DNMT1-HDAC double-target inhibitor as well as preparation method and application thereof

Abstract

The invention relates to a DNMT1-HDAC double-target inhibitor as shown in a formula (I) as well as a preparation method and application thereof, and the DNMT1-HDAC double-target inhibitor not only shows good DNMT1 and HDAC enzyme inhibition activity, but also has remarkable anti-tumor activity on solid tumors and hematoma. As a double-target inhibitor based on DNMT1 and HDACs reported for the first time, the compound has a great prospect in the aspect of cancer treatment; # imgabs0 #

Description

Technical Field

[0001] The present invention relates to a DNMT1-HDAC dual-target inhibitor and a preparation method and application thereof, belonging to the technical field of medicine. Background Art

[0002] Cancer is a disease characterized by genetic abnormalities, which is characterized by resistance to treatment and high mortality. However, epigenetics has gradually been recognized as a factor that contributes to cancer behavior and cannot be explained solely by genetic alterations. In general, the heterogeneity of tumor cells in terms of growth potential, invasiveness, malignant transformation ability, and resistance to therapeutic intervention can be attributed to the disturbance of epigenetic regulation. Compared with normal cells, cancer cells cause extensive and profound changes in the epigenome, including DNA methylation, histone acetylation, histone methylation, and chromatin remodeling. Among them, abnormal lysine acetylation of histone tails and hypermethylation of CpG islands in the promoter region of tumor suppressor genes are drivers and biomarkers of tumor progression and poor prognosis.

[0003] DNA methylation is regulated by DNA methyltransferases (DNMTs). The human DNMT family includes five members, which can be further divided into two groups, including typical members (DNMT1, DNMT3A, and DNMT3B), which can add methyl groups derived from s-adenosyl-l-methionine (SAM) to the C-5 position of cytosine in CpG islands, and atypical members (DNMT2 and DNMT3L), which do not have catalytic activity. Unlike DNMT3A and DNMT3B, which establish methylation patterns from scratch, DNMT1 is responsible for maintaining DNA methylation and transmitting methylation patterns from patterns to daughter strands. In addition, compared with other DNMTs, the level of DNMT1 is dominant in both normal tissues and metastatic cancer tissues. At present, DNMT inhibitors are divided into covalent nucleoside inhibitors and non-nucleoside reversible inhibitors. The former is represented by azacitidine (AZA), decitabine (DAC), and sibulalin, and the latter is represented by RG108 and SGI-1027.

[0004] Histone deacetylases (HDACs) can remove acetyl groups from lysine in histone tails or non-histone substrates. 18 HDACs have been discovered in humans and are divided into four categories based on their catalytic mechanism and intracellular localization, including zinc-dependent class I (HDAC1, 2, 3, 8), class IIa (HDAC4, 5, 7, 9), class IIb (HDAC 6, 10), class IV (HDAC11) and NAD +Class III (Sirtuin 1-7) that are dependent on HDACs. To date, HDAC inhibitors are commonly used to treat hematological malignancies, and some inhibitors have been approved by the FDA, including vorinostat (SAHA), belinostat (PDX101), panobinostat (LBH589), cedabenamide (CS055), and romidepsin (FK 228).

[0005] DNMT inhibitors and HDAC inhibitors are used in combination to treat a variety of cancers, such as acute myeloid leukemia (AML), multiple myeloma (MM), diffuse large B-cell lymphoma (DLBCL), T-cell lymphoma (TCL), and myelodysplastic syndrome. For example, the clinical efficacy observed in AML patients treated with azacitidine and panobinostat combined is superior to panobinostat monotherapy. Although the combination of DNMT and HDAC inhibitors has certain advantages in the treatment of AML and tumor immunotherapy, the disadvantage is that drug-drug interactions limit the realization of synergistic effects, such as complex dosing regimens and negative effects on pharmacokinetics. In contrast, the design of DNMT / HDAC dual-target inhibitors can circumvent these potential disadvantages. However, the currently reported DNMT / HDAC dual-target inhibitors have poor inhibition effects on DNMT or HDAC, and lack subtype selectivity for DNMT1, resulting in no obvious therapeutic advantages compared with monotherapy.

