Anilinopyrimido heterocyclic compound, and preparation method therefor, pharmaceutical composition thereof and use thereof

Abstract

An anilinopyrimido heterocyclic compound, and a preparation method therefor, a pharmaceutical composition thereof and a use thereof. The compound has a structure as shown in formula I, further comprises a pharmaceutically acceptable salt thereof, and is capable of effectively inhibiting WEE1 and HDAC, with the optimal inhibition rate value reaching 90% or above, and further capable of effectively inhibiting tumor cell proliferation, with the inhibition rate reaching 50% or above; the compound has a wide range of applications and can exert efficacy at both the molecular level and the cellular level, and optimally, the nanomolar concentration level can be achieved; in addition, the preparation method is conducive to structure expansion and has high universality.

Description

Anilinopyrimidoheterocyclic compound and its preparation method, pharmaceutical composition and application Technical Field

[0001] The present invention relates to an anilino pyrimido heterocyclic compound, a preparation method thereof, a pharmaceutical composition and applications thereof, and in particular to an anilino pyrimido heterocyclic compound having WEE1 and HDAC dual-target inhibitory activity, a preparation method thereof, a pharmaceutical composition and applications thereof. Background Art

[0002] The primary mechanism of action of WEE1 inhibitors is through synthetic lethality with the p53 gene. Because most tumors harbor mutations or deletions in the p53 gene, they are reliant on the WEE1 kinase-mediated G2 / M cell cycle checkpoint. This arrests cells with DNA damage in the G2 / M phase, allowing time for DNA repair. Inhibition of the WEE1 kinase causes cells with DNA damage to enter mitosis prematurely, triggering mitotic catastrophe and ultimately leading to apoptosis. Several WEE1 inhibitors are currently in clinical trials, including Zentalis' ZN-C3 (Phase II), Debio's Debio-0123 (Phase I / II), Shanghai Yingpi Pharmaceutical's IMP7068 (Phase I), and SC0191 (Phase I), jointly developed by Zhikang Hongren and WuXi AppTec. However, tumor pathogenesis is complex, often involving multiple signaling pathways, and single-target drugs often have unsatisfactory results. Literature reports that acute myeloid leukemia cells treated with WEE1 inhibitors may produce compensatory pathway activation mediated by CHK1, thereby reducing the anti-proliferative effect of WEE1 inhibitors.

[0003] HDACs are classified into four categories, I, II, III, and IV, with a total of 18 subtypes. Overexpression of types I, II, and IV can alter normal cell cycle and metabolic behavior, thereby inducing tumorigenesis. Based on this, several HDAC inhibitors have been approved for marketing, including Vorinostat, Romidepsin, Belinostat, and Panobinostat, all approved by the US FDA. my country's independently developed cedaranid was also launched in China in 2015. These drugs have played a significant role in the treatment of hematological malignancies. Studies have shown that HDACs are involved in regulating DNA damage repair pathways and exhibit significant synergistic effects with WEE1 inhibitors. Inhibiting HDACs in various tumor cells can suppress the compensatory activation pathways caused by WEE1 inhibitors, thereby resensitizing tumor cells to WEE1 inhibitors.

[0004] Considering that combination therapy often has problems such as toxic side effects caused by drug-drug interactions and heterogeneous drug metabolic kinetic properties, the development of WEE1 / HDAC dual-target drugs is of great significance and potential application value.

[0005] Summary of the Invention

[0006] Objectives of the invention: The first objective of the present invention is to provide an anilinopyrimidoheterocyclic compound, the second objective is to provide a method for preparing the compound, the third objective is to provide a pharmaceutical composition comprising the compound, and the fourth objective is to provide an application of the compound and its pharmaceutical composition.

[0007] Technical solution: The anilinopyrimido heterocyclic compound of the present invention has a structure of Formula I, and further includes a pharmaceutically acceptable salt thereof:

[0008] in:

[0009] R1 is selected from substituted or unsubstituted C 1-4 Alkyl, C 2-4 Alkenyl, 5-7 membered aryl, 5-7 membered heteroaryl containing 1-3 N, O, S, the substituents are selected from one or more hydrogen, halogen, C 1-6 Alkyl, C 1-6 Halogenated alkyl, C 1-6 Alkoxy, cyano, nitro, hydroxy, amino;

[0010] R2 is selected from one or more of hydrogen, halogen, C 1-6 Alkyl, C 1-6 Halogenated alkyl, C 1-6 Alkoxy, cyano, nitro, hydroxy, amino;

[0011] R3 is selected from hydroxyl, substituted or unsubstituted 5-7 membered aryl, 5-7 membered heteroaryl containing 1-3 N, O, S, and the substituent is selected from one or more hydrogen, halogen, C 1-6 Alkyl, C 1-6 Halogenated alkyl, C 1-6 Alkoxy, cyano, nitro, hydroxy, amino;

[0012] X is selected from where R a Selected from hydrogen, halogen, C 1-4 Alkoxy, 3-7 membered cycloalkoxy, C 1-4 Halogenated alkyl, C 1-4 Hydroxyalkyl, C 1-4 Alkoxy-substituted C 1-4 Alkyl, cyano, nitro, amino, hydroxyl, Q is selected from CH, N, R b Selected from substituted or unsubstituted C1-4 Alkyl, C 2-4 Alkenyl, 5-7 membered aryl, 5-7 membered aromatic hetero groups containing 1-3 N, O, S, wherein the substituents are selected from one or more hydrogen, halogen, C 1-6 Alkyl, C 1-6 Halogenated alkyl, C 1-6 Alkoxy, cyano, nitro, hydroxy, amino;

[0013] Y is selected from -(CH2) n -、-(CH2) n O-, -(CH2) n S-, -(CH2) n CONH-, -(CH2) n NHCO-, -(CH2) n SO2NH-、-(CH2) n NHSO2-, Where n=0-7.

[0014] Preferably, in the structure:

[0015] R1 is selected from substituted or unsubstituted C 1-4 Alkyl, C 2-4 Alkenyl, phenyl, the substituent is selected from one or more hydrogen, halogen, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 alkoxy;

[0016] R2 is selected from one or more of hydrogen, halogen, C 1-4 alkyl;

[0017] R3 is selected from hydroxyl, substituted phenyl, and the substituent is selected from halogen, amino;

[0018] X is selected from where R a Selected from hydrogen, halogen, C 1-4 Alkoxy, 3-5 membered cycloalkoxy, C 1-4 Halogenated alkyl, 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 alkyl;

[0019] Y is selected from -(CH2) n -、-(CH2) n O-, -(CH2) n CONH-、 Where n=0-7.

[0020] Preferably, in the structure:

[0021] R1 is selected from

[0022] R2 is selected from one or more hydrogen, methyl, halogen;

[0023] R3 is selected from hydroxy, o-aminophenyl, 2-amino-4-fluorophenyl;

[0024] X is selected from

[0025] Y is selected from -(CH2) n -、-(CH2) n O-, -(CH2) n CONH-、 Where n=0-7.

[0026] Preferably, in the structure:

[0027] R1 is selected from

[0028] R2 is selected from hydrogen;

[0029] R3 is selected from hydroxyl;

[0030] X is selected from

[0031] Y is selected from -(CH2) n O-, -(CH2) n CONH-、 Where n=6,7.

[0032] Specifically, the anilinopyrimidoheterocyclic compound is selected from any one of the following compounds:

[0033] Preferably, the pharmaceutically acceptable salt is a salt formed by the anilinopyrimidoheterocyclic compound and any one of the following acids: 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, and ferulic acid.

[0034] The preparation method of the compound of the present invention is selected from any one of the following methods:

[0035] (1) When R3 is a hydroxyl group, the methyl mercapto compound is coupled and aminolyzed to obtain the compound I:

[0036] Compound IV is prepared by reacting compound V with mCPBA, and the solvent used is selected from chloroform, dichloromethane, toluene, tetrahydrofuran, 1,4-dioxane, ethyl acetate, acetone, N,N-dimethylformamide or a mixed solvent of any two thereof, preferably dichloromethane.

