2, 4, 7-trisubstituted quinazoline derivative, synthetic method and application

Abstract

The invention discloses a 2, 4, 7-trisubstituted quinazoline derivative as well as a synthesis method and application thereof, and belongs to the field of organic synthesis and medicinal chemistry. The compound has an innovative molecular structure, can simultaneously inhibit histone lysine demethylase (KDM) and histone deacetylase (HDAC) in a targeted manner, and represents a novel double-target epigenetic regulating agent. The preparation method adopts a modular synthesis strategy, constructs a core structure through a three-step continuous functionalization reaction, has the characteristics of simple and convenient process, easily available raw materials, mild reaction conditions and the like, and is particularly suitable for industrial enlarged production. In-vitro activity evaluation shows that part of the compounds have nanomole-level inhibitory activity on double targets, can be used as lead compounds for developing novel antitumor drugs after structural modification and pharmacophore optimization, and show important application value in the field of epigenetic targeted therapy.

Description

Technical Field

[0001] The present application relates to 2,4,7-trisubstituted quinazoline derivatives, synthesis methods, applications, and drugs, and belongs to the field of chemistry and medicine.

[0002] Background Field

[0003] Histone methylation primarily occurs on lysine or arginine residues of histones H3 and H4. Lysine can be unmodified, monomethylated, dimethylated, or trimethylated; arginine can be unmodified, monomethylated, symmetrically dimethylated, or asymmetrically dimethylated. Furthermore, methylation can occur at various sites. The complexity of methylation states and sites allows for flexible regulation of gene transcriptional activation and repression. Methylation is catalyzed by histone methyltransferases (HMTs), and the process can also be reversed by demethylases. However, relatively little research has been conducted on arginine demethylation, with research primarily focusing on lysine demethylases (KDMs).

[0004] KDMs are divided into two categories. One is the lysine-specific demethylases (LSDs), including LSD1 and LSD2, which primarily catalyze the demethylation of mono- and dimethylated H3K4 and H3K9. The other is the Jumonji C domain-containing (JmjC domain-containing, JMJD) family, which belongs to the 2-oxoglutarate (2-OG) and ferrous ion-dependent dioxygenase superfamily and catalyzes the demethylation of mono-, di-, and tri-methylated lysine residues at various histone sites. The JMJD family includes KDM2, KDM3, KDM4, KDM5, and KDM6. Numerous studies have demonstrated that KDMs are involved in the regulation of DNA replication and DNA damage repair and are associated with the development and progression of cancer. Genetic deletion or drug inhibition of KDMs can achieve tumor suppressive effects, and KDMs have been identified as potential targets for cancer therapy.

[0005] Histone deacetylases (HDACs) are a family of epigenetic enzymes that regulate gene expression and cell function by catalyzing the deacetylation modification of histone and non-histone substrates. Based on their structure, subcellular localization, and cofactor dependence, the HDAC family can be divided into four categories: Class I (HDAC1, 2, 3, 8), Class II (IIa: HDAC4, 5, 7, 9; IIb: HDAC6, 10), Class III (Sirtuins family, SIRT1-7), and Class IV (HDAC11). Among them, Class I, Class II, and Class IV belong to the classic Zn 2+ dependent enzymes, while class III Sirtuins are NAD dependent+ As a cofactor, HDAC's primary function is to remove acetyl groups from histones H3 and H4, leading to chromatin compaction and inhibition of gene transcription. Furthermore, HDAC regulates the acetylation status of non-histone proteins, such as p53, STAT3, HSP90, and α-tubulin, affecting protein stability, localization, and signaling pathway activity, thereby participating in processes such as the cell cycle, apoptosis, metabolism, and immune response.

[0006] Studies have shown that combined inhibition of KDM and HDAC significantly improves therapeutic efficacy in malignancies such as breast cancer, surpassing monotherapy. Therefore, KDM and HDAC are closely linked, and the combined use of KDM and HDAC inhibitors has yielded improved therapeutic effects in some diseases, providing guidance for the development of dual-target inhibitors.

[0007] The present invention designs and synthesizes a series of 2,4,7-trisubstituted quinazoline derivatives, whose structures are different from the KDM inhibitors and HDAC inhibitors reported in previous studies, and belong to a new type of inhibitor structure. Through structural modification and in-depth pharmacological studies on the compounds of the present invention, it is expected that new KDM / HDAC dual-target inhibitors will be developed. Summary of the Invention

[0008] In view of the shortcomings of the existing technology, the present invention provides a 2,4,7-trisubstituted quinazoline-based dual-target inhibitor of lysine demethylase and histone deacetylase, and also provides a preparation method and application of the compound.

[0009] The technical solutions of the present invention are as follows:

[0010] 1. 2,4,7-Trisubstituted quinazoline-based dual-target inhibitors of lysine demethylase and histone deacetylase

[0011] A 2,4,7-trisubstituted quinazoline-based dual-target inhibitor of lysine demethylase and histone deacetylase, or a pharmaceutically acceptable salt, ester or prodrug thereof, having the following general structural formula I:

[0012]

[0013] wherein R1 is independently selected from: a carboxyl group or a hydroxamic acid group;

[0014] n=2-8;

[0015] The R2 is independently selected from: C5-C10 aryl group, mono- or poly-substituted C5-C10 aryl group, dioxaspiro substituent, and halogen atom.

[0016] The R3 is independently selected from: N-methylpiperazinyl, C5-C10 heterocycle containing N / O / S, aliphatic amine, C5-C10 aryl, mono- or poly-substituted C5-C10 aryl, halogen atom, olefin, C5-C8 cyclic olefin.

[0017] Preferably, the compound of the above general formula I is one of the following

[0018]

[0019]

[0020]

[0021] 2. Preparation method of 2,4,7-trisubstituted quinazoline KDM / HDAC dual-target inhibitors

[0022] The commercially available raw material 2,4-dichloro-7-bromoquinazoline is used as a raw material. First, it undergoes a nucleophilic substitution reaction with a chain amine under an alkaline environment to obtain a 4-monosubstituted quinazoline intermediate; the 4-monosubstituted quinazoline intermediate undergoes a Suzuki coupling reaction to modify the 7-position to generate a 4,7-disubstituted quinazoline intermediate; the 4,7-disubstituted quinazoline intermediate can be optionally subjected to a Suzuki coupling reaction with a substituted boronic acid, or undergoes a nucleophilic substitution reaction with various amines or heterocycles, thereby obtaining a 2,4,7-trisubstituted quinazoline intermediate; the 2,4,7-trisubstituted quinazoline intermediate undergoes a hydrolysis reaction under strong alkaline conditions or undergoes a hydroxylamine hydroximation reaction under alkaline conditions to obtain a derivative I.

[0023] Synthesis route such as Figure 1 shown.

[0024] (a) N,N-diisopropylethylamine, tetrahydrofuran, 60°C, 2h;

[0025] (b) Methanesulfonyloxy(diadamantyl-n-butylphosphino)-2'-amino-1,1'-biphenyl-2-yl)palladium(II), potassium phosphate, 1,4-dioxane, 80°C, 12h;

[0026] (c) N,N-diisopropylethylamine, cuprous iodide, 1,4-dioxane, 80°C, 12h;

[0027] (d) Methanesulfonyloxy(diadamantyl-n-butylphosphino)-2'-amino-1,1'-biphenyl-2-yl)palladium(II), potassium phosphate, 1,4-dioxane, 100°C, 12h;

[0028] (e) sodium hydroxide, methanol, room temperature, 3 h;

[0029] (f) Hydroxylamine aqueous solution, potassium hydroxide, methanol, room temperature, 2h.

[0030] According to a preferred method of the present invention, the preparation method of 2,4,7-trisubstituted quinazoline derivatives comprises the following steps:

[0031] (1) 1 mmol of 2,4-dichloro-7-bromoquinazoline was added to a 50 mL round-bottom flask and dissolved in 5 mL of tetrahydrofuran. The mixture was stirred and 1.1 mmol of aminoalkylcarboxylic acid methyl ester hydrochloride and 2.4 mmol of N,N-diisopropylethylamine were added in sequence. The mixture was refluxed at 65°C for 3 h. After the reaction was completed by TLC monitoring, 150 mL of ethyl acetate was added for dilution. The organic phase was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL), and dried over anhydrous sodium sulfate. After removing the solvent by distillation under reduced pressure, a light yellow oily intermediate 1 (crude product) was obtained, which was used directly in the subsequent reaction step without purification.

[0032] (2) 1 mmol of 1 was placed in a 100 mL thick-walled pressure-resistant bottle and dissolved in 5 mL of 1,4-dioxane. 1.5 mmol of substituted boronic acid, 2.5 mmol of potassium phosphate, and 0.1 mmol of methanesulfonyloxy(diadamantyl-n-butylphosphino)-2'-amino-1,1'-biphenyl-2-yl)palladium(II) were added. The mixture was refluxed at 80°C for 12 h under a nitrogen atmosphere. The reaction was monitored by TLC. 150 mL of ethyl acetate was added for dilution. The organic phase was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL), and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure, and the mixture was purified by silica gel column chromatography (DCM:EA=30:1) to obtain a yellow solid 2.

[0033] (3) 1 mmol of 1 was added to a 50 mL round-bottom flask, dissolved in 5 mL of 1,4-dioxane, and then 10 mmol of N-methylpiperazine, 2.4 mmol of N,N-diisopropylethylamine, and 1 mmol of cuprous iodide were added. The mixture was refluxed at 85°C for 12 h. After TLC monitoring, the reaction was completed. 150 mL of ethyl acetate was poured into the system, and the mixture was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL) and dried over anhydrous sodium sulfate. The solvent was removed by vacuum distillation, and the mixture was purified by silica gel column chromatography (DCM:MeOH=30:1) to obtain a yellow solid 4.

[0034] (4) 1 mmol of compound 2 was added to a 50 mL round-bottom flask and dissolved in 5 mL of 1,4-dioxane. 10 mmol of a substituted amine, 2.4 mmol of N,N-diisopropylethylamine, and 1 mmol of cuprous iodide were added, and the mixture was refluxed at 85°C for 12 h. After TLC monitoring, the reaction was completed. 150 mL of ethyl acetate was poured into the system, and the mixture was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL) and dried over anhydrous sodium sulfate. The solvent was removed by vacuum distillation, and the mixture was purified by silica gel column chromatography (DCM:MeOH=30:1) to obtain a yellow solid 3.