[0006] Therefore, the development of an effective DNMT1 / HDAC dual inhibitor is of great clinical significance. Summary of the invention

[0007] In view of the deficiencies in the prior art, the present invention provides a DNMT1-HDAC dual-target inhibitor and a preparation method and application thereof.

[0008] The technical solution of the present invention is as follows:

[0009] DNMT1-HDAC dual-target inhibitor, the structural formula of the inhibitor is shown in formula (I) or a pharmaceutically acceptable salt of formula (I):

[0010]

[0011] Wherein, X is an aromatic heterocycle or one of the following linking groups:

[0012]

[0013] A is one of the following structures:

[0014]

[0015] R1 is hydroxyl or 2-aminophenyl;

[0016] R2 is one of the following structures

[0017]

[0018] R3 is a hydrogen atom and is one of the following structures:

[0019]

[0020] The DNMT1-HDAC dual-target inhibitor is selected from one of the following compounds:

[0021] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -Hydroxysuccinamide (11a);

[0022] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (11b);

[0023] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 - Hydroxyadipoic acid amide (11c);

[0024] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 7 -Hydroxypimelamide (11d);

[0025] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 8 -Hydroxyoctanamide (11e);

[0026] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 9 -Hydroxyazelazone (11f);

[0027] N 1-(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 - Hydroxy-p-toluene diamide (11 g);

[0028] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-4-((4-(hydroxycarbamoyl)benzyl)oxy)benzamide (11h);

[0029] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(Hydroxycarbamoyl)phenyl)succinamide (11i);

[0030] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(Hydroxycarbamoyl)phenyl)glutaramide (11j);

[0031] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 -(4-(Hydroxycarbamoyl)phenyl)adipamide (11k);

[0032] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(Hydroxycarbamoyl)phenyl)succinamide (111);

[0033] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(Hydroxycarbamoyl)phenyl)glutaramide (11m);

[0034] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N4 -(4-(3-(Hydroxyamino)-3-oxopropyl-1-en-1-yl)phenyl)succinamide (11n);

[0035] 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-hydroxybenzamide (11o);

[0036] (E)-3-4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-hydroxyacrylamide (11p);

[0037] 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(hydroxycarbamoyl)phenyl)carbamoyl)phenyl)piperidinium 4-ammonium salt (11q);

[0038] (E)-1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(3-hydroxyamino)-3-oxopropyl-1-en-1-yl)benzyl)carbamoyl)benzyl)piperidin-4-ammonium salt (11r);

[0039] (E)-1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(3-hydroxyamino)-3-oxopropyl-1-en-1-yl)benzyl)oxy)benzyl)piperidin-4-ammonium salt (11s);

[0040] 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((7-(hydroxyamino)-7-oxoheptyl)oxy)benzyl)piperidin-4-ammonium salt (11t);

[0041] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-N-hydroxypyrimidine-5-carboxamide (11u);

[0042] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)(methyl)amino)-N-hydroxypyrimidine-5-carboxamide (11v);

[0043] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)piperidin-1-yl)-N-hydroxypyrimidine-5-carboxamide (15a)

[0044] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)(methyl)amino)piperidin-1-yl)-N-hydroxypyrimidine-5-carboxamide (15b);

[0045] (E)-3-(4-((4-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)methyl)phenyl)-N-hydroxyacrylamide (19a);

[0046] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)-N-hydroxypyrimidine-5-carboxamide (19b);

[0047] (E)-3-(4-((((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-hydroxyacrylamide (23a);

[0048] (R,E)-3-(4-((((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-hydroxyacrylamide ((R)-23a);

[0049] (S,E)-3-(4-((((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-hydroxyacrylamide ((S)-23a);

[0050] 2-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)-N-hydroxypyrimidine-5-carboxamide (23b);

[0051] (E)-3-(4-((4-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)methyl)phenyl)-N-hydroxyacrylamide (27);

[0052] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(2-aminophenyl)glutaramide (30a);

[0053] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(2-aminophenyl)terephthalamide (30b);