[0037] Compound II is prepared by reacting compound IV with compound III, the base used is selected from triethylamine and N,N-diisopropylethylamine, preferably N,N-diisopropylethylamine; the solvent used is selected from chloroform, dichloromethane, toluene, tetrahydrofuran, 1,4-dioxane, ethyl acetate, acetone, N,N-dimethylformamide or a mixed solvent of any two, preferably toluene.

[0038] Compound IA is prepared by reacting compound II with hydroxylamine hydrochloride, where the base used is selected from sodium ethoxide, potassium acetate, sodium hydroxide, potassium hydroxide, potassium carbonate, sodium carbonate, preferably potassium hydroxide; and the solvent used is selected from methanol, ethanol, n-propanol, isopropanol, tert-butanol, n-butanol, isobutanol, preferably methanol.

[0039] (2) When R3 is not a hydroxyl group, the methyl mercapto compound is coupled, hydrolyzed, and acylated to obtain the compound I:

[0040] Compound IIB is prepared by hydrolysis of compound II, wherein the base used is selected from lithium hydroxide, sodium hydroxide, potassium hydroxide, potassium carbonate, sodium carbonate, preferably sodium hydroxide; and the solvent used is selected from a mixed solvent of tetrahydrofuran, 1,4-dioxane, methanol, ethanol and water, preferably a mixed solvent of tetrahydrofuran and water.

[0041] Compound IB is prepared by reacting compound IIB with R3-NH2. The condensing agent used is selected from carbonyldiimidazole (CDI), dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDCI), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) or benzotriazol-1-yl-oxytripyrrolidinophosphine hexafluorophosphate (PyBOP), preferably HATU; the acid binding agent is selected from N,N-diisopropylethylamine or triethylamine, preferably N,N-diisopropylethylamine; and the solvent used is selected from dichloromethane, tetrahydrofuran, 1,4-dioxane, ethyl acetate, acetone, N,N-dimethylformamide, preferably N,N-dimethylformamide.

[0042] Wherein, R1, R2, R3, X, and Y are as defined above. c is methyl or ethyl;

[0043] The corresponding acid is reacted with the compound I prepared by the above method to form a salt to obtain a pharmaceutically acceptable salt of the compound I.

[0044] The pharmaceutical composition of the present invention comprises the anilinopyrimidoheterocyclic compound and a pharmaceutically acceptable carrier, and is prepared into common pharmaceutical preparations such as tablets, capsules, syrups, suspensions or injections by adding common pharmaceutical excipients such as spices, sweeteners, liquid / solid fillers, diluents, etc.

[0045] The anilinopyrimidoheterocyclic compounds and drug combinations thereof of the present invention are used to prepare WEE1 and HDAC dual-target inhibitor drugs, and are specifically used to prepare anti-tumor drugs.

[0046] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0047] This type of compound can effectively inhibit WEE1 and HDAC, with the optimal inhibition rate reaching more than 90%. It can also effectively inhibit tumor cell proliferation, with an inhibition rate of more than 50%. It has a wide range of applications and can exert its efficacy at both the molecular and cellular levels, with the optimal concentration reaching nanomolar levels. Its preparation method is easy to expand the structure and has good versatility. DETAILED DESCRIPTION

[0048] The technical solution of the present invention will be further described below in conjunction with embodiments.

[0049] Example 1: Preparation of N 1 -(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenyl)-N 8 Hydroxysuccinamide (Compound I-1)

[0050] Step 1: Synthesis of Intermediate 3

[0051] To an eggplant-shaped flask, starting materials 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) were added and dissolved. The mixture was stirred at room temperature for 3 hours, and the reaction was monitored for completion by TLC (DCM:MeOH = 35:1). The reaction solution was diluted with water and extracted three times with DCM. 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. Silica gel column chromatography afforded 2.84 g of a white solid, Intermediate 3, in a 94.2% yield. 1H NMR(400MHz,Chloroform-d)δ7.45(d,J=8.9Hz,2H),7.32(d,J=8.9Hz,2H),3.69(s,3H),2.3 9-2.29(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).

[0052] Step 2: Synthesis of Intermediate 4: To an eggplant-shaped flask, add Intermediate 3 (2.80 g, 7.40 mmol), TFA (19 mL), and DCM (19 ml). Stir at room temperature for 12 hours. TLC (dichloromethane:methanol = 20:1) monitors the reaction for completion. The reaction solution is concentrated under reduced pressure, quenched with saturated sodium bicarbonate, and extracted three times with ethyl acetate. The organic layers are combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and filtered. The filtrate is concentrated under reduced pressure to obtain 1.91 g of a pink solid, Intermediate 4, in a yield of 92.7%. 1 H NMR (300MHz, DMSO-d6) 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).

[0053] Step 3: Synthesis of Intermediate 7

[0054] To an eggplant-shaped flask, starting materials 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) were added and refluxed at 75°C for 12 hours. The reaction was monitored for completion by TLC (petroleum ether:ethyl acetate = 10:1). The reaction solution was concentrated under reduced pressure, slurried with ether, filtered, and the filtrate was concentrated under reduced pressure. Trifluoroacetic acid (100 mL) was added, and the mixture was stirred at room temperature for 1 hour, then at 70°C for 1 hour. The mixture was concentrated under reduced pressure. Sodium hydroxide (6 M, 200 mL) and ethanol (100 mL) were added, and the mixture was stirred at room temperature for 15 minutes. The reaction was monitored for completion by TLC (dichloromethane:methanol = 20:1). Concentrated hydrochloric acid was added to adjust the pH to acidic, and the mixture was concentrated under reduced pressure. A mixed solvent of DCM and MeOH was added to dissolve the product, and the product was filtered. The filtrate was concentrated under reduced pressure and purified by silica gel column chromatography to obtain 14.60 g of yellow solid, intermediate 7, with a yield of 59.4%. 1H NMR (400MHz, DMSO-d6) 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).

[0055] Step 4: Synthesis of Intermediate 9

[0056] To a three-necked flask, add starting material 8 (25.00 g, 115.72 mmol) and diethyl ether (200 mL). The atmosphere was replaced with nitrogen. Methylmagnesium iodide (3 M in diethyl ether, 100 mL) was added via syringe at 0°C and the temperature was slowly warmed to room temperature. Completion of the reaction was monitored by TLC (petroleum ether:ethyl acetate = 10:1). The reaction was quenched with 1 M hydrochloric acid and extracted three times with EA. The organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure to afford 25.00 g of a yellow oil (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).

[0057] Step 5: Synthesis of Intermediate 10

[0058] To a three-necked flask were 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). The atmosphere was replaced with argon, and trans-N,N'-dimethyl-1,2-cyclohexanediamine (10.28 g, 72.25 mmol) was added via syringe. The reaction was allowed to react at 85°C for 16 hours, and completion was monitored by TLC (dichloromethane:methane = 20:1). The reaction solution was cooled to room temperature, quenched with aqueous ammonia, and extracted three times with EA. The organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate concentrated under reduced pressure. The filtrate was purified by silica gel column chromatography to afford 12.30 g of a white solid, Intermediate 10, in a 52.4% yield. 1HNMR(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).

[0059] Step 6: Synthesis of Intermediate 11

[0060] To an eggplant-shaped flask, intermediate 10 (600 mg, 1.68 mmol), mCPBA (1.04 g, 6.04 mmol), and toluene (10 mL) were added and reacted at room temperature for 1 hour. The reaction was monitored for completion by TLC (petroleum ether:ethyl acetate = 1:1). Intermediate 4 (708 mg, 2.01 mmol) and DIEA (2.00 mL, 11.75 mmol) were then added and reacted at room temperature for 2 hours. The reaction was monitored for completion by TLC (dichloromethane:methanol = 25:1). The reaction mixture was quenched with saturated sodium bicarbonate solution and extracted three times with EA. 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. 366 mg of a yellow solid, intermediate 11, was obtained by silica gel column chromatography. The yield was 37.1%. 1 H NMR(300MHz,Chloroform-d)δ8.85(s,1H),7.92(t,J=7.9Hz,1H),7.75(d,J=8.0 Hz,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).