[0035] (5) 1 mmol of 2 was placed in a 100 mL thick-walled pressure-resistant bottle and dissolved in 5 mL of 1,4-dioxane. 1.5 mmol of substituted boronic acid, 2.5 mmol of potassium phosphate, and 0.1 mmol of methanesulfonyloxy(diadamantyl-n-butylphosphino)-2'-amino-1,1'-biphenyl-2-yl)palladium(II) were added. The mixture was refluxed at 100°C for 12 h under a nitrogen atmosphere. The reaction was monitored by TLC. 150 mL of ethyl acetate was added for dilution. The organic phase was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL), and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure, and the mixture was purified by silica gel column chromatography (DCM:EA=30:1) to obtain a yellow solid 3.

[0036] (6) 1 mmol of the esterified intermediate 3 or 2 was added to a 50 mL round-bottom flask, followed by the addition of 5 mL of methanol and 1 mL of a 2N aqueous sodium hydroxide solution. The mixture was stirred at room temperature for 4 h. After completion of the reaction, the pH was adjusted to a weakly acidic state using 1N hydrochloric acid. The reaction was allowed to stand until a solid precipitated. The precipitate was filtered, washed with water, and recrystallized from ethanol to obtain the target compounds A01-A14.

[0037] (7) 1 mmol of the esterified intermediate 3, 2, or 4 was added to a 50 mL round-bottom flask, followed by the addition of 5 mL of methanol, 30 mmol of aqueous hydroxylamine solution, and 4 mmol of potassium hydroxide. The mixture was stirred at room temperature for 4 h. After completion of the reaction, the pH was adjusted to a weakly acidic state using 1 N hydrochloric acid. The solvent was evaporated under reduced pressure and purified by silica gel column chromatography (DCM:MeOH = 5:1) to obtain the target compounds A15-A37.

[0038] The room temperature described in the present invention refers to 20-30°C.

[0039] III. KDM and HDAC Inhibitory Activities of 2,4,7-Trisubstituted Quinazoline Derivatives

[0040] Several 2,4,7-trisubstituted quinazoline derivatives synthesized using the above method were tested for their inhibitory activity, and the results showed that most compounds exhibited significant inhibitory activity against both KDM and HDAC. Therefore, 2,4,7-trisubstituted quinazoline derivatives are worthy of further development as dual-target inhibitors for the treatment of various malignancies mediated by KDM or HDAC.

[0041] The present invention will be further described below with reference to the accompanying drawings and specific embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Figure 1 The synthetic route of the present invention is shown in FIG. DETAILED DESCRIPTION

[0043] The following examples are provided to help understand the present invention, but they are not intended to limit the present invention.

[0044] Example 1: 3-((2-(4-methoxyphenyl))-7-(4-vinylphenyl)quinazolin-4-yl)amino)propanoic acid (A01)

[0045] 1 mmol of 2,4-dichloro-7-bromoquinazoline was added to a 50 mL round-bottom flask and dissolved in 5 mL of tetrahydrofuran. The mixture was stirred and then 1.1 mmol of methyl 3-aminopropionate hydrochloride and 2.4 mmol of N,N-diisopropylethylamine were added sequentially. The mixture was refluxed at 65°C for 3 h. After TLC monitoring, the reaction was diluted with 150 mL of ethyl acetate. The organic phase was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL) and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure to obtain intermediate 1 (crude product) as a pale yellow oil, which was used directly in the subsequent reaction step without purification.

[0046] 1 mmol of 1 was placed in a 100 mL thick-walled pressure bottle and dissolved in 5 mL of 1,4-dioxane. 1.5 mmol of 4-vinylphenylboronic acid, 2.5 mmol of potassium phosphate, and 0.1 mmol of methanesulfonyloxy(diadamantyl-n-butylphosphino)-2'-amino-1,1'-biphenyl-2-yl)palladium(II) were added. The mixture was refluxed at 80°C for 12 h under a nitrogen atmosphere. TLC monitored the reaction completion. The reaction was diluted with 150 mL of ethyl acetate, washed with water (3 × 30 mL), then with saturated brine (3 × 30 mL), and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure, and the mixture was purified by silica gel column chromatography (DCM:EA = 30:1) to obtain 2 as a yellow solid.

[0047] 1 mmol of compound 2 was placed in a 100 mL thick-walled pressure bottle and dissolved in 5 mL of 1,4-dioxane. 1.5 mmol of 4-methoxyphenylboronic acid, 2.5 mmol of potassium phosphate, and 0.1 mmol of methanesulfonyloxy(diadamantyl-n-butylphosphino)-2'-amino-1,1'-biphenyl-2-yl)palladium(II) were added. The mixture was refluxed at 100°C for 12 h under a nitrogen atmosphere. TLC monitored the reaction completion. The reaction was diluted with 150 mL of ethyl acetate, washed with water (3 × 30 mL), then with saturated brine (3 × 30 mL), and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure, and the mixture was purified by silica gel column chromatography (DCM:EA = 30:1) to obtain 3 as a yellow solid.

[0048] Add 1 mmol of the esterified intermediate 3 to a 50 mL round-bottom flask, followed by 5 mL of methanol and 1 mL of 2N sodium hydroxide solution. Stir at room temperature for 4 hours. After completion of the reaction, monitor the reaction by TLC. Adjust the pH to a weakly acidic state with 1N hydrochloric acid. Allow the reaction to stand until a solid precipitates. Filter the precipitate, wash it with water, and recrystallize it from ethanol to obtain the target compound A01. This product is a white solid with a 50% yield and an MP of 178–179°C. 1 H NMR(400MHz,DMSO-d6,ppm)δ10.21(t,J=5.3Hz,1H),8.50–8.48(m,2H),8.44(d, J=8.6Hz,2H),7.93(d,J=8.6Hz,1H),7.72(d,J=8.0Hz,2H),7.57(d,J=8.1Hz,2H) ,7.10(d,J=8.6Hz,2H),6.77(dd,J=17.6,10.9Hz,1H),5.91(d,J=17.7Hz,1H),5 .34(d,J=10.9Hz,1H),3.94(q,J=6.6Hz,2H),3.84(s,3H),2.80(t,J=7.0Hz,2H). 13 C NMR (100MHz, DMSO-d6, ppm) δ173.1,164.0,159.7,156.8,146.0,140.3,138.5,137.1,136.3,131. 7,127.7,127.4,126.1,125.3,123.4,116.9,116.1,114.8,111.2,56.1,38.3,33.3.HRMS-ESI:m / z calcd for C 26 H 24 N3O3[M+H] + 426.1812, found 426.1816.

[0049] Example 2: Preparation of 4-((2-(4-methoxyphenyl))-7-(4-vinylphenyl)quinazolin-4-yl)amino)butyric acid (A02)

[0050] A02 was prepared using the same method as A01, except that 4-aminobutyric acid methyl ester hydrochloride was used instead of 3-aminopropionic acid methyl ester hydrochloride. White solid; 55% yield; MP: 156–157°C; 1 H NMR (400MHz, DMSO-d6, ppm) δ12.19(s,1H),9.53(s,1H),8.50(d,J=8.7Hz,2H),8.46(d,J=8. 8Hz,1H),8.32(s,1H),7.92(dd,J=8.5,1.8Hz,1H),7.80(d,J=8.1Hz,2H),7.63(d,J=8.1Hz,2 H),7.12(d,J=8.7Hz,2H),6.80(dd,J=17.6,10.9Hz,1H),5.94(d,J=17.7Hz,1H),5.35(d,J=1 0.9Hz, 1H), 3.86 (s, 3H), 3.77 (q, J = 6.7Hz, 2H), 2.41 (t, J = 7.2Hz, 2H), 1.99 (p, J = 7.2Hz, 2H). 13 C NMR (100MHz, DMSO-d6, ppm) δ174.3,162.6,162.6,159.5,157.6,144.8,137.7,137.5,136.0,130.6,1 27.3,127.1,127.0,124.9,124.9,124.4,115.4,114.1,111.6,55.6,40.7,31.2,23.9.HRMS-ESI:m / z calcd for C 27 H 26 N3O3[M+H] + 440.1969,found 440.1969.

[0051] Example 3: Preparation of 5-((2-(4-methoxyphenyl))-7-(4-vinylphenyl)quinazoline-4-amino)pentanoic acid (A03)

[0052] A03 was prepared using the same method as A01, except that 5-aminovaleric acid methyl ester hydrochloride was used instead of 3-aminopropionic acid methyl ester hydrochloride. White solid; 52% yield; MP: 155–156°C; 1H NMR (400MHz, DMSO-d6, ppm) δ12.12 (s, 1H), 10.01 (s, 1H), 8.54 (d, J = 8.7Hz, 1H), 8.52-8.49 (ove rlap,3H),7.99(dd,J=8.6,1.8Hz,1H),7.80(d,J=8.1Hz,2H),7.65(d,J=8.1Hz,2H),7.19-7.14 (m,2H),6.81(dd,J=17.6,10.9Hz,1H),5.96(d,J=17.7Hz,1H),5.37(d,J=11.0Hz,1H),3.88(s, 3H), 3.77 (q, J = 6.6Hz, 2H), 2.32 (t, J = 7.2Hz, 2H), 1.78 (p, J = 7.0Hz, 2H), 1.66 (q, J = 7.3Hz, 2H). 13 CNMR(100MHz,DMSO-d6,ppm)δ174.4,163.3,163.3,159.4,156.9,145.4,137.9,137.1,136.0,131.0,127.3,12 7.2,127.1,125.5,125.5,124.7,115.6,114.4,111.2,55.7,41.3,40.1,39.9,33.3,27.8,22.1.HRMS-ESI:m / z calcd for C 28 H 28 N3O3[M+H] + 454.2125,found454.2121.