[0054] 4-(2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-2-oxoethyl)-N-(2-aminophenyl)benzamide (30c);

[0055] 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-(2-aminophenyl)benzamide (30d);

[0056] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)(methyl)amino)-N-(2-aminophenyl)pyrimidine-5-carboxamide (30e);

[0057] N 1 -(2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N 4 -Hydroxysuccinamide (40a);

[0058] N 1 -(2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N 5 -Hydroxyglutaramide (40b);

[0059] N 1-(3-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)propyl)-N 4 -Hydroxysuccinamide (40c);

[0060] 2-((2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)(methyl)amino)-N-hydroxypyrimidine-5-carboxamide (40d);

[0061] N 1 -(1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (44a);

[0062] (E)-(1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (44b);

[0063] (E)-3-(4-(((1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-hydroxyacrylamide (44c);

[0064] 2-(4-(3,5-Dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperazin-1-yl)-N-hydroxypyrimidine-5-carboxamide (48).

[0065] The DNMT1-HDAC dual-target inhibitor is selected from one of the following structures:

[0066]

[0067]

[0068]

[0069]

[0070]

[0071]

[0072] The second object of the present invention is to provide a method for preparing a DNMT1-HDAC dual-target inhibitor, wherein the preparation method comprises any of the following methods:

[0073] The preparation methods of compounds 11a-11V are as follows:

[0074]

[0075] Reagents and conditions: (a) p-toluenesulfonyl chloride, N,N-diisopropylethylamine, 4-dimethylaminopyridine, acetonitrile, 80°C, 6 hours; (b) n-propionaldehyde, N-methylmorpholine, anhydrous ethanol, room temperature, 30 hours; (c) N,N-diisopropylethylamine, N,N-dimethylformamide, 85°C, 5 hours; (d) isoamyl nitrite, cupric chloride, acetonitrile, 80°C, 2.5 hours; (e) aminopiperidine derivatives, carbonic acid Potassium, dichloromethane, 4 hours; (f) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (g) 1) 9a-n, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 9o-t, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 3) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours;

[0076] The preparation methods of compounds 15a, 15b, 19a and 19b are as follows:

[0077]

[0078] Reagents and conditions: (a) aminopiperidine derivative, potassium carbonate, dichloromethane, 4 hours; (b) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (c) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (d) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (e) piperazine-1-tert-butylcarboxylate, potassium carbonate, dichloromethane, 4 hours; (f) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (g) 1) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 2) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours;

[0079] The preparation methods of compounds 23a, (R)-23a, (S)-23a, 23b, 27, and 30a-e are as follows:

[0080]

[0081] Reagents and conditions: (a) piperidine-4-aminomethyl tert-butyl carbamate, potassium carbonate, dichloromethane, 4 hours; (b) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (c) 1) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 2) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (d) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (e) tert-butyl (2-(methylamino)ethyl)carbamate, potassium carbonate, dichloromethane, 4 hours; (f) 4M hydrochloric acid / acetic acid Ethyl ester, room temperature, 12 hours; (g) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (i) 1) 28a-c, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 28d, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 3) 28e, potassium carbonate, acetonitrile, 80°C, 6 hours; (j) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours;

[0082] The preparation method of compound 40a-d is as follows:

[0083]

[0084] Reagents and conditions: (a) methanesulfonyl chloride, triethylamine, dichloromethane, room temperature, 4 hours; (b) diisobutylaluminum hydride, tetrahydrofuran, -20°C, 6 hours; (c) p-toluenesulfonyl chloride, triethylamine, DMAP, acetonitrile, 80°C, 4 hours; (d) 4, triethylamine, DMF, 85°C, 5 hours; (e) isoamyl nitrite, cupric chloride, acetonitrile, 80°C, 2.5 hours; (f) 2-methylamino derivative, potassium carbonate, dichloromethane, 4 hours; (g) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (h) 1) 9a or 9b, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (i) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours;

[0085] The preparation methods of compounds 44a-c and 48 are as follows:

[0086]

[0087] Reagents and conditions: (a) tert-butyl 4-carbamate, potassium carbonate, dichloromethane, 4 hours; (b) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (c) 1) 9b, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 3) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (d) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (e) piperazine-1-tert-butylcarboxylate, potassium carbonate, dichloromethane, 4 hours; (f) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (g) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours.