[0061] Step 7: Synthesis of Target Compound I-1: Hydroxylamine hydrochloride (710 mg, 10.21 mmol), methanol (6 mL), and potassium hydroxide (860 mg, 15.31 mmol) were added to an eggplant-shaped flask and reacted at 40°C for 10 minutes. The mixture was filtered with suction, and the filtrate was added with intermediate 11 (300 mg, 0.51 mmol). The mixture was reacted at room temperature for 1 hour. TLC (dichloromethane:methanol = 10:1) monitored the completion of the reaction. 1 M hydrochloric acid was added to the reaction solution to adjust the pH to neutral. The mixture was extracted three times with DCM. 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. PTLC yielded 62 mg of a yellow solid, the target compound I-1, in a yield of 20.6%. 1 H NMR (300MHz, DMSO-d6) 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=5.9 Hz,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).

[0062] Example 2: Preparation of 2-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenyl)-N-hydroxyacetamide (I-2)

[0063] Step 1: Synthesis of intermediate 13

[0064] By referring to the method of Example 1, intermediate 10 (600 mg, 1.68 mmol) and 12 (277 mg, 1.68 mmol) were used as raw materials to obtain intermediate 13 (264 mg) with a yield of 33.1%. 1H 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).

[0065] Step 2: Synthesis of target compound I-2

[0066] Referring to the method of Example 1, intermediate 13 (264 mg, 0.56 mmol) was used as raw material to prepare compound I-2 (102 mg) with a yield of 38.6%. 1 H NMR (400MHz, DMSO-d6+D2O) 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).

[0067] Example 3: Preparation of 3-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenyl)-N-hydroxypropanamide (I-3)

[0068] Step 1: Synthesis of Intermediate 15

[0069] To an eggplant-shaped flask, starting material 14 (2.70 g, 16.42 mmol), tetrabutylammonium nitrate (10.00 g, 32.84 mmol), and DCM (40 mL) were added. Trifluoromethanesulfonic anhydride (4.63 g, 16.41 mmol) was slowly added at -30°C. The reaction was incubated for 30 minutes and monitored for completion by TLC (petroleum ether:ethyl acetate = 10:1). Sodium bicarbonate was added to the reaction solution, filtered, and the filtrate was concentrated under reduced pressure. Silica gel column chromatography afforded 940 mg of a yellow solid, Intermediate 15, in a 27.4% yield. 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).

[0070] Step 2: Synthesis of Intermediate 16

[0071] To an eggplant-shaped flask, intermediate 15 (940 mg, 4.49 mmol), Pd / C (100 mg), methanol (10 mL), and ethyl acetate (1 mL) were added. The mixture was allowed to react at room temperature for 12 hours. TLC (petroleum ether:ethyl acetate = 10:1) monitored the reaction for completion. The reaction solution was filtered through celite, and the filtrate was concentrated under reduced pressure. Silica gel column chromatography afforded 572 mg of a yellow solid, intermediate 16, in a 71.0% yield. 1 H NMR (300MHz, DMSO-d6) 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).

[0072] Step 3: Synthesis of Intermediate 17

[0073] Referring to the method of Example 1, intermediate 10 (600 mg, 1.68 mmol) and 16 (333 mg, 2.01 mmol) were used as raw materials to prepare intermediate 17 (242 mg) with a yield of 29.5%. 1H 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).

[0074] Step 4: Synthesis of target compound I-3

[0075] Referring to the method of Example 1, intermediate 17 (242 mg, mmol) was used as starting material to obtain the target compound I-3 (152 mg) in a yield of 62.8%. 1 H NMR(400MHz,DMSO-d6)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).

[0076] Example 4: Preparation of 2-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxyacetamide (I-4)

[0077] Step 1: Synthesis of Intermediate 20

[0078] To an eggplant-shaped flask were 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). The mixture was stirred at room temperature for 12 hours. The reaction was monitored for completion by TLC (dichloromethane:methanol = 35:1). 200 mL of water was added to the reaction solution, which was filtered with suction and dried over infrared spectroscopy to obtain 6.05 g of a white solid, intermediate 20, in a 90.0% yield. 1 H NMR (400MHz, Chloroform-d) δ7.28 (d, J = 9.0 Hz, 2H), 6.88 (d, J = 9.0 Hz, 2H), 4.62 (s, 2H), 3.82 (s, 3H), 1.53 (s, 9H).

[0079] Step 2: Synthesis of Intermediate 21

[0080] To an eggplant-shaped flask, 20 (6.00 g, 21.33 mmol), trifluoroacetic acid (20 mL), and dichloromethane (20 mL) were added and stirred at room temperature for 3 hours. The reaction was monitored for completion by TLC (dichloromethane:methanol = 35:1). The reaction solution was concentrated under reduced pressure, saturated sodium bicarbonate solution was added, and extraction was performed three times with EA. The organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain 3.81 g of a yellow liquid, intermediate 21, in a 98.6% yield. 1 H NMR (400MHz, Chloroform-d) δ6.79 (d, J = 8.8 Hz, 2H), 6.66 (d, J = 8.9 Hz, 2H), 4.58 (s, 2H), 3.82 (s, 3H), 3.23 (s, 2H).

[0081] Step 3: Synthesis of Intermediate 22

[0082] Referring to the method of Example 1, intermediate 22 (477 mg) was prepared using intermediate 10 (600 mg, 1.68 mmol) and 21 (365 mg, 2.01 mmol) as raw materials with a yield of 57.9%. 1H NMR (300MHz, Chloroform-d) δ8.85 (s, 1H), 7.87 (t, J = 7.9 Hz, 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).

[0083] Step 4: Synthesis of target compound I-4

[0084] Referring to the method of Example 1, intermediate 22 (466 mg, 0.95 mmol) was used as the starting material to obtain the target compound I-4 (192 mg) with a yield of 41.1%. 1 H NMR(400MHz,DMSO-d6)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).

[0085] Example 5: Preparation of 3-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxypropanamide (I-5)

[0086] Step 1: Synthesis of intermediate 24

[0087] To an eggplant-shaped flask were added starting materials 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). The mixture was refluxed at 105°C for 12 hours. TLC (dichloromethane:methanol = 10:1) monitored the reaction for completion. The reaction solution was cooled to room temperature, and the pH was adjusted to acidic with 6 M hydrochloric acid. The mixture was extracted three times with EA. The organic layers were combined, and the mixture was extracted three times with saturated sodium bicarbonate solution. The aqueous layers were combined, and the pH was adjusted to acidic with 6 M hydrochloric acid. The mixture was filtered, and the filter cake was dried under vacuum to obtain 2.74 g of a white solid, intermediate 24, in a yield of 24.7%. 1 H NMR (300MHz, DMSO-d6) 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).

[0088] Step 2: Synthesis of Intermediate 25

[0089] Intermediate 24 (2.74 g, 12.98 mmol) and anhydrous methanol (30 mL) were added to a three-necked flask. Acetyl chloride (0.51 g, 6.49 mmol) was added via syringe at -4°C. The mixture was kept warm for 10 h. The reaction was monitored by TLC (dichloromethane:methanol = 10:1) to determine completion. The mixture was filtered and the filter cake was dried under vacuum to obtain 2.62 g of a gray solid, intermediate 25, in a yield of 89.7%. 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).

[0090] Step 3: Synthesis of Intermediate 26

[0091] Referring to Example 3, intermediate 25 (2.6 g, 11.55 mmol) was used as starting material to obtain intermediate 26 (2.24 g) with a yield of 99.3%. 1 HNMR (300MHz, DMSO-d6) 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).

[0092] Step 4: Synthesis of target compound I-5

[0093] Referring to the method of Example 1, intermediates 26 and 10 were used as raw materials to prepare the target compound I-5. 1 H NMR(400MHz,DMSO-d6)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).

[0094] Example 6: Preparation of 4-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxybutanamide (I-6)

[0095] Referring to the method of Example 4, the target compound 6 was prepared using the intermediate methyl 4-bromobutyrate as the starting material. 1 H NMR (300MHz, DMSO-d6) 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).

[0096] Example 7: Preparation of 5-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxypentanamide (I-7)

[0097] Referring to the method of Example 4, compound I-7 was prepared using the intermediate 5-bromovalerate methyl ester as the starting material. 1 H NMR(300MHz,DMSO-d6)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=18.7 Hz,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).

