ethyl acetate=3:1) to completion. 300mL of water was added to the reaction mixture, suction filtration was performed, and the filter cake was dried under vacuum to give 6.63g of a yellow solid as intermediate 100 in a yield of 91.5%. ¹ H NMR(400MHz,Chloroform-d)δ8.68(s,1H),8.47(d,J＝8.3Hz,1H),8.00(s,1H),7.45(d,J＝9.5Hz,1H),7.41-7.32(m,1H),7.20-7.11(m,1H),3.05(q,J＝7.5Hz,2H),2.64(s,3H),1.37(t,J＝7.5Hz,3H).

Step 4: synthesis of intermediate 101

To a eggplant-shaped bottle was added intermediate 100 (5.50 g,19.41 mmol), (chloromethylene) and dimethyl ammonium chloride (7.45 g,58.23 mmol) and DMF (30 mL), nitrogen substitution, reaction at 50 ℃ monitored by TLC (dichloromethane: methanol=35:1) to complete the reaction, quenched with saturated sodium bicarbonate solution, followed by addition of 100mL of water, suction filtration, and vacuum drying of the filter cake to give 4.00g of white solid as intermediate 101 in 69.3% yield by silica gel column chromatography. ¹ H NMR(300MHz,Chloroform-d)δ9.42(s,1H),7.67-7.56(m,1H),7.53-7.37(m,2H),7.23(q,J＝1.2Hz,1H),2.70(s,3H),2.34(d,J＝1.2Hz,3H).

Step 5: synthesis of target Compound II-17

The procedure of example 1 was followed, starting from intermediates 101 and 40, to give the desired compound II-17. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.34(s,1H),10.10(s,1H),9.12(s,1H),8.68(s,1H),7.92-7.67(m,2H),7.59(s,1H),7.45-7.27(m,4H),6.94(d,J＝8.9Hz,2H),3.09(t,J＝4.4Hz,4H),2.49(t,J＝3.6Hz,4H),2.30(t,J＝7.4Hz,2H),2.21(s,3H),2.10(s,3H),1.94(t,J＝7.3Hz,2H),1.53-1.41(m,4H),1.31-1.21(m,6H).

Example 53: preparation of 8- (4- (6- (o-tolyl) -8-methyl-5-oxo-5, 6-dihydropyridin [4,3-d ] pyrimidin-2-yl) amino) phenoxy) -N-hydroxyoctanoamide (II-18)

The procedure of example 1 was followed, starting from intermediate 101 and 37, to give the desired compound II-18. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.32(s,1H),10.15(s,1H),9.14(s,1H),8.67(s,1H),7.93-7.72(m,2H),7.60(s,1H),7.44-7.25(m,4H),6.93(d,J＝9.1Hz,2H),3.94(t,J＝6.5Hz,2H),2.21(s,3H),2.10(s,3H),1.95(t,J＝7.3Hz,2H),1.70(p,J＝6.6Hz,2H),1.50(p,J＝7.4Hz,2H),1.45-1.21(m,2H).

Example 54: preparation of N- (2-aminophenyl) -8- (4- (8-methyl-5-oxo-6- (o-tolyl) -5, 6-dihydropyridin [4,3-d ] pyrimidin-2-yl) amino) phenoxy) octanamide (II-19)

Step 1: synthesis of intermediate 103

Referring to the procedure of step 2 of example 52, starting from intermediate 102 (100 mg,0.19 mmol), intermediate 103 (80 mg) was produced in a yield of 84.6%. ¹ H NMR(300MHz,DMSO-d ₆ )δ10.15(s,1H),9.14(s,1H),7.83(d,J＝7.2Hz,2H),7.60(s,1H),7.46-7.26(m,4H),6.93(d,J＝9.1Hz,2H),3.95(t,J＝6.5Hz,2H),2.24-2.18(m,5H),2.10(s,3H),1.70(p,J＝6.4Hz,2H),1.51(p,J＝7.2Hz,2H),1.44-1.21(m,6H).

Step 2: synthesis of target Compound II-19

According to the method of example 52, step 3, starting from intermediate 103 (80 mg,0.16 mmol) and o-phenylenediamine (15 mg,0.16 mmol), the objective compound II-19 (38 mg) was produced in 40.2% yield. ¹ H NMR(400MHz,Chloroform-d)δ9.34(s,1H),8.82(s,1H),7.69(d,J＝8.0Hz,2H),7.46-7.14(m,7H),7.08-6.98(m,2H),6.91(d,J＝9.0Hz,2H),3.94(t,J＝6.6Hz,2H),2.43(t,J＝7.7Hz,2H),2.28(s,3H),2.19(s,3H),1.76(p,J＝6.5Hz,2H),1.73-1.63(m,2H),1.50-1.40(m,2H),1.39-1.31(m,4H).