[0088] A pharmaceutical composition comprises a therapeutically effective amount of the DNMT1 and HDAC dual-target inhibitor and pharmaceutically acceptable excipients.

[0089] The use of the DNMT1-HDAC dual-target inhibitor in the preparation of drugs for treating tumor diseases related to the target.

[0090] The tumor is lung cancer, liver cancer, non-small cell lung cancer, colon cancer, skin cancer, pancreatic cancer, ovarian cancer, breast cancer, bladder cancer, lymphoma, esophageal cancer, gastrointestinal cancer, nasopharyngeal cancer, leukemia or myeloma.

[0091] Technical features and advantages of the present invention:

[0092] 1. The compounds of the present invention not only exhibit good DNMT1 and HDAC enzyme inhibitory activity, can inhibit DNMT1 and HDAC activity in cells, but also have strong in vitro and in vivo anti-tumor activity, and can significantly delay tumor growth.

[0093] 2. As the first reported dual-target anti-tumor drug based on DNMT1 and HDAC, the compound of the present invention has further development and research value. BRIEF DESCRIPTION OF THE DRAWINGS

[0094] Figure 1 The graph shows the in vivo antitumor activity of compound (R)-23a and the combination drug group (GSK5032 and SAHA) on human acute monocytic leukemia cells MV4-11, wherein A: weight changes of tumor-bearing mice; B: tumor volume growth curve of tumor-bearing mice.

[0095] Figure 2 The graph shows the in vivo antitumor activity of compound (R)-23a and the combination drug group (GSK5032 and SAHA) on mouse colon cancer cell MC38, wherein A: weight change of tumor-bearing mice; B: tumor volume growth curve of tumor-bearing mice. DETAILED DESCRIPTION

[0096] The features of the present invention are further described below by way of examples, but the present invention is not limited to the following examples.

[0097] The reagents, raw materials or intermediates in the examples are all commercially available or can be prepared according to literature methods. The experimental methods in the following examples without specifying specific conditions are usually carried out under conventional conditions or under conditions recommended by the manufacturers.

[0098] Example 1: Preparation of Intermediate 6

[0099] 2-Amino-2-oxo-1-phenylethyl-4-methylbenzenesulfonate (2)

[0100] 2-Hydroxy-2-phenylacetamide (10 g, 66.15 mmol), N,N-diisopropylethylamine (DMAP, 25.65 g, 198.46 mmol) and 4-dimethylaminopyridine (1.21 g, 9.92 mmol) were added to acetonitrile (100 mL) with 4-methylbenzenesulfonyl chloride (18.92 g, 99.23 mmol), stirred at room temperature for 6 h, and then the solvent was removed by rotary evaporation. The obtained residue was washed with 100 mL of water, and the mixture was extracted with ethyl acetate (30 mL×3). The separated organic phase was dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography to obtain compound 2. The synthesis methods of compounds (R)-2, (S)-2 and 34 were the same as the operation steps.

[0101] White solid, yield 77.2%, 1 H NMR (400MHz, DMSO-d6) δ7.77(s,2H),7.48(d,J=7.9Hz,1H),7.43(d,J=8.1Hz,4H),7.34(s,4H),5.68(s,1H),2.40(s,3H).

[0102] (R)-2-Amino-2-oxo-1-phenylethyl-4-methylbenzenesulfonate ((R)-2)

[0103] White solid, yield 69.1%, 1 H NMR (400MHz, DMSO) δ7.79–7.73(m,3H),7.45–7.38(m,3H),7.38–7.28(m,5H),5.68(s,1H),2.40(s,3H).

[0104] (S)-2-Amino-2-oxo-1-phenylethyl-4-methylbenzenesulfonate ((S)-2)

[0105] White solid, yield 71.5%, 1H NMR (400MHz, DMSO) δ7.80–7.72(m,3H),7.47–7.38(m,3H),7.37–7.28(m,5H),5.68(s,1H),2.40(s,3H).