[0098] Example 8: Preparation of 6-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxyhexanamide (I-8)

[0099] Referring to the method of Example 4, compound I-8 was prepared using methyl 6-bromohexanoate as the starting material. 1 H NMR (300MHz, DMSO-d6) δ10.39(s,1H),10.22(s,1H),8.85(s,1H),8.71(s,1H),8.05(t,J=7.8Hz,1 H),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).

[0100] Example 9: Preparation of 7-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxyheptylamide (I-9)

[0101] Referring to the method of Example 4, compound I-9 was prepared using methyl 7-bromoheptanoate as the starting material. 1 H NMR (400MHz, DMSO-d6) δ10.34(s,1H),10.22(s,1H),8.85(s,1H),8.66(s,1H),8.05(t,J=7.7Hz,1 H),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).

[0102] Example 10: Preparation of 8-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxyoctanamide (I-10)

[0103] Referring to the method of Example 4, compound I-10 was prepared using ethyl 8-bromooctanoate as the starting material. 1H NMR (400MHz, DMSO-d6) δ10.36(s,1H),10.23(s,1H),8.85(s,1H),8.68(s,1H),8.05(t,J=8.1 Hz,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).

[0104] 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)

[0105] Step 1: Synthesis of intermediate 29

[0106] To an eggplant-shaped flask were added starting materials 18 (4.43 g, 28.95 mmol), 28 (5 g, 24.13 mmol), DIEA (7.80 g, 60.32 mmol) and DMF (50 mL). The mixture was stirred at room temperature overnight. The reaction was monitored for completion by TLC (dichloromethane:methanol = 15:1). 200 mL of water was added, the mixture was filtered, and the filter cake was dried under IR to obtain 6.04 g of a yellow solid, intermediate 29, in a yield of 89.6%. 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).

[0107] Step 2: Synthesis of Intermediate 30

[0108] To an eggplant-shaped flask, intermediate 29 (6.00 g, 24.07 mmol), Pd / C (300 mg), and methanol (50 mL) were added. The atmosphere was replaced with hydrogen and the reaction was allowed to proceed at room temperature for 12 hours. TLC (dichloromethane:methanol = 15:1) monitored the reaction for completion. The reaction solution was filtered through celite, concentrated under reduced pressure, and purified by silica gel column chromatography to afford 5.24 g of a brown solid, intermediate 30, in an 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).

[0109] Step 3: Synthesis of target compound I-11

[0110] Referring to the method of Example 1, intermediates 30 and 10 were used as raw materials to prepare the target compound I-11. 1 H NMR (400MHz, DMSO-d6) 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=8.1H z,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).

[0111] Example 12: Preparation of 3-(4-(4-((2-allyl-1-(6-(2-hydroxypropan-2-yl)pyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenyl)piperazin-1-yl)-N-hydroxypropanamide (I-12)

[0112] Referring to the method of Example 11, the target compound I-12 was prepared using methyl 3-bromopropionate as the starting material. 1H NMR (400MHz, DMSO-d6) 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),3.0 9(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).

[0113] 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-hydroxybutanamide (I-13)

[0114] Referring to the method of Example 11, the target compound I-13 was prepared using methyl 4-bromobutyrate as the starting material. 1 H NMR (300MHz, DMSO-d6) 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).

[0115] Example 14: Preparation of 5-(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)

[0116] Referring to the method of Example 11, the target compound I-14 was prepared using methyl 5-bromopentanoate as the starting material. 1 H NMR (400MHz, DMSO-d6) δ10.37(s,1H),10.15(s,1H),8.83(s,1H),8.68(s,1H),8.05(t,J=6.3 Hz,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=16.5,1 0.2,5.9Hz,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).

[0117] 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-hydroxyhexanamide (I-15)

[0118] Referring to the method of Example 11, the target compound I-15 was prepared using methyl 6-bromohexanoate as the starting material. 1H NMR (300MHz, DMSO-d6) 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),5.3 6(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),3.1 9-2.97(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).

[0119] 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-hydroxyheptylamide (I-16)

[0120] Referring to the method of Example 11, the target compound I-16 was prepared using methyl 7-bromoheptanoate as the starting material. 1 H NMR (300MHz, DMSO-d6) 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).

[0121] 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-hydroxyoctanamide (I-17)

[0122] Referring to the method of Example 11, the target compound I-17 was prepared using 8-bromooctanoic acid methyl ester as the starting material. 1 H NMR (300MHz, DMSO-d6) 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),2.5 0-2.40(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).

[0123] 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-hydroxyoctanamide (I-18)

[0124] Step 1: Referring to the method of Example 4, intermediate 35 and 19 were used as starting materials to prepare intermediate 37. 1 H NMR (300MHz, DMSO-d6) 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).

[0125] Step 2: Referring to the method of Example 1, intermediate 32 is used as the starting material to prepare intermediate 34.1 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,1 H),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).

[0126] Step 3: Referring to the method of Example 1, intermediates 34 and 37 were used as raw materials to prepare the target compound I-18. 1 H NMR(300MHz,DMSO-d6)δ10.37(s,1H),10.14(s,1H),8.83(s,1H),8.70(s,1H),7.7 4-7.42(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).

[0127] 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-hydroxyoctanamide (I-19)

[0128] Referring to the method of Example 11, intermediate 40 was prepared using intermediates 35 and 28 as raw materials. 1H 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).

[0129] Referring to the method of Example 1, intermediates 34 and 40 were used as raw materials to prepare the target compound I-19. 1 H NMR(300MHz,DMSO-d6)δ10.35(s,1H),10.10(s,1H),8.81(s,1H),8.71(s,1H),7.68-7.42(m,5 H),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=5.0H z,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).

[0130] Example 20: Preparation of N-hydroxy-8-(4-(1-(6-(2-hydroxypropane-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)

[0131] Step 1: Synthesis of intermediate 42

[0132] To an eggplant-shaped flask, starting material 5 (10.00 g, 42.98 mmol) and ethanol (200 mL) were added. Hydrazine hydrate (5.24 mL, 86.96 mmol) was added at 0°C and allowed to react for 1 hour. Completion of the reaction was monitored by TLC (petroleum ether:ethyl acetate = 5:1). The reaction solution was concentrated under reduced pressure, slurried with methyl tert-butyl ether, and filtered. The filtrate was concentrated under reduced pressure to afford 5.19 g of a white solid, intermediate 42, in a 52.9% yield. 1H NMR (300MHz, DMSO-d6) 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).

[0133] Step 2: Synthesis of Intermediate 43

[0134] To an eggplant-shaped flask, intermediate 42 (5.00 g, 21.92 mmol) and acetone (100 mL) were added. The mixture was reacted at 70°C for 12 hours, concentrated under reduced pressure, and dissolved in methanol (100 mL). Sodium cyanoborohydride (2.07 g, 32.89 mmol) and concentrated hydrochloric acid (2 mL) were added at 0°C and allowed to react at room temperature for 12 hours. The reaction was monitored for completion by TLC (petroleum ether:ethyl acetate = 3:1). The reaction was quenched by the addition of saturated sodium bicarbonate solution and extracted three 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. Purification by silica gel column chromatography afforded 4.15 g of a white solid, intermediate 43, in a yield of 70.0%. 1 H NMR(300MHz,DMSO-d6)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).

[0135] Step 3: Synthesis of Intermediate 44

[0136] To an eggplant-shaped flask, intermediate 43 (4.10 g, 15.17 mmol), sodium hydroxide solution (5 M, 50 mL), and methanol (25 mL) were added and stirred at room temperature for 12 hours. The reaction was monitored for completion by TLC (dichloromethane:methanol = 3:1). The reaction solution was adjusted to acidic pH with concentrated hydrochloric acid and concentrated under reduced pressure. Dissolved in a mixed solvent of DCM and MeOH, filtered, and the filtrate was concentrated under reduced pressure. Silica gel column chromatography afforded 1.73 g of a white solid, intermediate 44, in a 50.8% yield. 1 H NMR (300MHz, DMSO-d6) δ8.22(s,1H),4.57(hept,J=6.6Hz,1H),2.39(s,3H),1.22(d,J=6.7Hz,6H).