[0053] Example 4: Preparation of 3-((7-(4-methoxyphenyl))-2-(2-nitrophenyl)quinazolin-4-yl)amino)propionic acid (A04)

[0054] A04 was prepared using the same method as A01, except that 4-nitrophenylboronic acid was used instead of 4-methoxyphenylboronic acid to obtain the 4,7-disubstituted quinazoline intermediate 2, which was then replaced with 2-nitrophenylboronic acid to obtain the 2,4,7-trisubstituted quinazoline intermediate 3. Yellow solid; 35% yield; MP: 138–139°C. 1H NMR (400MHz, DMSO-d6, ppm) δ12.34(s,1H),9.85(s,1H),8.56(d,J=8.7Hz,1H),8.16(dd,J=7.7,1.5Hz,1H),8.14–8.09(m,2H),8.05(dd,J=8.7,1.9Hz ,1H),7.95–7.92(m,1H),7.86(dd,J=7.8,1.5Hz,1H),7.83–7.79(m,2H),7. 13–7.09(m,2H),3.83(s,3H),3.73(t,J=6.4Hz,2H),2.66(t,J=7.0Hz,2H). 13 CNMR(100MHz,DMSO-d6,ppm)δ172.6,160.3,159.5,157.3,149.0,145.8,145.8,133.1,132.2,132. 1,131.6,130.1,128.6,126.1,126.1,124.6,124.4,114.8,111.0,55.4,37.5,32.8.HRMS-ESI:m / z calcd for C 24 H 21 N4O5[M+H] + 445.1506,found445.1506.

[0055] Example 5: Preparation of 4-((7-(4-methoxyphenyl))-2-(2-nitrophenyl)quinazolin-4-yl)amino)butyric acid (A05)

[0056] A05 is prepared using the same method as A02, except that 4-nitrophenylboronic acid is used instead of 4-methoxyphenylboronic acid to obtain the 4,7-disubstituted quinazoline intermediate 2, which is then replaced with 2-nitrophenylboronic acid to obtain the 2,4,7-trisubstituted quinazoline intermediate 3. Yellow solid; 29% yield; MP: 145°C. 1 H NMR(400MHz,DMSO-d6,ppm)δ10.40(s,1H),8.68(d,J=8.8Hz,1H),8.24–8.15(m,3H),8.11(dd,J=8.7,1.8Hz,1H),8.0–7.95(m,1H),7.9 4–7.90(m,1H),7.82–7.77(m,2H),7.15–7.11(m,2H),3.83(s,3H),3.61(d,J=6.5Hz,2H),2.34(t,J=7.3Hz,2H),1.88(q,J=7.2Hz,2H). 13C NMR (100MHz, DMSO-d6, ppm) δ174.1,160.5,159.6,156.7,148.6,148.6,146.4,133.5,132.9,131.7,131 .7,129.7,128.6,126.6,125.1,124.7,124.7,114.9,110.6,55.4,41.1,31.1,23.8.HRMS-ESI:m / zcalcd for C 25 H 23 N4O5[M+H] + 459.1663, found 459.1662.

[0057] Example 6: Preparation of 5-((7-(4-methoxyphenyl))-2-(2-nitrophenyl)quinazolin-4-yl)amino)pentanoic acid (A06)

[0058] A06 is prepared using the same method as A03, except that 4-nitrophenylboronic acid is used instead of 4-methoxyphenylboronic acid to obtain the 4,7-disubstituted quinazoline intermediate 2, which is then replaced with 2-nitrophenylboronic acid to obtain the 2,4,7-trisubstituted quinazoline intermediate 3. Yellow solid; 37% yield; MP: 145–147°C. 1 H NMR (400MHz, DMSO-d6, ppm) δ10.47(s,1H),8.69(d,J=8.8Hz,1H),8.24(d,J=1.9Hz, 1H), 8.19 (ddd, J=14.0, 7.8, 1.4Hz, 2H), 8.11 (dd, J=8.7, 1.8Hz, 1H), 7.99 (td, J=7. 6,1.3Hz,1H),7.92(td,J=7.7,1.5Hz,1H),7.83–7.78(m,2H),7.13(d,J=8.9Hz,2H) ,3.84(s,3H),3.60–3.57(t,J=5.8Hz,2H),2.26(t,J=7.2Hz,2H),1.68–1.54(m,4H). 13 C NMR (100MHz, DMSO-d6, ppm) δ174.3,160.5,159.4,156.6,148.6,148.6,146.5,133.4,132.9,131.8,131 .8,129.7,128.6,126.6,125.1,124.8,124.8,114.9,110.6,55.4,41.4,33.3,27.8,22.0.HRMS-ESI:m / z calcd for C 26 H25 N4O5[M+H] + 473.1819, found 473.1817.

[0059] Example 7: Preparation of 5-((7-(4-methoxyphenyl))-2-(propylamino)quinazolin-4-yl)amino)pentanoic acid (A07)

[0060] The preparation method for A07 is similar to that for A06, except that after obtaining the 4,7-disubstituted quinazoline intermediate 2, 1 mmol of 2 is added to a 50 mL round-bottom flask and dissolved in 5 mL of 1,4-dioxane. 10 mmol of n-propylamine, 2.4 mmol of N,N-diisopropylethylamine, and 1 mmol of cuprous iodide are then added, and the mixture is refluxed at 85°C for 12 h. Upon completion of the reaction, TLC is monitored. 150 mL of ethyl acetate is poured into the system, and the system is washed sequentially with water (3 × 30 mL) and saturated brine (3 × 30 mL), then dried over anhydrous sodium sulfate. The solvent is removed by distillation under reduced pressure, and the product is purified by silica gel column chromatography (DCM:MeOH = 30:1) to yield 3 as a yellow solid.

[0061] 1 mmol of the esterified intermediate 3 was added to a 50 mL round-bottom flask. 5 mL of methanol and 1 mL of 2N sodium hydroxide solution were then added, and the mixture was stirred at room temperature for 4 h. After completion of the reaction, the pH was adjusted to a weakly acidic state with 1N hydrochloric acid. The reaction was allowed to stand for precipitation of a solid, which was filtered, washed with water, and recrystallized from ethanol to yield the target compound A07. This product was a brown solid with a 25% yield and an MPa of 221–223°C. 1 H NMR(400MHz,DMSO-d6,ppm)δ9.76(t,J=6.1Hz,1H),8.36(d,J=8.6Hz,1H),8.30–8.17(m,1H),7.73–7.60(m,3H),7.54(s,1H) ,7.06(d,J=8.4Hz,2H),3.80(s,3H),3.55(t,J=6.6Hz,4H),2.27(t,J=7.3Hz,2H),1.70–1.53(m,6H),0.92(t,J=7.5Hz,3H). 13 CNMR(100MHz,DMSO-d6,ppm)δ174.4,160.1,159.5,153.2,145.8,139.5,130.1,128.3,125.0,1 22.1,114.7,112.8,108.0,55.4,42.4,40.9,33.4,27.7,22.3,22.1,11.3.HRMS-ESI:m / zcalcd for C 23 H 29 N4O3[M+H]+ 409.2234.3578, found 409.2236.

[0062] Example 8: Preparation of 5-((2-(cyclohexylamino)-7-(4-methoxyphenyl)quinazolin-4-yl)amino)pentanoic acid (A08) The preparation method of A08 was the same as A07, except that cyclohexylamine was used instead of n-propylamine. Brown solid; 30% yield; Mp: 200–202°C; 1 H NMR (400MHz, DMSO-d6, ppm) δ9.72(s,1H),8.32(s,1H),8.25(s,1H),7.69(d,J=8.2Hz,2H),7.50(s,1H),7.06(d,J=8.1Hz,2H),3 .81(s,3H),3.59–3.54(m,2H),2.29–2.26(m,2H),1.91(m,2H),1.77–1.64(overlap,5H),1.60–1.56(m,4H),1.38–1.33(m,4H). 13 CNMR(100MHz,DMSO-d6,ppm)δ174.8,160.6,159.9,152.8,146.3,139.7,130.5,128.8,125.4,122 .6,115.1,113.2,108.4,55.8,50.3,41.4,33.8,32.4,28.2,25.5,24.6,22.5.HRMS-ESI:m / zcalcd for C 26 H 33 N4O3[M+H] + 449.2547, found 449.2545.

[0063] Example 9: Preparation of 4-((2-(dimethylamino)-7-(4-methoxyphenyl)quinazolin-4-yl)amino)butanoic acid (A09) The preparation method of A09 was similar to that of A05, except that after obtaining the 4,7-disubstituted quinazoline intermediate 2, 1 mmol of 2 was added to a 50 mL round-bottom flask and dissolved in 5 mL of 1,4-dioxane. 10 mmol of dimethylamine hydrochloride, 2.4 mmol of N,N-diisopropylethylamine, and 1 mmol of cuprous iodide were added, and the mixture was refluxed at 85°C for 12 h. The reaction was monitored for completion by TLC. 150 mL of ethyl acetate was poured into the system, and the mixture was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL), and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure, and the mixture was purified by silica gel column chromatography (DCM:MeOH = 30:1) to obtain 3 as a yellow solid.

[0064] 1 mmol of the esterified intermediate 3 was added to a 50 mL round-bottom flask. 5 mL of methanol and 1 mL of 2N sodium hydroxide solution were then added, and the mixture was stirred at room temperature for 4 h. After completion of the reaction, the pH was adjusted to a weakly acidic state with 1N hydrochloric acid. The reaction was allowed to stand for precipitation of a solid, which was filtered, washed with water, and recrystallized from ethanol to yield the target compound A09. This yellow solid was obtained in a 37% yield and had an MP of 280–282°C. 1 H NMR (400MHz, DMSO-d6, ppm) δ9.59(d,J=5.9Hz,1H),8.34(d,J=8.6Hz,1H),8.25(s,1H),7.67(dd,J=13.8,7.7Hz, 3H),7.07(d,J=8.4Hz,2H),3.81(s,3H),3.59(q,J=6.6Hz,2H),3.28(s,6H),2.33(t,J=7.1Hz,2H),1.89(m,2H). 13 C NMR(100MHz,DMSO-d6,ppm)δ174.2,160.1,158.6,152.5,145.2,140.5,130.2,128 .2,124.7,122.1,114.7,114.1,107.8,55.4,40.6,37.9,31.1,23.6.HRMS-ESI:m / z calcd for C 21 H 25 N4O3[M+H] + 381.1921,found381.1918.