Example 55: preparation of N- (2-amino-4-fluorophenyl) -8- (4- (8-methyl-5-oxo-6- (o-tolyl) -5, 6-dihydropyridin [4,3-d ] pyrimidin-2-yl) amino) phenoxy) octanamide (II-20)

According to the method of example 52, step 3, starting from intermediate 103 (100 mg,0.20 mmol) and p-fluorophenylenediamine (26 mg,0.20 mmol), the objective compound II-20 (92 mg) was produced in a yield of 75.6%. ¹ H NMR(400MHz,Chloroform-d)δ9.35(s,1H),7.77-7.62(m,3H),7.41-7.30(m,3H),7.24(d,J＝7.4Hz,1H),7.20(d,J＝1.3Hz,1H),7.10-7.02(m,1H),6.95(d,J＝9.0Hz,2H),6.54-6.43(m,2H),4.00(t,J＝6.4Hz,2H),2.42(t,J＝7.6Hz,2H),2.30(s,3H),2.21(s,3H),1.86-1.73(m,4H),1.54-1.42(m,6H).

Example 56: preparation of N ¹ - (4- (6- (2-chloro-6-fluorophenyl) -8-methyl-5-oxo-5, 6,7, 8-tetrahydropyrimidine [4, 5-d)]Pyrimidin-2-yl) amino) phenyl) -N ⁸ Hydroxy octanediamide (II-21)

The procedure of example 36 was followed, starting from intermediate 5, 2-chloro-6-fluoroaniline and 4, to give the title compound II-21. ¹ H NMR(400MHz,DMSO-d ₆ )δ10.35(s,1H),9.74(s,1H),9.17(s,1H),8.68(s,1H),8.46(s,1H),7.66-7.58(m,4H),7.38-7.27(m,3H),4.96(q,J＝9.6Hz,2H),3.19(s,3H),2.31(t,J＝7.4Hz,2H),1.94(t,J＝7.3Hz,2H),1.69-1.50(m,4H),1.39-1.31(m,4H).

Example 57: determination of in vitro cell proliferation (H1299) inhibitory Activity of Compounds

1. Experimental materials

H1299 non-small cell lung cancer cell lineIs obtained by purchasing from the company of Biotechnology, inc. of state Ji Ni European, RPMI-1640 plus 10% fetal bovine serum (Gibcol Co., ltd.) and is placed at 37℃with 5% CO ₂ Culturing under 95% humidity.

Cell lines	Cell type	Cell number/well	Culture medium
				H1299	Suspending	10000	RPMI-1640+10％FBS

2. Experimental procedure

Cells in the logarithmic growth phase were harvested, examined for cell viability by trypan blue staining, counted by a cell counter (thermo countness II) to give appropriate cell amounts, cell concentrations were adjusted, 50 μl of cell suspension was added to 96-well plates, 5000 cells per well, and cultured overnight. Preparing a solution of the compound to be tested, adding 50 mu L of the solution of the compound to be tested into each cell of each hole, wherein the final concentration of each compound to be tested is 5 mu M, arranging three compound concentration holes, and continuously culturing in an incubator for 72 hours. After 72 hours of incubation after dosing, 10 μl of CCK-8 solution was added to each well, and the 96-well plate was placed in an incubator, after further incubation for 2 hours, absorbance was read at 450nm and relative inhibition was calculated.

3. Experimental results

The results of the antiproliferative effect of some compounds on the H1299 cell line are shown in table 1 below.

TABLE 1 inhibitory Activity of certain compounds of the invention against H1299 cell proliferation

Inhibition rate: 50-100% (marked as A), 20-50% (marked as B) and <20% (marked as C).

Example 58: in vitro WEE1 kinase Activity inhibition assay

All compounds tested were dissolved in 100% dmso and formulated into 10mM stock solutions. And freezing at-20deg.C in dark place. Preparation of 1 Xkinase buffer 1x kinase buffer,50mM HEPES,pH 7.5, 10mM MgCl ₂ 2mM DTT,0.01%Tween-20,0.01% BSA. The compound was diluted with 100% dmso to 100 times the final desired highest inhibitor concentration in the reaction. For 0.1. Mu.M compound, a 10. Mu.M solution of compound in DMSO was prepared in this step. For 0.01. Mu.M compound, a 1. Mu.M solution of compound in DMSO was prepared in this step. To two wells in the same 96-well plate, 100 μl of 100% dmso was added for the compound-free control and enzyme-free control, and the plates were labeled as source plates. Transfer 40 μl of compound from the source plate into a new 384 well Echo plate as an intermediate plate. Transfer 200nL to 384 well assay plates from each well of a 384 well Echo plate. WEE1 inhibitor solutions were prepared in 1 Xkinase buffer at a concentration of 2 times the final concentration of each reagent in the assay. To each well of the assay plate, except for the control well without enzyme, 10 μl of kinase solution (instead 10 μl of 1x kinase buffer) was added. And (5) shaking the dish. Incubate at room temperature for 10 min. Substrate solutions of fluorescein-polygt and ATP were prepared in 1x kinase reaction buffer at a concentration 2 times the final concentration of each reagent required in the assay. To each well of the assay plate 10 μl of substrate solution was added to start the reaction. And (5) shaking the dish. The assay plate was covered and incubated at room temperature for a period of time. Preparation of 2-fold final concentration of detection solution in antibody dilution buffer And (3) liquid. mu.L of detection solution was added to each well of the assay plate to stop the reaction. Mix briefly with a centrifuge and incubate for 60 minutes at room temperature before reading fluorescence on a microplate reader. Data for excitation at 340nm, emission at 520nm and 495nm were collected on Envision.