[0137] Step 4: Synthesis of target compound I-20

[0138] Referring to the method of Example 1, compound I-20 was prepared using intermediate 44 as raw material. 1 H NMR (300MHz, DMSO-d6) 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=8.4 Hz,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).

[0139] 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)

[0140] Referring to the method of Example 1, compound I-21 was prepared using intermediates 45 and 40 as raw materials. 1 H NMR(300MHz,DMSO-d6)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).

[0141] 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-hydroxyoctanamide (I-22)

[0142] Step 1: Synthesis of intermediate 48

[0143] To an eggplant-shaped flask, intermediate 9 (5.50 g, 25.45 mmol), iodomethane (10.84 g, 76.36 mmol), and anhydrous tetrahydrofuran (100 mL) were added, along with sodium hydride (2.93 g, 76.36 mmol). The mixture was stirred at room temperature for 12 hours, and the reaction was complete as monitored by TLC (petroleum ether:ethyl acetate = 5:1). The reaction was quenched with water and extracted three times with EA. 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 to afford 5.42 g of a yellow oil, intermediate 48, in a yield of 92.6%. 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).

[0144] Step 2: Synthesis of target compound I-22

[0145] Referring to the method of Example 1, intermediates 48 and 37 were used as raw materials to prepare the target compound I-22. 1 H NMR (300MHz, DMSO-d6) 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=1 6.4,10.1,5.9Hz,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).

[0146] 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-hydroxyoctanamide (I-23)

[0147] Referring to the method of Example 1, the target compound I-23 was prepared using intermediates 49 and 40 as starting materials. 1H NMR (300MHz, DMSO-d6) 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,4 H),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).

[0148] Example 24: Preparation of N-hydroxy-8-(4-(1-(6-(2-hydroxypropane-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)

[0149] Step 1: Synthesis of Intermediate 51

[0150] To an eggplant-shaped flask were 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). The mixture was stirred at room temperature overnight. The reaction was monitored by TLC (petroleum ether:ethyl acetate = 3:1) for completion. The mixture was diluted with water and extracted three times with EA. 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. 2.85 g of a white solid, intermediate 51, was obtained by silica gel column chromatography. The yield was 43.6%. 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).

[0151] Step 2: Synthesis of target compound I-24

[0152] Referring to the method of Example 20, compound I-24 was prepared using intermediate 51 as raw material. 1H NMR (300MHz, DMSO-d6) δ10.44-10.29(m,2H),8.93(s,1H),8.69(s,1H),8.02(t,J=7.8 Hz,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=7.2 Hz,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).

[0153] 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)

[0154] The target compound I-25 was prepared by following the method of Example 1 using intermediates 53 and 40 as starting materials. LC / MS: m / z = 680.37 [M+H] + .

[0155] 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-hydroxyoctanamide (I-26)

[0156] Referring to the method of Example 1, intermediates 56 and 7 were used as raw materials to prepare the target compound I-26. 1H NMR (400MHz, DMSO-d6) 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(d dt,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).

[0157] 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-hydroxyoctanamide (I-27)

[0158] Referring to the method of Example 1, intermediates 57 and 40 were used as raw materials to prepare the target compound I-27. 1 H NMR (400MHz, DMSO-d6) 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).

[0159] 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-hydroxyoctanamide (I-28)

[0160] Following the method of Example 1, intermediates 60 and 7 were used as starting materials to obtain the target compound I-28. LC / MS: m / z = 586.24 [M+H] + .

[0161] 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-hydroxyoctanamide (I-29)

[0162] Referring to the method of Example 1, intermediates 61 and 40 were used as starting materials to prepare the target compound I-29. LC / MS: m / z = 654.30 [M+H] + .

[0163] 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-hydroxyoctanamide (I-30)

[0164] Referring to the method of Example 1, compound I-30 was prepared using intermediates 64 and 7 as raw materials. 1 H NMR (400MHz, DMSO-d6) 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,2 H),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).

[0165] 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-hydroxyoctanamide (I-31)

[0166] Referring to the method of Example 1, compound I-31 was prepared using the intermediate as raw material. 1 H NMR(400MHz,DMSO-d6)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).

[0167] 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-hydroxyoctanamide (I-32)

[0168] Step 1: Synthesis of Intermediate 69

[0169] Anhydrous ethanol (5.8 g, 126.64 mmol) and tetrahydrofuran (50 mL) were added to an eggplant-shaped flask. Sodium hydride was added at 0°C, and a tetrahydrofuran solution (50 mL) of the starting material 68 (10 g, 42.21 mmol) was added dropwise. The reaction was allowed to react at room temperature for 12 h. The reaction was completed as monitored by TLC (petroleum ether:ethyl acetate = 10:1). The reaction was quenched with water and extracted three times with EA. 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. 7.11 g of a transparent oil was obtained by silica gel column chromatography, which was intermediate 69 in a yield of 83.4%. 1H 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).

[0170] Step 2: Synthesis of target compound I-32

[0171] Referring to the method of Example 30, compound I-32 was prepared using intermediates 69 and 7 as raw materials. 1 H NMR (400MHz, DMSO-d6) 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=16.4,10.2,5.8H z,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).

[0172] 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-hydroxyoctanamide (I-33)

[0173] Referring to the method of Example 1, compound I-33 was prepared using intermediates 70 and 40 as raw materials. 1H NMR (300MHz, DMSO-d6) 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=16.4,10.3,5.8H z,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).

[0174] Example 34: Preparation of 8-(4-(2-allyl-1-(6-cyclopropyloxypyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenoxy)-N-hydroxyoctanamide (I-34)

[0175] Referring to the method of Example 32, compound I-34 was prepared using intermediates 68 and 74 as raw materials. 1 H NMR (400MHz, DMSO-d6) 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).

[0176] Example 35: Preparation of 8-(4-(4-(2-allyl-1-(6-cyclopropyloxypyridin-2-yl)-3-oxo-2,3-dihydro-1H-pyrazolo[3,4-d]pyrimidin-6-yl)amino)phenyl)piperazin-1-yl)-N-hydroxyoctanamide (I-35)

[0177] Referring to the method of Example 1, compound I-35 was prepared using intermediates 76 and 40 as raw materials. 1 H NMR (400MHz, DMSO-d6) 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).

[0178] Example 36: Preparation of 8-(4-(4-(6-(2,6-dichlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenyl)piperazin-1-yl)-N-hydroxyoctanamide (Compound II-1)

[0179] Step 1: Synthesis of Intermediate 78

[0180] To an eggplant-shaped flask, 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) were added. The mixture was reacted at 70°C for 12 hours. Completion of the reaction was monitored by TLC (petroleum ether:ethyl acetate = 5:1). The filtrate was filtered and concentrated under reduced pressure to afford 9.77 g of a white solid, Intermediate 78, in a 100% yield. 1H 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).

[0181] Step 2: Synthesis of Intermediate 79

[0182] To an eggplant-shaped flask, intermediate 78 (9.77 g, 42.98 mmol), sodium hydroxide (2.06 g, 51.58 mmol), tetrahydrofuran (100 mL), and water (100 mL) were added and stirred at 50°C for 12 hours. Completion of the reaction was monitored by TLC (dichloromethane:methanol = 10:1). The pH was adjusted to acidic with concentrated hydrochloric acid. The tetrahydrofuran was removed by concentration under reduced pressure, and the mixture was filtered and the filter cake dried under vacuum to afford 8.56 g of a white solid, intermediate 79, in a 100% yield. 1 H NMR (300MHz, DMSO-d6) δ8.96 (s, 1H), 8.51 (s, 1H), 3.05 (d, J = 4.8Hz, 3H), 2.57 (s, 3H).