[0065] Example 10: Preparation of 4-((2-(dimethylamino)-7-(4-methoxyphenyl)quinazolin-4-yl)amino)propanoic acid (A10)

[0066] A10 was prepared using the same method as A07, except that dimethylamine hydrochloride was used instead of n-propylamine. Yellow solid; 35% yield; MP: 243–245°C; 1 H NMR (400MHz, Methanol-d4, ppm) δ8.13–7.93(m,1H),7.78(s,1H),7.62(d,J=8.5Hz,3H),6.99(d,J=7 .3Hz,2H),3.79(s,3H),3.63(t,J=6.5Hz,2H),3.27(s,6H),2.35(d,J=8.0Hz,2H),1.82–1.61(m,4H). 13C NMR(100MHz,Methanol-d4,ppm)δ162.2,160.6,154.1,148.4,141.3,131.9,12 9.4,125.1,124.3,115.7,114.9,109.3,55.9,42.6,29.1,23.6.HRMS-ESI:m / z calcd for C 22 H 27 N4O3[M+H] + 395.2078,found 395.2074.

[0067] Example 11: Preparation of 5-((7-(4-methoxyphenyl))-2-(4-methylpiperazin-1-yl)quinazolin-4-yl)amino)pentanoic acid (A11)

[0068] A11 was prepared using the same method as A07, except that N-methylpiperazine was used instead of n-propylamine. Pale yellow solid; yield 70%; MP: 260°C; 1 H NMR (400MHz, DMSO-d6, ppm) δ11.60(s,1H),9.86(s,1H),8.45–8.33(m,2H),7.70(t,J=9.7Hz,3H),7.11–7.03(m,2H), 4.98(s,2H),3.81(s,3H),3.54(q,J=6.6Hz,4H),3.43(s,4H),2.79(s,3H),2.27(t,J=7.1Hz,2H),1.72–1.50(m,4H). 13 C NMR (100MHz, DMSO-d6, ppm) δ174.8,160.6,159.4,152.3,145.8,140.6,130.5,128.6,125.1,1 23.1,115.1,114.6,108.6,55.8,51.9,42.7,42.4,41.5,33.7,27.9,22.4.HRMS-ESI:m / zcalcd for C 25 H 32 N5O3[M+H] + 450.2500,found 450.2495.

[0069] Example 12: Preparation of 5-((2-(cyclopent-1-en-1-yl))-7-(4-methoxyphenyl)quinazolin-4-yl)amino)pentanoic acid (A12)

[0070] A12 was prepared using the same method as A06, except that cyclopentene-1-boronic acid was used instead of 2-nitrophenylboronic acid. White solid; 30% yield; MP: 110–111°C. 1 H NMR (600MHz, DMSO-d6, ppm) δ12.10 (s, 1H), 10.26 (t, J = 5.8Hz, 1H), 8.49 (d, J = 8.7Hz, 1H),8.47(s,1H),7.90(d,J=8.2Hz,1H),7.72–7.69(m,2H),7.61(t,J=2.6Hz,1H),7. 07(d,J=8.5Hz,2H),3.81(s,3H),3.66–3.63(m,2H),2.82–2.74(m,2H),2.66–2.58(m ,2H),2.28(t,J=7.3Hz,2H),1.99–1.94(m,2H),1.72–1.67(m,2H),1.60–1.55(m,2H). 13 C NMR (150MHz, DMSO-d6, ppm) δ174.4,160.4,159.4,154.3,145.8,145.7,139.2,137.2,129.6,128.4 ,125.4,124.7,115.2,114.7,109.9,55.4,41.3,34.1,33.3,31.7,27.7,22.4,22.0.HRMS-ESI:m / z calcd for C 25 H 28 N3O3[M+H] + 418.2125,found418.2125.

[0071] Example 13: Preparation of 5-((2-(cyclohexen-1-yl))-7-(4-methoxyphenyl)quinazolin-4-yl)amino)pentanoic acid (A13)

[0072] A13 was prepared using the same method as A06, except that cyclohexene-1-boronic acid was used instead of 2-nitrophenylboronic acid. White solid; 34% yield; MP: 160–162°C. 1H NMR (600MHz, DMSO-d6, ppm) δ12.10 (s, 1H), 10.26 (s, 1H), 8.53 (d, J = 8.7Hz, 1H), 8.46(d,J=1.9Hz,1H),7.92(dd,J=8.7,1.8Hz,1H),7.72(d,J=8.8Hz,2H),7.49(t ,J=3.7Hz,1H),7.08(d,J=8.8Hz,2H),3.82(s,3H),3.70–3.64(m,2H),2.52–2.5 1(m,2H),2.37–2.25(m,4H),1.73–1.69(overlap,4H),1.66–1.56(overlap,4H). 13 C NMR (150MHz, DMSO-d6, ppm) δ174.8,160.8,159.6,158.4,146.2,141.1,139.9,131.3,130.1,128.8,125. 8,125.2,116.0,115.2,110.6,55.8,41.7,33.7,28.1,26.2,24.7,22.4,22.1,21.4.HRMS-ESI:m / zcalcd for C 26 H 30 N3O3[M+H] + 432.2282,found432.2281.

[0073] Example 14: Preparation of 5-((2-chloro-7-(4-methoxyphenyl))quinazolin-4-yl)amino)pentanoic acid (A14)

[0074] The preparation method for A14 is similar to that for A06, except that after obtaining the 4,7-disubstituted quinazoline intermediate 2, 1 mmol of intermediate 2 is added to a 50 mL round-bottom flask. 5 mL of methanol and 1 mL of 2N sodium hydroxide solution are then added, and the mixture is stirred at room temperature for 4 hours. After completion of the reaction, as monitored by TLC, the pH is adjusted to a weakly acidic state with 1N hydrochloric acid. The reaction is allowed to stand until a solid precipitates. The precipitate is filtered, washed with water, and recrystallized from ethanol to yield the target compound A14. This product is a white solid with a 50% yield and an MPa of 206–208°C. 1HNMR(400MHz,DMSO-d6,ppm)δ9.64(s,1H),8.49(d,J=8.6Hz,1H),7.89–7.88(overlap,2H),7.75(d,J=8.3H z,2H),7.07(d,J=8.3Hz,2H),3.81(s,3H),3.62–3.53(m,2H),2.23–2.26(m,2H),1.71–1.54(overlap,4H). 13 C NMR(100MHz,DMSO-d6,ppm)δ174.4,160.6,160.1,155.1,147.0,145.4,130.2,128 .5,125.2,124.6,119.7,114.7,111.5,55.4,41.0,33.4,27.7,22.0.HRMS-ESI:m / z calcd for C 20 H 21 N3O3Cl[M+H] + 386.1266, found 386.1273.

[0075] Example 15: Preparation of 5-((2-(dimethylamino)-7-(4-methoxyphenyl)quinazolin-4-yl)amino)-N-hydroxypentanamide (A15)

[0076] The preparation method for A15 is similar to that for A10, except that after obtaining the 2,4,7-trisubstituted quinazoline intermediate 3, 1 mmol of intermediate 3 is added to a 50 mL round-bottom flask. Then, 5 mL of methanol, 30 mmol of aqueous hydroxylamine solution, and 4 mmol of potassium hydroxide are added, and the mixture is stirred at room temperature for 4 hours. After completion of the reaction, as monitored by TLC, the pH is adjusted to a weakly acidic state with 1N hydrochloric acid. The solvent is then evaporated under reduced pressure and purified by silica gel column chromatography (DCM:MeOH = 5:1) to yield the target compound A15. Yellow solid; 29% yield; MP: 93–95°C. 1 HNMR(400MHz,DMSO-d6,ppm)δ10.43(s,1H),8.61(s,1H),8.17(s,1H),7.85-7.60(overlap,3H),7.46(s,1H), 7.15-6.92(m,2H),3.81(s,3H),3.56-3.46(m,2H),3.20(s,6H),2.11-1.93(m,2H),1.75-1.44(overlap,4H). 13C NMR(100MHz,DMSO-d6,ppm)δ169.1,159.6,159.6,159.1,156.7,144.2,131.3,128.2, 123.9,120.1,118.4,114.5,108.5,55.3,40.5,37.1,32.1,28.1,23.0.HRMS-ESI:m / z calcd for C 22 H 28 N5O3[M+H] + 410.2187,found410.2188.

[0077] Example 16: Preparation of N-hydroxy-5-((7-(4-methoxyphenyl)-2-(2-nitrophenyl)quinazolin-4-yl)amino)pentanamide (A16)

[0078] The preparation method for A16 is similar to that for A06, except that after obtaining the 2,4,7-trisubstituted quinazoline intermediate 3, 1 mmol of intermediate 3 is added to a 50 mL round-bottom flask. Then, 5 mL of methanol, 30 mmol of aqueous hydroxylamine solution, and 4 mmol of potassium hydroxide are added, and the mixture is stirred at room temperature for 4 hours. After completion of the reaction, as monitored by TLC, the pH is adjusted to a weakly acidic state with 1N hydrochloric acid. The solvent is then evaporated under reduced pressure and purified by silica gel column chromatography (DCM:MeOH = 5:1) to yield the target compound A16. Yellow solid; 25% yield; MP: 105–107°C. 1 H NMR (600MHz, DMSO-d6, ppm) δ10.37(s,1H),8.68(s,1H),8.54–8.48(m,1H),8.33(d,J=8 .6Hz,1H),8.20(dd,J=7.7,1.4Hz,1H),7.91(d,J=1.9Hz,1H),7.87–7.84(m,2H),7.83– 7.81(m,2H),7.78(td,J=7.6,1.2Hz,1H),7.69(td,J=7.7,1.4Hz,1H),7.08(d,J=8.8Hz ,2H),3.82(s,3H),3.48(d,J=5.9Hz,2H),2.03(q,J=7.1,6.6Hz,2H),1.66–1.56(m,4H). 13CNMR(150MHz,DMSO-d6,ppm)δ169.1,159.7,159.3,158.5,150.2,150.0,144.0,133.1,131.8,131.1,131 .0,130.4,128.4,124.6,123.8,123.5,123.5,114.6,112.1,55.3,40.4,32.1,28.3,22.9.HRMS-ESI:m / z calcd for C 26 H 26 N5O5[M+H] + 488.1928, found 488.1929.