RFU values are copied from the Envision program. The ratio of RFU 520nm/RFU 495nm was calculated. The ratio values are converted to percent inhibition values. Percent inhibition = (max-sample ratio)/(max-min) ×100."min" refers to the ratio of the enzyme-free control, and "max" refers to the ratio of the DMSO control. The data are presented in MS Excel and the curve is fitted by XLFit Excel plug-in 5.4.0.8. IC (integrated circuit) ₅₀ The calculation formula is as follows: y=bottom+ (Top-Bottom)/(1+ (IC) ₅₀ /X)^HillSlope。

Example 59: in vitro HDAC1 enzyme activity inhibition assay

1 Xassay buffer (modified Tris buffer) was prepared. Serial dilution of the compound: compounds were transferred to assay plates by Echo in 100% dmso. The final fraction of DMSO was 1%. Enzyme solutions were prepared in 1x assay buffer. Trypsin and Ac-peptide substrate were added to 1x assay buffer to prepare a substrate solution. Transfer 15 μl of enzyme solution to assay plate, or for low concentration controls, transfer 15 μl of 1x assay buffer. Incubate for 15 minutes at room temperature. To each well 10 μl of substrate solution was added to start the reaction. Plates were read on Envision, excited at 355nm and emitted at 460 nm. Curve fitting: data were fitted in Excel and inhibition was calculated: inh% = (Max-Signal)/(Max-Min) ×100, fitting data in XL-Fit, and calculating IC ₅₀ Value: y=bottom+ (Top-Bottom)/(1+ (IC) ₅₀ X) HillSlope), Y is inhibition, X is compound concentration. The in vitro enzyme activity inhibition results are shown in Table 2 below.

TABLE 2 results of percent inhibition test of WEE1 kinase and HDAC1 enzymes by partial compounds of the present invention

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As shown by the above activity results, most of the compounds designed by the invention have strong inhibition effect on WEE1 kinase and HDAC1 enzyme, and the antitumor cell proliferation activity is considerable, so that the compounds can be used for preparing medicines for treating diseases caused by over-expression of WEE1 kinase and/or HDAC enzyme.

Claims

1. An anilinopyrimido heterocyclic compound having a structure according to formula I and further comprising a pharmaceutically acceptable salt thereof:

wherein:

R ₃ selected from hydroxy, substituted or unsubstituted 5-7 membered aryl, 5-7 membered heteroaryl containing 1-3N, O, S, said substituents being selected from one or more hydrogen, halogen, C _1-6 Alkyl, C _1-6 Haloalkyl, C _1-6 Alkoxy, cyano, nitro, hydroxy, amino;

2. The anilinopyrimidino heterocycle compound according to claim 1, wherein in the structure:

R ₂ selected from one or more of hydrogen, halogen, C _1-4 An alkyl group;

3. The anilinopyrimidino heterocycle compound according to claim 1, wherein in the structure:

R ₁ selected from the group consisting of

R ₂ Selected from one or more of hydrogen, methyl, halogen;

R ₃ selected from hydroxy, o-aminophenyl, 2-amino-4-fluorophenyl;

x is selected from

4. The anilinopyrimidino heterocycle compound according to claim 1, wherein in the structure:

R ₁ selected from the group consisting of

R ₂ Selected from hydrogen;

R ₃ selected from hydroxyl groups;

x is selected from

Y is selected from- (CH) ₂ ) _n O-、-(CH ₂ ) _n CONH-、Where n=6, 7.

5. The anilinopyrimidino-heterocycle compound according to claim 1, characterized in that it is selected from any one of the following compounds:

6. the anilinopyrimidino heterocycle compound according to claim 1, wherein the pharmaceutically acceptable salt is a salt of the compound with an acid selected from any one of the following: hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, carbonic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, citric acid, malic acid, tartaric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, succinic acid, fumaric acid, salicylic acid, phenylacetic acid, mandelic acid, ferulic acid.

7. A process for the preparation of an anilinopyrimidino heterocycle compound according to claim 1, characterized in that it is selected from any one of the following processes:

wherein R is ₁ 、R ₂ 、R ₃ X, Y are defined in claim 1, R _c Methyl or ethyl;

8. A pharmaceutical composition comprising an anilinopyrimidino heterocycle compound of claim 1 and a pharmaceutically acceptable carrier.

9. Use of an anilinopyrimido heterocyclic compound according to claim 1 or a pharmaceutical composition according to claim 8 for the preparation of a WEE1 and HDAC dual target inhibitor drug.

10. The use according to claim 9, wherein the medicament is an anti-tumour medicament.