[0183] Step 3: Synthesis of Intermediate 80

[0184] To an eggplant-shaped flask, intermediate 79 (8.56 g, 42.98 mmol), DMF (5d), and thionyl chloride (100 mL) were added and reacted at 90°C for 12 hours. The reaction solution was concentrated under reduced pressure, slurried with toluene, and filtered. The filtrate was concentrated under reduced pressure and transferred to a three-necked flask. 2,6-Dichloroaniline (6.96 g, 42.98 mmol), pyridine (4.25 g, 53.73 mmol), and dichloromethane (100 mL) were added to the three-necked flask and stirred at room temperature for 12 hours. Completion of the reaction was monitored by TLC (petroleum ether:ethyl acetate = 5:1). The reaction solution was concentrated under reduced pressure and purified by silica gel column chromatography to obtain 6.67 g of a yellow solid, intermediate 80, in a 45.2% yield. 1 H NMR (300MHz, DMSO-d6) 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).

[0185] Step 4: Synthesis of Intermediate 81

[0186] 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) were added to an eggplant-shaped flask and reacted at 80°C for 7 days. TLC (petroleum ether:ethyl acetate = 5:1) monitored the reaction for completion. The reaction solution was concentrated under reduced pressure, diluted with water, and extracted three times with EA. The organic layers were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure and purified by silica gel column chromatography to obtain 2.31 g of a yellow solid, Intermediate 81, in a yield of 33.4%. 1 H NMR (300MHz, DMSO-d6) 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).

[0187] Step 5: Synthesis of target compound II-1

[0188] Referring to the method of Example 1, intermediates 81 and 40 were used as raw materials to prepare the target compound II-1. 1 H NMR (400MHz, DMSO-d6) 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,2 H),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).

[0189] Example 37: Preparation of 8-(4-(6-(2,6-dichlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)-N-hydroxyoctanamide (II-2)

[0190] Referring to the method of Example 1, intermediates 81 and 37 were used as raw materials to prepare the target compound II-2. 1H 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).

[0191] Example 38: Preparation of 8-(4-(6-(2-chlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)-N-hydroxyoctanamide (II-3)

[0192] Referring to the method of Example 36, compound II-3 was prepared using intermediate 79 and 2-chloroaniline as raw materials. 1 H NMR (400MHz, DMSO-d6) 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).

[0193] Example 39: Preparation of 8-(4-(4-(6-(2-chlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenyl)piperazin-1-yl)-N-hydroxyoctanamide (II-4)

[0194] Referring to the method of Example 37, the target compound II-4 was prepared using intermediates 85 and 40 as raw materials. 1H NMR(300MHz,DMSO-d6)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=42.8 Hz,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).

[0195] Example 40: Preparation of 8-(4-(6-(2-fluorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)-N-hydroxyoctanamide (II-5)

[0196] Referring to the method of Example 36, the target compound II-5 was prepared using intermediate 5, 2-fluoroaniline and 37 as raw materials. 1 H NMR (300MHz, DMSO-d6) 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,2 H),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).

[0197] Example 41: Preparation of 8-(4-(6-(2-fluorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)-N-hydroxyoctanamide (II-6)

[0198] Referring to the method of Example 36, the target compound II-6 was prepared using intermediate 5, 2-fluoroaniline and 40 as raw materials. 1H NMR (400MHz, DMSO-d6) 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).

[0199] Example 42: Preparation of N-hydroxy-8-(4-(8-methyl-5-oxo-6-(o-tolyl)-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)octanamide (II-7)

[0200] Referring to the method of Example 36, intermediate 5, 2-methylaniline and 37 were used as raw materials to prepare the target compound II-7. LC / MS: m / z = 519.28 [M+H] + . 1 H NMR(400MHz,DMSO-d6)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).

[0201] Example 43: Preparation of N-hydroxy-8-(4-((8-methyl-5-oxo-6-(o-tolyl)-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenyl)piperazin-1-yl)octanamide (II-8)

[0202] Referring to the method of Example 36, the target compound II-8 was prepared using intermediate 5, 2-methylaniline and 40 as raw materials. 1H NMR(400MHz,DMSO-d6)δ10.34(s,1H),9.63(s,1H),8.68(s,1H),8.43(s,1H),7.6 2(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=7.4H z,2H),2.19(s,3H),1.94(t,J=7.3Hz,2H),1.54-1.39(m,4H),1.32-1.21(m,6H).

[0203] Example 44: Preparation of 8-(4-(6-(3-chlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)-N-hydroxyoctanamide (II-9)

[0204] Referring to the method of Example 36, the target compound II-9 was prepared using intermediate 5, 3-chloroaniline and 37 as raw materials. 1 H NMR(400MHz,DMSO-d6)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).

[0205] Example 45: Preparation of 8-(4-(4-(6-(3-chlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenyl)piperazin-1-yl)-N-hydroxyoctanamide (II-10)

[0206] Referring to the method of Example 36, the target compound II-10 was prepared using intermediate 5, 3-chloroaniline and 40 as raw materials. 1H NMR(400MHz,DMSO-d6)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).

[0207] Example 46: Preparation of 8-(4-(6-(4-chlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)-N-hydroxyoctanamide (II-11)

[0208] Referring to the method of Example 36, the target compound II-11 was prepared using intermediate 5, 4-chloroaniline and 37 as raw materials. 1 H NMR(300MHz,DMSO-d6)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).

[0209] Example 47: Preparation of 8-(4-(4-(6-(4-chlorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenyl)piperazin-1-yl)-N-hydroxyoctanamide (II-12)

[0210] Referring to the method of Example 36, the target compound II-12 was prepared using intermediate 5, 4-chloroaniline and 40 as raw materials. 1H NMR(300MHz,DMSO-d6)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=8.9H z,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).

[0211] Example 48: Preparation of 8-(4-(6-(2-chloro-6-fluorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenoxy)-N-hydroxyoctanamide (II-13)

[0212] Referring to the method of Example 36, the target compound II-13 was prepared using intermediate 5, 2-chloro-6-fluoroaniline and 37 as raw materials. 1 H NMR (400MHz, DMSO-d6) 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).

[0213] Example 49: Preparation of 8-(4-(4-(6-(2-chloro-6-fluorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenyl)piperazin-1-yl)-N-hydroxyoctanamide (II-14)

[0214] Referring to the method of Example 36, the target compound II-14 was prepared using intermediate 5, 2-chloro-6-fluoroaniline and 40 as raw materials. 1H NMR(400MHz,DMSO-d6)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=9.0 Hz,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).

[0215] 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-hydroxyoctanamide (II-15)

[0216] Step 1: Synthesis of Intermediate 88

[0217] Triphosgene (9.16 g, 30.87 mmol) and DCM (50 mL) were added to an eggplant-shaped flask, followed by the slow addition of Intermediate 87 (5 g, 30.87 mmol). Triethylamine (9.01 mL, 6.48 mmol) was then added dropwise at -4°C. The mixture was allowed to react at room temperature for 12 hours. Completion of the reaction was monitored by TLC (petroleum ether:ethyl acetate = 10:1). The product was concentrated under reduced pressure to afford 5.80 g of a yellow solid, Intermediate 88, in a 99% yield. This product was used directly in the next step without purification.

[0218] Step 2: Synthesis of Intermediate 90

[0219] To an eggplant-shaped flask was added a solution of intermediate 88 (5.80 g, 30.86 mmol) in diethyl ether (90 mL), and a solution of intermediate 89 (15.27 g, 95.28 mmol) in diethyl ether (90 mL) was added dropwise at 0°C. The mixture was reacted at room temperature for 4 hours. The reaction was monitored by TLC (dichloromethane:methanol=20:1) until completion. The filter cake was filtered and purified by silica gel column chromatography to obtain 8.86 g of a white solid, intermediate 90, with a yield of 82.4%. 1 H NMR(300MHz,DMSO-d6)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).

[0220] Step 3: Synthesis of Intermediate 91

[0221] To an eggplant-shaped flask, intermediate 90 (8.86 g, 25.36 mmol), DCM (100 mL), and a 4 M solution of hydrogen chloride in 1,4-dioxane (100 mL) were added. The mixture was allowed to react at room temperature for 12 hours. Completion of the reaction was monitored by TLC (dichloromethane:methanol = 20:1). The reaction mixture was filtered, and the filter cake was dried under vacuum to afford 7.22 g of a white solid, intermediate 91, in a 100% yield. This product was used directly in the next step without further purification. 1 H NMR (300MHz, DMSO-d6) 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).