[0079] Example 17: Preparation of N-hydroxy-5-((7-(4-methoxyphenyl)-2-(4-methylpiperazin-1-yl)quinazoline-4-yl)amino)pentanamide (A17)

[0080] A17 was prepared the same way as A15, except that cycloheximide was used instead of dimethylamine hydrochloride. Yellow solid; 52% yield; MP: 130–132°C; 1 H NMR (400MHz, DMSO-d6, ppm) δ10.47(s,1H),9.59(s,1H),8.72(s,1H),8.34(d,J=8.2Hz,1H),8.25(s,1H),7.73–7.65(overlap,3H),7.08( d,J=7.9Hz,2H),3.88–3.79(overlap,7H),3.58–3.53(m,2H),2.01(t,J=7.0Hz,2H),1.86–1.74(overlap,4H),1.66–1.50(overlap,8H). 13 C NMR(100MHz,DMSO-d6,ppm)δ169.4,160.5,159.0,152.1,145.7,140.9,130.7,128.6,125.0 ,122.6,115.1,114.6,108.4,55.8,48.5,41.4,32.4,28.3,27.2,26.4,23.3.HRMS-ESI:m / z calcd for C 26 H 34 N5O3[M+H] + 464.2656,found464.2659.

[0081] Example 18: Preparation of 5-((2-(cyclohexylimino-1-yl))-7-(4-methoxyphenyl)quinazolin-4-yl)amino)-N-hydroxypentanamide (A18)

[0082] A18 was prepared as A15, except that N-methylpiperazine was used instead of dimethylamine hydrochloride. Yellow solid; 28% yield; MP: 210–212°C; 1 H NMR (400MHz, DMSO-d6, ppm) δ10.48(s,1H),8.73(s,1H),8.38–8.28(m,1H),7.72(d,J=8.4Hz,2H),7.70–7.61(overlap,1H),7.09(d,J =8.3Hz,2H),3.82(s,3H),3.60–3.52(overlap,6H),3.32–3.16(overlap,4H),2.77(s,3H),2.02(t,J=6.8Hz,2H),1.66–1.58(m,4H). 13 C NMR (150MHz, DMSO-d6, ppm) δ169.5,160.2,159.8,159.1,152.3,143.9,132.3,128.5,123.9 ,121.7,119.7,114.9,109.8,55.7,54.6,45.6,43.3,40.6,32.6,28.6,23.4.HRMS-ESI:m / z calcd for C 25 H 33 N6O3[M+H] + 465.2609,found465.2608.

[0083] Example 19: Preparation of N-hydroxy-5-((7-(4-methoxyphenyl)-2-morpholinoquinazolin-4-yl)amino)pentanamide (A19)

[0084] A19 was prepared the same way as A15, except that morpholine was used instead of dimethylamine hydrochloride. White solid; yield 38%; MP: 155°C; 1H NMR(600MHz,Methanol-d4,ppm)δ8.10(d,J=8.5Hz,1H),7.83(d,J=1.8Hz,1H),7.67–7.63(m,3H),7.03(d,J=8.6Hz,2H),3 .91(t,J=4.8Hz,4H),3.85(s,3H),3.83(t,J=4.7Hz,4H),3.67(t,J=6.7Hz,2H),2.19(t,J=6.9Hz,2H),1.82–1.70(m,4H). 13 C NMR (150MHz, Methanol-d4, ppm) δ172.5,162.2,160.9,153.4,148.4,141.1,131.7,129.4, 125.2,124.5,115.7,114.9,109.5,67.2,55.9,46.4,42.6,33.2,29.0,24.2.HRMS-ESI:m / z calcd for C 24 H 30 N5O4[M+H] + 452.2292, found 452.2287.

[0085] Example 20: Preparation of N-hydroxy 5-((7-(4-methoxyphenyl)-2-(4-methylpiperidin-1-yl)quinazolin-4-yl)amino)pentanamide (A20)

[0086] A20 was prepared using the same method as A15, except that 4-methylpiperidine was used instead of dimethylamine hydrochloride. Brown solid; 20% yield; Mp: 108–110°C; 1 H NMR(400MHz,DMSO-d6,ppm)δ8.16(s,1H),8.04–7.97(m,1H),7.79–7.64(m,2H),7.51–7.30(overlap,2H),7.08–6.98(m,2H),3 .81(s,3H),2.82(t,J=12.3Hz,2H),2.06–1.90(m,2H),1.73–1.45(overlap,8H),1.29–1.20(m,1H),1.10–0.83(overlao,6H). 13C NMR (100MHz, DMSO-d6, ppm) δ169.5,160.0,159.8,158.5,151.7,144.0,132.2,128.5,123.9,121. 1,119.5,114.9,109.5,55.7,44.3,40.7,34.2,32.6,31.3,28.6,25.3,23.5,22.4.HRMS-ESI:m / z calcd for C 26 H 34 N5O3[M+H] + 464.2656, found 464.2658.

[0087] Example 21: Preparation of N-hydroxy-5-((7-(4-methoxyphenyl)-2-thiomorpholinoquinazolin-4-yl)amino)pentanamide (A21)

[0088] A21 was prepared using the same method as A15, except that thiomorpholine was used instead of dimethylamine hydrochloride. Brown solid; 29% yield; Mp: 156–158°C; 1 H NMR(600MHz,DMSO-d6,ppm)δ10.46(s,1H),9.50–9.22(m,1H),8.72(s,1H) ,8.30(d,J=7.6Hz,1H),8.14(s,1H),7.71(d,J=7.9Hz,2H),7.64(d,J=7.7H z,1H),7.07(d,J=8.0Hz,2H),4.27–4.14(m,4H),3.81(s,3H),3.55–3.51( m,2H),2.77–2.68(m,4H),2.01(t,J=7.1Hz,2H),1.66–1.55(overlap,4H). 13 C NMR (150MHz, DMSO-d6, ppm) δ169.0,163.1,160.0,159.0,152.8,145.0,130.5,128.2,12 4.4,121.9,121.8,114.7,108.4,55.4,47.7,40.9,32.0,27.8,26.3,22.9.HRMS-ESI:m / z calcd forC 24 H 30 N5O3S[M+H] + 468.2064,found468.2060.

[0089] Example 22: Preparation of 5-((2-(cyclopent-1-en-1-yl))-7-(4-methoxyphenyl)quinazolin-4-yl)amino)pentanoic acid (A22)

[0090] The preparation method for A22 is similar to that for A12, except that after obtaining the 2,4,7-trisubstituted quinazoline intermediate 3, 1 mmol of intermediate 3 is added to a 50 mL round-bottom flask. Then, 5 mL of methanol, 30 mmol of aqueous hydroxylamine solution, and 4 mmol of potassium hydroxide are added, and the mixture is stirred at room temperature for 4 hours. After completion of the reaction, as monitored by TLC, the pH is adjusted to a weakly acidic state with 1N hydrochloric acid. The solvent is then evaporated under reduced pressure and purified by silica gel column chromatography (DCM:MeOH = 5:1) to yield the target compound A22. It is a white solid; yield 33%; MP: 158–160°C. 1 H NMR (600MHz, DMSO-d6, ppm) δ10.54(s,1H),10.41(s,1H),8.64(d,J=8.9Hz,1H),8.07(d,J=1.9Hz,1H),8.03(dd,J=8.7,1.8Hz,1H),7.79–7 .75(m,2H),7.14–7.10(m,2H),4.44–4.36(m,1H),3.83(s,3H),3.69– 3.62(m,2H),2.34–2.16(m,2H),2.10–1.85(m,6H),1.73–1.56(m,4H). 13 C NMR (150MHz, DMSO-d6, ppm) δ174.4,169.1,164.7,160.5,159.8,146.3,139.6,129.7,128.6,126. 0,125.1,115.2,114.9,110.5,64.5,55.5,41.4,33.3,31.9,27.9,27.8,24.6,22.8.HRMS-ESI:m / z calcd for C 25 H 29 N4O3[M+H] + 433.2234,found433.2242.

[0091] Example 23: Preparation of 5-((2-(cyclohex-1-en-1-yl))-7-(4-methoxyphenyl)quinazolin-4-yl)amino)pentanoic acid (A23)

[0092] The preparation method for A23 is similar to that for A13, except that after obtaining the 2,4,7-trisubstituted quinazoline intermediate 3, 1 mmol of intermediate 3 is added to a 50 mL round-bottom flask. Then, 5 mL of methanol, 30 mmol of aqueous hydroxylamine solution, and 4 mmol of potassium hydroxide are added, and the mixture is stirred at room temperature for 4 hours. After completion of the reaction, as monitored by TLC, the pH is adjusted to a weakly acidic state with 1N hydrochloric acid. The solvent is then evaporated under reduced pressure and purified by silica gel column chromatography (DCM:MeOH = 5:1) to yield the target compound A23. It is a white solid; yield: 31%; MP: 97–99°C. 1 HNMR(600MHz,DMSO-d6,ppm)δ10.53(s,1H),10.34(s,1H),8.54(d,J=8.8Hz,1H),8.46(s,1H),7.88(d,J=8.6Hz,1H),7.69(d,J=8.6Hz,2H),7.47( s,1H),7.06(d,J=8.4Hz,2H),3.81(s,3H),3.70–3.61(m,2H),2.49–2.41 (m,2H),2.34–2.25(m,2H),2.11–1.96(m,2H),1.76–1.50(overlap,8H). 13 C NMR(150MHz,DMSO-d6)δ169.0,160.4,159.2,157.8,145.8,141.0,139.1,130.7,129.6,128.4,125 .4,124.8,115.2,114.8,110.0,55.4,41.3,32.0,27.8,25.9,24.3,22.8,21.7,20.9.HRMS-ESI:m / z calcd for C 26 H 31 N4O3[M+H] + 447.2391,found447.2391.