[0222] Step 4: Synthesis of Intermediate 92

[0223] To an eggplant-shaped flask were 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). The atmosphere was replaced with nitrogen and the reaction was carried out at 80°C for 3 h. The reaction was monitored by TLC (dichloromethane:methanol = 20:1) until completion. The product was filtered and dried by infrared drying to obtain 9.77 g of a white solid, intermediate 92, with a yield of 86.7%. 1 H NMR (400MHz, DMSO-d6) 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).

[0224] Step 5: Synthesis of Intermediate 93

[0225] Intermediate 92 (9.77 g, 21.99 mmol) and phosphorus oxychloride (100 mL) were added to an eggplant-shaped flask and reacted at 110°C for 12 hours. TLC (petroleum ether:ethyl acetate = 1:1) monitored the reaction for completion. The reaction solution was concentrated under reduced pressure, saturated sodium bicarbonate solution was added at 0°C, and the mixture was filtered. The filter cake was slurried with ethyl acetate, filtered, and dried under IR to obtain 5.86 g of a white solid, Intermediate 93, in a yield of 70.1%. 1H NMR (400MHz, DMSO-d6) 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).

[0226] Step 6: Synthesis of target compound II-15

[0227] Referring to the method of Example 1, intermediates 93 and 37 were used as raw materials to prepare the target compound II-15. 1 H NMR (400MHz, DMSO-d6) 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).

[0228] 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-hydroxyoctanamide (II-16)

[0229] Referring to the method of Example 50, the target compound II-16 was prepared using intermediates 40 and 93 as raw materials. 1 H NMR(400MHz,DMSO-d6)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=7.0H z,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).

[0230] 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-hydroxyoctanamide (II-17)

[0231] Step 1: Synthesis of Intermediate 98

[0232] To an eggplant-shaped flask, starting materials 95 (95.00 g, 658.94 mmol) and 96 (94.21 g, 790.73 mmol) were added and reacted at 80°C for 10 minutes. Intermediate 97 (59.40 g, 658.94 mmol) was then added and reacted at 110°C for 15 hours. The reaction was complete as monitored by TLC (petroleum ether:ethyl acetate = 10:1). The reaction solution was cooled to room temperature, diluted with ethyl acetate, washed with saturated sodium bicarbonate solution and saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. 93.04 g of a white solid, intermediate 98, was obtained by silica gel column chromatography in a yield of 62.4%. 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).

[0233] Step 2: Synthesis of Intermediate 99: To an eggplant-shaped flask, intermediate 98 (93.04 g, 411.15 mmol), lithium hydroxide (19.69 g, 822.31 mmol), tetrahydrofuran (600 mL), and water (600 mL) were added. The mixture was reacted at room temperature for 3 hours. TLC (dichloromethane:methanol = 10:1) monitored the reaction for completion. The reaction solution was concentrated under reduced pressure, the pH was adjusted to acidic with 1 M hydrochloric acid, and the filter cake was dried under vacuum to obtain 44.86 g of a white solid, intermediate 99, in a 100% yield. 1 H NMR (300MHz, DMSO-d6) δ8.90 (s, 1H), 3.08 (q, J = 7.4Hz, 2H), 2.56 (s, 3H), 1.21 (t, J = 7.4Hz, 3H).

[0234] Step 3: Synthesis of Intermediate 100

[0235] To an eggplant-shaped flask, 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) were added. The mixture was allowed to react at room temperature for 3 hours. Completion of the reaction was monitored by TLC (petroleum ether:ethyl acetate = 3:1). 300 mL of water was added to the reaction mixture, which was filtered and the filter cake was dried under vacuum to afford 6.63 g of a yellow solid, Intermediate 100, in a 91.5% yield. 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).

[0236] Step 4: Synthesis of Intermediate 101

[0237] To an eggplant-shaped flask were added intermediate 100 (5.50 g, 19.41 mmol), (chloromethylene)dimethylammonium chloride (7.45 g, 58.23 mmol) and DMF (30 mL). The atmosphere was replaced with nitrogen and the reaction was carried out at 50°C. The reaction was monitored by TLC (dichloromethane:methanol = 35:1) to determine completion. The reaction was quenched by adding saturated sodium bicarbonate solution, followed by the addition of 100 mL of water and filtration. The filter cake was vacuum dried and purified by silica gel column chromatography to obtain 4.00 g of a white solid, intermediate 101, in a yield of 69.3%. 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).

[0238] Step 5: Synthesis of target compound II-17

[0239] Referring to the method of Example 1, intermediates 101 and 40 were used as raw materials to prepare the target compound II-17. 1H NMR(400MHz,DMSO-d6)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,3 H),2.10(s,3H),1.94(t,J=7.3Hz,2H),1.53-1.41(m,4H),1.31-1.21(m,6H).

[0240] 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-hydroxyoctanamide (II-18)

[0241] Referring to the method of Example 1, the target compound II-18 was prepared using intermediates 101 and 37 as raw materials. 1 H NMR (400MHz, DMSO-d6) 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=9.1 Hz,2H),3.94(t,J=6.5Hz,2H),2.21(s,3H),2.10(s,3H),1.95(t,J=7.3 Hz,2H),1.70(p,J=6.6Hz,2H),1.50(p,J=7.4Hz,2H),1.45-1.21(m,2H).

[0242] 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)

[0243] Step 1: Synthesis of Intermediate 103

[0244] By referring to the method of step 2 of Example 52, intermediate 102 (100 mg, 0.19 mmol) was used as starting material to obtain intermediate 103 (80 mg) in a yield of 84.6%. 1H NMR (300MHz, DMSO-d6) 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).

[0245] Step 2: Synthesis of target compound II-19

[0246] Referring to the method of step 3 of Example 52, intermediate 103 (80 mg, 0.16 mmol) and o-phenylenediamine (15 mg, 0.16 mmol) were used as raw materials to prepare the target compound II-19 (38 mg) in a yield of 40.2%. 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).

[0247] 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)

[0248] By referring to the method of step 3 of Example 52, intermediate 103 (100 mg, 0.20 mmol) and p-fluoro-o-phenylenediamine (26 mg, 0.20 mmol) were used as raw materials to prepare the target compound II-20 (92 mg) in a yield of 75.6%. 1H 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).

[0249] Example 56: Preparation of N 1 -(4-(6-(2-chloro-6-fluorophenyl)-8-methyl-5-oxo-5,6,7,8-tetrahydropyrimidin[4,5-d]pyrimidin-2-yl)amino)phenyl)-N 8 -Hydroxyoctanamide (II-21)

[0250] Referring to the method of Example 36, the target compound II-21 was prepared using intermediate 5, 2-chloro-6-fluoroaniline and 4 as raw materials. 1 H NMR (400MHz, DMSO-d6) 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).

[0251] Example 57: Determination of the Inhibitory Activity of Compounds on In vitro Cell Proliferation (H1299)

[0252] 1. Experimental Materials

[0253] H1299 non-small cell lung cancer cell line was purchased from Guangzhou Geneo Biotechnology Co., Ltd. and cultured in RPMI-1640 supplemented with 10% fetal bovine serum (Gibcol) at 37° C., 5% CO 2 , and 95% humidity.

[0254] 2. Experimental steps

[0255] Harvest the cells in the logarithmic growth phase and detect cell viability with trypan blue staining, and carry out cell counting with a cell counter (thermo countness II) to take appropriate cell amount, adjust cell concentration, add 50 μL cell suspensions to 96-well plates, 5000 cells per well, and culture overnight. Prepare test compound solution, add 50 μL test compound solution to each well cell, every kind of test compound final concentration 5 μM, and each compound concentration sets three replicates, and continues to culture in an incubator for 72 hours. After administration and culturing for 72 hours, 10 μL CCK-8 solutions are added to each well, and then 96-well plates are placed in an incubator, and after continuing to culture for 2 hours, absorbance is read at 450 nm and its relative inhibition rate is calculated.

[0256] 3. Experimental results

[0257] The anti-proliferative effects of some compounds on H1299 cell lines are shown in Table 1 below.

[0258] Table 1. Inhibitory activity of some compounds of the present invention on H1299 cell proliferation

[0259] Inhibition rate: 50%-100% (denoted as A), 20%-50% (denoted as B), <20% (denoted as C).