[0093] Example 24: Preparation of 5-((2-chloro-7-(4-methoxyphenyl))quinazolin-4-yl)amino)-N-hydroxypentanamide (A24)

[0094] The preparation method for A24 is similar to that for A14, except that after obtaining the 2,4,7-trisubstituted quinazoline intermediate 3, 1 mmol of intermediate 3 is added to a 50 mL round-bottom flask. Then, 5 mL of methanol, 30 mmol of aqueous hydroxylamine solution, and 4 mmol of potassium hydroxide are added, and the mixture is stirred at room temperature for 4 hours. After completion of the reaction, as monitored by TLC, the pH is adjusted to a weakly acidic state with 1N hydrochloric acid. The solvent is then evaporated under reduced pressure and purified by silica gel column chromatography (DCM:MeOH = 5:1) to yield the target compound A24. It is a white solid; yield 35%; MP: 253–255°C. 1 H NMR (400MHz, DMSO-d6, ppm) δ10.41(s,1H),8.78(t,J=5.4Hz,1H),8.72(s,1H),8.32(d,J=8.6Hz,1H),7.81(d,J=1.9Hz, 1H),7.80–7.75(m,2H),7.06(d,J=8.6Hz,2H),3.81(s,3H),3.52–3.48(m,2H),2.02(t,J=6.6Hz,2H),1.63–1.56(m,4H). 13 C NMR(100MHz,DMSO-d6,ppm)δ169.1,160.9,159.8,157.5,150.9,144.6,130.7,128 .5,124.5,123.8,122.8,114.6,112.1,55.3,40.6,32.1,28.0,22.9.HRMS-ESI:m / z calcd for C 20 H 22 N4O3Cl[M+H] + 401.1375, found 401.1382.

[0095] Example 25: Preparation of 5-((7-bromo-2-(4-methylpiperazin-1-yl))quinazolin-4-yl)amino)-N-hydroxypentanamide (A25)

[0096] 1 mmol of 2,4-dichloro-7-bromoquinazoline was added to a 50 mL round-bottom flask and dissolved in 5 mL of tetrahydrofuran. The mixture was stirred and then 1.1 mmol of methyl 5-aminopentanoate hydrochloride and 2.4 mmol of N,N-diisopropylethylamine were added sequentially. The mixture was refluxed at 65°C for 3 h. After TLC monitoring, the reaction was diluted with 150 mL of ethyl acetate. The organic phase was washed with water (3 × 30 mL) and saturated brine (3 × 30 mL) and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure to obtain intermediate 1 (crude product) as a pale yellow oil, which was used directly in the subsequent reaction step without purification.

[0097] 1 mmol of compound 1 was added to a 50 mL round-bottom flask and dissolved in 5 mL of 1,4-dioxane. 10 mmol of N-methylpiperazine, 2.4 mmol of N,N-diisopropylethylamine, and 1 mmol of cuprous iodide were added, and the mixture was refluxed at 85°C for 12 h. Following completion of the reaction, TLC was monitored. 150 mL of ethyl acetate was poured into the system, and the mixture was washed with water (3 × 30 mL) and then with saturated brine (3 × 30 mL), and dried over anhydrous sodium sulfate. The solvent was removed by distillation under reduced pressure, and the mixture was purified by silica gel column chromatography (DCM:MeOH = 30:1) to obtain 4 as a yellow solid.

[0098] 1 mmol of intermediate 4 was added to a 50 mL round-bottom flask, followed by 5 mL of methanol, 30 mmol of aqueous hydroxylamine solution, and 4 mmol of potassium hydroxide. The mixture was stirred at room temperature for 4 h. After TLC monitoring, the pH was adjusted to a weakly acidic state with 1N hydrochloric acid. The solvent was evaporated under reduced pressure and purified by silica gel column chromatography (DCM:MeOH = 5:1) to obtain the target compound A25. The product was a white solid; yield: 28%; MP: 141-143°C. 1 H NMR (400MHz, DMSO-d6, ppm) δ10.43(s,1H),8.16(t,J=5.4Hz,1H),7.95(d,J=8.7Hz,1H),7.39(d,J=1.9Hz,1H),7.16(dd,J=8.6,2.1H z,1H),3.75(t,J=4.8Hz,4H),3.54–3.45(overlap,2H),2.32(t,J=4.9Hz,4H),2.19(s,3H),1.98(t,J=6.9Hz,2H),1.65–1.49(m,4H). 13 C NMR(100MHz,DMSO-d6,ppm)δ169.0,159.6,159.0,153.2,126.8,125.9,125.0,1 23.0,109.6,54.7,46.0,43.3,40.2,32.1,28.1,23.0,14.0.HRMS-ESI:m / zcalcd for C 25 H 33 N6O2Br[M+H] + 437.1295, found 437.1300.

[0099] Example 26: Preparation of 5-((7-(4-acetylphenyl))-2-(4-methylpiperazin-1-yl)quinazolin-4-yl)amino)-N-hydroxypentanamide (A26)

[0100] A26 was prepared using the same method as A18, except that 4-acetylphenylboronic acid was used instead of 4-methoxyphenylboronic acid. Yellow solid; 23% yield; Mp: 165–167°C; 1 H NMR(600MHz,DMSO-d6,ppm)δ11.40(s,1H),10.54(s,1H),10.06(s,1H),8.77-8.64(m,1H),8.62–8.41(m,2H),7.81(d,J=8.3Hz,2H),7.77 (d,J=8.4Hz,2H),3.72–3.50(m,6H),3.28–3.07(overlap,4H),2.78(s,3H),2.19(s,3H),2.03(t,J=7.2Hz,2H),1.71–1.56(overlap,4H). 13 C NMR (150MHz, DMSO-d6, ppm) δ197.6,169.0,159.0,152.4,152.4,145.0,137.9,137.5,129.1,12 7.3,126.9,126.4,125.0,109.0,51.5,42.3,42.0,41.0,31.9,27.6,22.7,11.5.HRMS-ESI:m / z calcd for C 26 H 33 N6O3[M+H] + 477.2609,found477.2617.

[0101] Example 27: Preparation of 5-((7-(4-fluorophenyl))-2-(4-methylpiperazin-1-yl)quinazolin-4-yl)amino)-N-hydroxypentanamide (A27)

[0102] A27 was prepared using the same method as A18, except that 4-fluorophenylboronic acid was used instead of 4-methoxyphenylboronic acid. White solid; 34% yield; MP: 225–226°C. 1 H NMR (400MHz, DMSO-d6, ppm) δ11.38(s,1H),10.49(s,1H),9.89(s,1H),8.45(d,J=8.7Hz,1H),8.40(s,1H),7.83–7.75(overlap,3H),7.40 (t,J=8.6Hz,2H),5.05–4.88(m,2H),3.71–3.52(overlap,6H),3.27–3.15(m,2H),2.80(s,3H),2.02(t,J=7.2Hz,2H),1.69–1.54(m,4H). 13C NMR(100MHz,DMSO-d6,ppm)δ169.0,163.6,162.0,159.0,151.9(d, 1 J CF =277.24Hz),144.9,134.5(d, 4 J CF =1.95Hz),129.2(d, 3 J CF =8.47Hz),124.9,123.4,123.4,116.3(d, 2 J CF =21.54Hz),108.8,51.5,42.4,42.0,41.1,31.9,27.6,22.7.HRMS-ESI:m / z calcd for C 24 H 30 N6O2F[M+H] + 453.2409,found453.2410.

[0103] Example 28: Preparation of N-hydroxy-5-((2-(4-methylpiperazin-1-yl))-7-(p-tolyl)quinazolin-4-yl)amino)pentanamide (A28)

[0104] A28 was prepared using the same method as A18, except that 4-methylphenylboronic acid was used instead of 4-methoxyphenylboronic acid. Yellow solid; 22% yield; MP: 225–227°C. 1 H NMR (600MHz, DMSO-d6, ppm) δ10.62(s,1H),8.78(s,1H),8.40(s,1H),7.64(d,J=7.6Hz,2H),7.57(s,1H),7. 30(d,J=7.7Hz,2H),3.56–3.47(m,6H),2.73(s,3H),2.34(s,3H),2.02(t,J=7.1Hz,2H),1.68–1.53(M,4H). 13 C NMR(150MHz,DMSO-d6,ppm)δ169.2,159.5,159.5,144.8,138.2,135.9,129.9,126.9,12 6.8,124.5,121.6,109.4,51.9,42.2,41.7,40.6,32.0,27.9,22.9,20.9.HRMS-ESI:m / z calcd forC 25 H 33 N6O2[M+H] +449.2660, found 449.2665.

[0105] Example 29: Preparation of N-hydroxy 5-((2-(4-methylpiperazin-1-yl))-7-(4-(methylthio)phenyl)quinazolin-4-yl)amino)pentanamide (A29)

[0106] A29 was prepared using the same method as A18, except that 4-methylthiophenylboronic acid was used instead of 4-methoxyphenylboronic acid. White solid; 30% yield; Mp: 176–178°C; 1 H NMR(400MHz,DMSO-d6,ppm)δ11.51(s,1H),10.54(s,1H),9.91(s,1H),8.44 (d,J=8.7Hz,1H),8.39(s,1H),7.77–7.71(m,1H),7.67(d,J=8.7Hz,2H),7. 38(dd,J=9.0,2.7Hz,2H),4.96(s,2H),3.63(s,2H),3.55(q,J=6.4Hz,4H), 3.23(s,2H),2.80(s,3H),2.52(s,3H),2.02(t,J=6.9Hz,2H),1.59(m,4H). 13 C NMR (100MHz, DMSO-d6, ppm) δ169.4,159.3,159.3,152.2,145.5,140.5,134.4,127.7,126.6,1 25.3,123.3,114.8,109.0,51.9,42.7,42.4,41.5,32.3,28.0,23.1,14.8.HRMS-ESI:m / zcalcd for C 25 H 33 N6O2S[M+H] + 481.2380,found481.2385.