[0260] Example 58: In vitro WEE1 kinase activity inhibition assay

[0261] All test compounds were dissolved in 100% DMSO to a 10 mM stock solution. Store frozen at -20°C in the dark. Prepare 1x kinase buffer: 1x kinase buffer, 50 mM HEPES, pH 7.5, 10 mM MgCl2, 2 mM DTT, 0.01% Tween-20, 0.01% BSA. Dilute the compound to 100x the maximum inhibitor concentration desired in the reaction with 100% DMSO. For a 0.1 μM compound, prepare a 10 μM compound DMSO solution in this step. For a 0.01 μM compound, prepare a 1 μM compound DMSO solution in this step. Add 100 μL of 100% DMSO to two empty wells in the same 96-well plate for the no-compound and no-enzyme controls and label this plate as the source plate. Transfer 40 μL of compound from the source plate to a new 384-well Echo plate as the intermediate plate. Transfer 200 nL from each well of a 384-well Echo plate to a 384-well assay plate. Prepare a WEE1 inhibitor solution in 1x kinase buffer at a concentration twice the final concentration of each reagent in the assay. Add 10 μL of kinase solution to each well of the assay plate, except for control wells without enzyme (instead, add 10 μL of 1x kinase buffer). Shake the plate. Incubate at room temperature for 10 minutes. Prepare substrate solutions of fluorescein-polyGT and ATP in 1x kinase reaction buffer at a concentration twice the final concentration of each reagent in the assay. Add 10 μL of substrate solution to each well of the assay plate to initiate the reaction. Shake the plate. Cover the assay plate and incubate at room temperature for the desired reaction time. Prepare a detection solution at 2x the final concentration in antibody dilution buffer. Add 20 μL of detection solution to each well of the assay plate to stop the reaction. Mix briefly by centrifuging and incubate at room temperature for 60 minutes before reading fluorescence on a microplate reader. Data were collected on an Envision at excitation at 340 nm and emission at 520 nm and 495 nm.

[0262] The RFU values ​​were copied from the Envision program. The ratio of RFU 520 nm / RFU 495 nm was calculated. The ratio values ​​were converted to percent inhibition values. Percent inhibition = (max - sample ratio) / (max - min) * 100. "Min" refers to the ratio of the no enzyme control, and "max" refers to the ratio of the DMSO control. Data were presented in MS Excel, and curves were fitted using the XLFit Excel plug-in version 5.4.0.8. IC 50 Calculation formula: Y = Bottom + (Top - Bottom) / (1 + (IC 50 / X)^HillSlope.

[0263] Example 59: In vitro HDAC1 enzyme activity inhibition assay

[0264] Prepare 1x assay buffer (modified Tris buffer). Compound serial dilution: Transfer the compounds to the assay plate in 100% DMSO by Echo. The final fraction of DMSO is 1%. Prepare the enzyme solution in 1x assay buffer. Prepare the substrate solution by adding trypsin and Ac-peptide substrate to 1x assay buffer. Transfer 15 μL of enzyme solution to the assay plate, or for low concentration controls, transfer 15 μL of 1x assay buffer. Incubate at room temperature for 15 minutes. Add 10 μL of substrate solution to each well to start the reaction. Read the plate on Envision with excitation at 355 nm and emission at 460 nm. Curve fitting: Fit the data in Excel and calculate the inhibition rate: Inh% = (Max-Signal) / (Max-Min) * 100, fit the data in XL-Fit and calculate the IC 50 Value: Y=Bottom+(Top-Bottom) / (1+(IC 50 = / X)*HillSlope), where Y is the inhibition rate and X is the compound concentration. The results of in vitro enzyme inhibition are shown in Table 2.

[0265] Table 2. Percent inhibition test results of some compounds of the present invention on WEE1 kinase and HDAC1 enzyme

[0266] From the above activity results, it can be seen that most of the compounds designed in the present invention have strong inhibitory effects on WEE1 kinase and HDAC1 enzyme, and have considerable anti-tumor cell proliferation activity, and can be used to prepare drugs for treating diseases caused by overexpression of WEE1 kinase and / or HDAC enzyme.

Claims

1. An anilinopyrimido heterocyclic compound, characterized in that: The structure of formula I also includes pharmaceutically acceptable salts thereof: in: R1 is selected from substituted or unsubstituted C 1-4 Alkyl, C 2-4 alkenyl, 5-7 membered aryl, 5-7 membered heteroaryl containing 1-3 N, O, S, wherein the substituent is selected from one or more hydrogen, halogen, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, cyano, nitro, hydroxyl, amino; R2 is selected from one or more of hydrogen, halogen, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, cyano, nitro, hydroxyl, amino; R3 is selected from hydroxyl, substituted or unsubstituted 5-7 membered aryl, 5-7 membered heteroaryl containing 1-3 N, O, S, and the substituent is selected from one or more hydrogen, halogen, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, cyano, nitro, hydroxyl, amino; X is selected from Where R 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, hydroxyl, Q is selected from CH, N, R b is selected from substituted or unsubstituted C 1-4 Alkyl, C 2-4 alkenyl, 5-7 membered aryl, 5-7 membered aromatic heteroyl containing 1-3 N, O, S, wherein the substituent is selected from one or more hydrogen, halogen, C 1-6 Alkyl, C 1-6 Haloalkyl, C 1-6 Alkoxy, cyano, nitro, hydroxyl, amino; Y is selected from -(CH2) n -、-(CH2) n O-, -(CH2) n S-, -(CH2) n CONH-、-(CH2) n NHCO-, -(CH2) n SO2NH-、-(CH2) n NHSO2-, Where n=0-7.

2. The anilinopyrimido heterocyclic compound according to claim 1, characterized in that: In the structure: R1 is selected from substituted or unsubstituted C 1-4 Alkyl, C 2-4 alkenyl, phenyl, the substituent being selected from one or more of hydrogen, halogen, C 1-4 Alkyl, C 1-4 Haloalkyl, C 1-4 Alkoxy; R2 is selected from one or more of hydrogen, halogen, C 1-4 alkyl; R3 is selected from hydroxyl, substituted phenyl, and the substituent is selected from halogen, amino; X is selected from Where R 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 alkyl; Y is selected from -(CH2) n -、-(CH2) n O-, -(CH2) n CONH-, Where n=0-7.

3. The anilinopyrimido heterocyclic compound according to claim 1, characterized in that: In the structure: R1 is selected from R2 is selected from one or more of hydrogen, methyl, halogen; R3 is selected from hydroxy, o-aminophenyl, 2-amino-4-fluorophenyl; X is selected from Y is selected from -(CH2) n -、-(CH2) n O-, -(CH2) n CONH-, Where n=0-7.

4. The anilinopyrimido heterocyclic compound according to claim 1, characterized in that: In the structure: R1 is selected from R2 is selected from hydrogen; R3 is selected from hydroxyl; X is selected from Y is selected from -(CH2) n O-, -(CH2) n CONH-, Where n=6,7.

5. The anilinopyrimido heterocyclic compound according to claim 1, characterized in that: Select any of the following compounds:

6. The anilinopyrimido heterocyclic compound according to claim 1, characterized in that: The pharmaceutically acceptable salt is a salt formed by the compound and any one of the following acids: 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, and ferulic acid.

7. A method for preparing anilinopyrimido heterocyclic compound according to claim 1, characterized in that: Choose from any of the following methods: (1) When R3 is a hydroxyl group, the methyl mercapto compound is coupled and aminolyzed to obtain the compound I: (2) When R3 is not a hydroxyl group, the methyl mercapto compound is coupled, hydrolyzed, and acylated to obtain the compound I: Wherein, R1, R2, R3, X, and Y are as defined in claim 1, R c is methyl or ethyl; The corresponding acid is reacted with the compound I prepared by the above method to form a salt, thereby obtaining a pharmaceutically acceptable salt of the compound I.

8. A pharmaceutical composition, characterized in that It comprises the anilinopyrimido heterocyclic compound according to claim 1 and a pharmaceutically acceptable carrier.

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

10. The use according to claim 9, characterized in that: The drug is an anti-tumor drug.

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