[0107] Example 30: Preparation of 5-((7-(Benzo[d][1,3]dioxol-5-yl))-2-(4-methylpiperazin-1-yl)quinazolin-4-yl)amino)-N-hydroxypentanamide (A30)

[0108] A30 was prepared using the same method as A18, except that 3,4-(methylenedioxy)phenylboronic acid was used instead of 4-methoxyphenylboronic acid. White solid; 25% yield; Mp: 200–202°C. 1H NMR (400MHz, DMSO-d6, ppm) δ10.43 (s, 1H), 8.10–7.96 (overlap, 2H), 7.45 (s, 1H), 7.39–7.30 (overlap, 2H), 7.25 (d, J = 7.8Hz, 1H), 7. 00(d,J=8.0Hz,1H),6.08(s,2H),4.02–3.67(m,8H),3.51–3.45(m,2H),2.30(s,3H),2.01(t,J=6.8Hz,2H),1.65–1.53(overlap,4H). 13 C NMR (100MHz, DMSO-d6, ppm) δ169.2,159.7,158.8,152.0,148.1,147.4,143.6,133.9,123.4,121.8 ,120.8,119.5,109.6,108.7,107.3,101.3,54.4,45.4,43.1,40.2,32.2,28.3,23.0.HRMS-ESI:m / z calcd forC 25 H 31 N6O4[M+H] + 479.2401, found 479.2408.

[0109] Example 31: Preparation of 5-((7-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl))-2-(4-methylpiperazin-1-yl)quinazolin-4-yl)amino)-N-hydroxypentanamide (A31)

[0110] A31 was prepared as A18, except that benzo-1,4-dioxane-6-boronic acid was used instead of 4-methoxyphenylboronic acid. White solid; 28% yield; Mp: 196–198°C. 1 H NMR (400MHz, DMSO-d6, ppm) δ10.50(s,1H),8.19(d,J=6.1Hz,1H),8.07(d,J=8.5Hz,1H),7.46(d,J=2.0Hz,1H),7.38–7.30(m,1H),7.22(d,J=10.1Hz ,2H),6.96–6.89(m,1H),4.27(s,4H),3.88(s,4H),3.51–3.42(m,2H),2.7 1(t,J=5.5Hz,4H),2.42(s,3H),2.01(t,J=6.7Hz,2H),1.65–1.51(m,4H). 13C NMR (100MHz, DMSO-d6, ppm) δ169.2,159.8,158.4,151.6,143.8,143.7,143.5,132.8,123.6,121.4 ,119.9,119.7,117.7,115.5,109.7,64.3,64.2,48.7,44.5,42.5,32.2,28.2,23.1.HRMS-ESI:m / z calcd for C 26 H 33 N6O4[M+H] + 493.2558, found 493.2559.

[0111] Example 32: Preparation of N-hydroxy-5-((2-(4-methylpiperazin-1-yl))-7-(thiophen-2-yl)quinazolin-4-yl)amino)pentanamide (A32)

[0112] A32 was prepared as A18, except that thiophene-2-boronic acid was used instead of 4-methoxyphenylboronic acid. Yellow solid; 33% yield; Mp: 208–210°C; 1 H NMR(600MHz,Methanol-d4,ppm)δ7.76(d,J=8.7Hz,1H),7.52(d,J=5.1Hz,1H),7.44–7.40(m,1H),7.38(d,J=5.1Hz,1H),7.35–7.30(m,1H ),7.04(t,J=4.2Hz,1H),3.85–3.78(m,4H),3.50–3.46(m,2H),2.52(t,J=4.9Hz,4H),2.31(s,3H),2.23–2.04(m,2H),1.68–1.62(m,4H). 13 C NMR (150MHz, Methanol-d4, ppm) δ175.7,161.2,159.3,151.0,144.3,139.9,129.4,127.3, 125.7,120.7,120.0,111.0,55.6,45.9,44.7,44.7,41.9,34.4,29.5,24.4.HRMS-ESI:m / z calcd for C 22 H 29 N6O2S[M+H] + 441.2067, found 441.2063.

[0113] Example 33: Preparation of N-hydroxy 5-((2-(4-methylpiperazin-1-yl))-7-(4-vinylphenyl)quinazolin-4-yl)amino)pentanamide (A33)

[0114] A33 was prepared as A18, except that 4-vinylphenylboronic acid was used instead of 4-methoxyphenylboronic acid. White solid; 32% yield; Mp: 200–202°C. 1 H NMR(600MHz,DMSO-d6,ppm)δ10.43(s,1H),8.19(t,J=5.6Hz,1H),8.11(d,J=8.5Hz,1H),7 .76(d,J=8.5Hz,2H),7.60-7.56(overlap,3H),7.45(dd,J=8.5,1.9Hz,1H),6.79(dd,J=17 .6,11.0Hz,1H),5.90(d,J=17.6Hz,1H),5.30(d,J=11.1Hz,1H),3.94–3.86(m,4H),3.51– 3.47(m,2H),2.78(t,J=5.3Hz,4H),2.48(s,3H),2.01(t,J=7.0Hz,2H),1.70–1.54(m,4H). 13 C NMR (150MHz, DMSO-d6, ppm) δ169.1,159.8,158.4,151.7,143.4,138.9,136.9,136.2,127.2,12 6.8,123.6,121.9,119.7,114.8,110.0,53.5,44.2,42.3,40.3,32.1,28.2,23.0.HRMS-ESI:m / z calcd for C 26 H 33 N6O2[M+H] + 461.2660, found 461.2665.

[0115] Example 34: Preparation of N-hydroxy 5-((2-(4-methylpiperazin-1-yl))-7-(1,4-dioxaspiro[4.5]dec-7-en-8-yl)quinazolin-4-yl)amino)pentanamide (A34)

[0116] A34 was prepared as A18, except that 4-boron-3-cyclohexene-1-one glycol acetal was used instead of 4-methoxyphenylboronic acid. White solid; 23% yield; MP: 96–98°C. 1H NMR (400MHz, DMSO-d6, ppm) δ10.44 (s, 1H), 8.05 (t, J = 5.5Hz, 1H), 7.94 (d, J = 9. 2Hz,1H),7.23–7.19(overlap,2H),6.22–6.18(m,1H),3.90(s,4H),3.81(t,J= 5.2Hz,4H),3.45(q,J=6.0Hz,2H),2.61–2.51(overlap,6H),2.40–2.36(m,2H) ,2.32(s,3H),1.99(t,J=6.8Hz,2H),1.81(t,J=6.4Hz,2H),1.62–1.51(m,4H). 13 C NMR (100MHz, DMSO-d6, ppm) δ169.1,159.7,158.6,151.6,144.3,134.7,123.6,122.7,120.3,118 .0,109.4,106.9,63.8,54.1,45.1,42.9,40.1,35.9,32.2,30.9,28.2,26.0,23.0.HRMS-ESI:m / z calcd for C 26 H 37 N6O4[M+H] + 497.2871,found497.2874.

[0117] Example 35: Preparation of N-hydroxy-6-((7-(4-methoxyphenyl)-2-(4-methylpiperazin-1-yl)quinazolin-4-yl)amino)hexanamide (A35)

[0118] A35 was prepared as A18, except that 6-aminohexanoic acid methyl ester hydrochloride was used instead of 5-aminovaleric acid methyl ester hydrochloride. White solid; 35% yield; MP: 222–224°C; 1 H NMR (400MHz, Methanol-d4, ppm) δ7.84(d,J=8.5Hz,1H),7.55(d,J=8.7Hz,2H),7.51(d,J=1.8Hz,1H),7.34(dd,J=8.5,1.8Hz,1H),6.94(d,J=8.8Hz,2 H),3.85(t,J=5.1Hz,4H),3.77(s,3H),3.52(t,J=7.1Hz,2H),2.53(s,4H) ,2.32(s,3H),2.05(t,J=7.4Hz,2H),1.71–1.58(m,4H),1.41–1.33(m,2H). 13C NMR (100MHz, Methanol-d4, ppm) δ172.7,161.5,161.4,159.0,150.4,146.7,133.4,129.3,124.1 ,122.1,120.7,115.4,110.5,55.8,55.7,46.0,44.8,42.0,33.7,29.7,27.7,26.5.HRMS-ESI:m / z calcd for C 26 H 35 N6O3[M+H] + 479.2765,found 479.2769.

[0119] Example 36: Preparation of N-hydroxy-7-((7-(4-methoxyphenyl)-2-(4-methylpiperazin-1-yl)quinazolin-4-yl)amino)heptylamide (A36)

[0120] A36 was prepared as A18, except that 7-aminoheptanoic acid methyl ester hydrochloride was used instead of 5-aminopentanoic acid methyl ester hydrochloride. White solid; 40% yield; MP: 227–229°C; 1 H NMR(400MHz,DMSO-d6,ppm)δ9.91(t,J=5.7Hz,1H),8.52–8.42(m,2H),7.77–7.69(overlap,3H),7.09(d,J=8.6Hz,2H),3.81(s,3H),3.64 –3.50(m,6H),3.35–3.21(overlap,4H),2.79(s,3H),2.20(t,J=7.3Hz,2H),1.65(t,J=7.1Hz,2H),1.52–1.45(m,2H),1.39–1.25(m,4H). 13 C NMR (100MHz, DMSO-d6, ppm) δ174.6,160.2,159.0,151.9,145.5,140.2,130.1,128.3,124.9,122. 9,114.8,114.1,108.3,55.4,51.5,42.4,42.0,41.3,33.7,28.3,28.0,26.3,24.5.HRMS-ESI:m / z calcdfor C 27 H 37 N6O3[M+H] + 493.2922,found493.2923.

[0121] Example 37: Preparation of N-hydroxy-3-((7-(4-methoxyphenyl)-2-(2-nitrophenyl)quinazolin-4-yl)amino)propanamide (A37)

[0122] A37 was prepared as A16, except that 3-aminopropionic acid methyl ester hydrochloride was used instead of 5-aminovaleric acid methyl ester hydrochloride. White solid; 72% yield; MP: 102–104°C; 1 H NMR (600MHz, DMSO-d6, ppm) δ10.55 (s, 1H), 10.34 (s, 1H), 8.66 (d, J = 8.8Hz, 1 H),8.22(d,J=8.0Hz,1H),8.20(s,1H),8.17(dd,J=7.6,1.4Hz,1H),8.12(dd, J=8.9,1.8Hz,1H),8.01–7.97(m,1H),7.94–7.90(m,1H),7.82–7.80(m,2H), 7.14–7.11(m,2H),3.84(s,3H),3.79(d,J=6.3Hz,2H),2.43(t,J=7.1Hz,2H). 13 C NMR (150MHz, DMSO-d6, ppm) δ166.8,160.5,159.6,156.9,148.3,146.4,140.9,133.6,132.7,131.8,1 29.7,128.6,128.2,126.6,125.1,124.8,124.7,114.9,110.6,55.4,38.4,31.2.HRMS-ESI:m / zcalcd for C 24 H 22 N5O5[M+H] + 460.1615, found 460.1618.

[0123] Example 38: Preparation of N-hydroxy-4-((7-(4-methoxyphenyl)-2-(2-nitrophenyl)quinazolin-4-yl)amino)-n-butyramide (A38)

[0124] A38 was prepared as A16, except that methyl 4-aminobutyrate hydrochloride was used instead of methyl 5-aminovalerate hydrochloride. White solid; 69% yield; MP: 105–107°C. 1H NMR (600MHz, DMSO-d6, ppm) δ10.42 (s, 1H), 8.72 (s, 1H), 8.56 (t, J = 6.5Hz, 1H), 8.33 (d ,J=8.6Hz,1H),8.19(dd,J=7.7,1.5Hz,1H),7.91(d,J=1.9Hz,1H),7.88–7.85(overlap ,2H),7.82(d,J=8.8Hz,2H),7.79–7.77(m,1H),7.69(td,J=7.7,1.5Hz,1H),7.08(d,J =8.8Hz,2H),3.82(s,3H),3.51–3.47(m,2H),2.10(t,J=7.5Hz,2H),1.88–1.83(m,2H). 13 C NMR (150MHz, DMSO-d6, ppm) δ169.0,159.7,159.4,158.5,150.2,150.0,144.1,133.1,131.9,131.1,1 31.0,130.4,128.4,128.4,124.7,123.8,123.5,114.6,112.1,55.3,40.3,30.0,24.8.HRMS-ESI:m / z calcd forC 25 H 24 N5O5[M+H] + 474.1772,found474.1771.

[0125] Example 39: Determination of KDM6B and HDAC Inhibitory Activity in Vitro

[0126] 1. The KDM6B inhibitory activity assay procedure is as follows:

[0127] The experimental design used GSK-J1 as a positive control compound. During the initial screening phase, all candidate compounds were tested for inhibition at a fixed concentration of 50 μM in duplicate. Potentially active compounds were further investigated by establishing a five-point concentration gradient using a systematic dilution method. The initial concentration was set at 50 μM, and working solutions of varying concentrations were prepared using a five-fold serial dilution method. The specific experimental procedure was as follows: To establish the experimental system, a substrate complex solution containing the characteristic peptide substrate, 2-oxoglutarate, ascorbic acid, and ferrous sulfate was precisely prepared in 1× reaction buffer. Subsequently, 5 μL of KDM6B enzyme solution was precisely pipetted into the wells of the pretreated assay plate and equilibrated at room temperature for 15 minutes. The enzymatic reaction was then initiated by adding an equal volume of substrate solution and terminated by incubation at constant temperature for 60 minutes. Following termination, 15 μL of each of the receptor and donor solutions prepared in 1× Alphalisa buffer was added to the system, and the immunobinding reaction was carried out for 60 minutes at room temperature in the dark. Finally, an EnSpire multifunctional plate reader equipped with an Alpha detection module was used for endpoint signal acquisition, and changes in enzyme activity were quantitatively analyzed by chemiluminescence intensity.

[0128] Inhibition rate and IC 50 The IC value is calculated using the following method: Inh% = (Max-Signal) / (Max-Min)*100, where Max is the maximum signal value, Min is the minimum signal value, and Signal is the actual detection signal value. 50 Nonlinear fitting calculations were performed using GraphPad software. The mathematical expression of the model was defined as: Y = Y min +(Y max -Y min ) / [1+10^((LogIC 50 -X)*n_H)], where Y represents the inhibition rate (%), Y max With Y min They correspond to the maximum and minimum response platform values, n_H corresponds to the Hill coefficient, and X represents the concentration of the compound.

[0129] 2. The HDAC inhibitory activity assay procedure is as follows:

[0130] In this study, vorinostat (SAHA) was used as a broad-spectrum positive control. A five-point concentration gradient dose-effect curve was established for the compound using a five-fold geometric dilution method, with the initial working concentration set at 50 μM. The specific experimental implementation plan is as follows: During the construction of the enzyme reaction system, trypsin and acetylated peptide substrate were first added to the Tris-HCl buffer system according to the experimental design concentration, and a homogenized substrate complex solution was prepared by ultrasonic vibration. 15 μL of the HDAC enzyme system working solution was accurately pipetted into the pretreated 384-well detection plate and equilibrated on a constant temperature oscillation platform for 15 minutes (25°C). Subsequently, 10 μL of substrate was accurately injected through a multi-channel pipetting system to initiate the reaction, and the deacetylation process was completed by incubating at room temperature for 60 minutes. The final detection was performed using a SynergyMX full-wavelength microplate reader, with an excitation wavelength of 355 nm and an emission wavelength of 460 nm set for fluorescence signal acquisition, and the relevant data were read for subsequent analysis.

[0131] Inhibition rate and IC 50 The IC value is calculated using the following method: Inh% = (Max-Signal) / (Max-Min)*100, where Max is the maximum signal value, Min is the minimum signal value, and Signal is the actual detection signal value. 50 Nonlinear fitting calculations were performed using GraphPad software. The mathematical expression of the model was defined as: Y = Y min +(Y max -Y min ) / [1+10^((LogIC 50 -X)*n_H)], where Y represents the inhibition rate (%), Y max With Y min They correspond to the maximum and minimum response platform values, n_H corresponds to the Hill coefficient, and X represents the concentration of the compound.

[0132] 3. Activity test results

[0133] Table 1. Inhibitory activity of compounds against KDM6B and HDAC enzymes

[0134]

[0135]

[0136] In terms of activity evaluation, carboxylic acid derivatives have certain inhibitory activity against KDM6B but the inhibitory activity against HDAC3 is not as expected. Hydroxamic acid compounds have significant inhibitory activity against KDM6B and HDAC3 / 8, and some compounds show selective inhibition of HDAC3 / 8 compared to HDAC2. Through activity screening, we found that 14 synthetic compounds showed good and comparable inhibitory activity against both KDM6B and HDAC3. The IC values of some target molecules for KDM6B and HDAC3 are 50 has reached nanomolar levels, with A38 having the best activity (KDM6B IC 50 =638nM; HDAC3 IC 50 =4.9nM). A38 is currently the most potent KDM6B / HDAC dual-target inhibitor.

[0137] In summary, the embodiments of the present invention provide a 2,4,7-trisubstituted quinazoline derivative and its preparation method and application. The compound has a novel structure, has a certain inhibitory effect on KDM and HDAC, and belongs to a new type of inhibitor structure. In addition, the preparation method of the compound is simple, easy to operate, has low requirements for equipment, and is suitable for large-scale industrial production in the later stage. The compound undergoes structural modification and in-depth pharmacological research, and has good application prospects in the preparation of KDM and HDAC dual-target inhibitors.

[0138] The above description is only a partial specific implementation case of the present invention, and the known specific contents or attempts in the scheme are not described in detail here. It should be pointed out that the above embodiments do not limit the present invention in any way. For those skilled in the art, any technical solution obtained by equivalent replacement or equivalent transformation falls within the protection scope of the present invention. The scope of protection claimed in this application shall be based on the content of its claims, and the specific implementation methods and other records in the specification can be used to interpret the content of the claims.

Claims

1. A 2,4,7-trisubstituted quinazoline derivative, characterized in that: The general structural formula of the derivative is Ⅰ: wherein R1 is independently selected from: a carboxyl group or a hydroxamic acid group; n=2-8; The R2 is independently selected from: C5-C10 aryl, mono- or poly-substituted C5-C10 aryl, dioxaspiro substituent, halogen atom; The R3 is independently selected from: N-methylpiperazinyl, C5-C10 heterocycle containing N / O / S, aliphatic amine, C5-C10 aryl, mono- or poly-substituted C5-C10 aryl, halogen atom, olefin, C5-C8 cyclic olefin.

2. The derivative according to claim 1, characterized in that The derivative is, 3. The method for preparing a 2,4,7-trisubstituted quinazoline derivative according to claim 1, wherein: The preparation method comprises the following steps: using commercially available raw material 2,4-dichloro-7-bromoquinazoline as a raw material, first subjecting it to a nucleophilic substitution reaction with a chain amine in an alkaline environment to obtain a 4-monosubstituted quinazoline intermediate; subjecting the 4-monosubstituted quinazoline intermediate to a Suzuki coupling reaction to modify the 7th position to generate a 4,7-disubstituted quinazoline intermediate; subjecting the 4,7-disubstituted quinazoline intermediate to a Suzuki coupling reaction with a substituted boronic acid, or subjecting it to a nucleophilic substitution reaction with various amines or heterocycles to obtain a 2,4,7-trisubstituted quinazoline intermediate; and subjecting the 2,4,7-trisubstituted quinazoline intermediate to a hydrolysis reaction under strong alkaline conditions or a hydroxylamine hydroximation reaction in an alkaline environment to obtain a 2,4,7-trisubstituted quinazoline derivative I.

4. Use of the 2,4,7-trisubstituted quinazoline derivative according to claim 1 as a dual-target inhibitor of histone lysine demethylase (KDM) and histone deacetylase (HDAC).

5. Use of the 2,4,7-trisubstituted quinazoline derivatives according to claim 1 as drugs for the development and treatment of diseases mediated by histone lysine demethylase (KDM) or histone deacetylase (HDAC).

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