[0106] 4-Methylmorpholine-4-ammonium salt-6-amino-3,5-dicyano-4-ethylpyridine-2-thioate (4)

[0107] Add propanal (870 mg, 14.98 mmol) to a solution of 2-cyanoacetamide (3 g, 29.96 mmol) and N-methyl phthalide (3.03 g, 29.96 mmol) in EtOH (40 mL), stir at room temperature for 30 hours, filter the reaction solution, collect the filter cake, wash it thoroughly with ethanol, and dry it overnight to obtain compound 4.

[0108] Brown solid, yield 30.4%, 1 H NMR (400MHz, DMSO-d6) δ7.58(s,2H),3.67(s,2H),2.72(s,2H),2.62(q,J=7.5Hz,2H),2.51(s,3H),1.19(t,J=7.6Hz,3H).

[0109] 2-((6-Amino-3,5-dicyano-4-ethylpyridin-2-yl)thio)-2-phenylacetamide (5)

[0110] Compound 4 (661 mg, 1.96 mmol) and N,N-diisopropylethylamine (760 mg, 5.88 mmol) were dissolved in DMF (5 mL) and compound 2 (598 mg, 1.96 mmol) was added thereto. The temperature was raised to 85°C for reaction for 5 hours, and then the solvent was removed by rotary evaporation. The obtained residue was washed with 50 mL of water, and the mixture was extracted with ethyl acetate (15 mL×3). The separated organic phase was dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography to obtain compound 5. The synthesis methods of compounds (R)-5, (S)-5 and 35 were the same as the operation steps. Pale yellow solid, yield 68.1%, 1 H NMR (400MHz, DMSO-d6) δ7.90(s,2H),7.73(s,1H),7.59(d,J=7.3Hz,2H),7.41–7.27(m,4H),5.56(s,1H),2.69(q,J=7.5Hz,2H),1.18(t,J=7.6Hz,3H).

[0111] (R)-2-((6-amino-3,5-dicyano-4-ethylpyridin-2-yl)thio)-2-phenylacetamide ((R)-5)

[0112] Yellow solid, yield 40.7%. 1 H NMR (400MHz, DMSO) δ7.90(s,2H),7.73(s,1H),7.63–7.55(m,2H),7.37–7.28(m,4H),5.57(s,1H),2.68(t,J=7.6Hz,2H),1.18(t,J=7.6Hz,3H).

[0113] (S)-2-((6-amino-3,5-dicyano-4-ethylpyridin-2-yl)thio)-2-phenylacetamide ((S)-5)

[0114] Yellow solid, yield 45.1%. 1 H NMR (400MHz, DMSO) δ7.90(s,2H),7.73(s,1H),7.62–7.55(m,2H),7.41–7.26(m,4H),5.56(s,1H),2.68(t,J=7.6Hz,2H),1.18(t,J=7.6Hz,3H).

[0115] 2-((6-Chloro-3,5-dicyano-4-ethylpyridin-2-yl)thio)-2-phenylacetamide (6)

[0116] Compound 5a (402 mg, 1.19 mmol) and copper chloride (797 mg, 5.93 mmol) were dissolved in acetonitrile (20 mL) and isoamyl nitrite (278 mg, 2.37 mmol) was added thereto. The temperature was raised to 80°C for reaction for 2.5 hours, and then the solvent was removed by rotary evaporation. The obtained residue was washed with 100 mL of water, and the mixture was extracted with ethyl acetate (30 mL×3). The separated organic phase was dried over anhydrous sodium sulfate and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography to obtain compound 6. The synthesis methods of compounds (R)-6, (S)-6 and 36 were the same as the operation steps. Pale yellow solid, yield 31.3%. 1 H NMR (400MHz, Chloroform-d) δ7.61–7.47(m,3H),7.40(d,J=6.6Hz,4H),5.63(s,1H),2.99(q,J=7.5Hz,2H),1.36(t,J=7.7Hz,3H).

[0117] (R)-2-((6-chloro-3,5-dicyano-4-ethylpyridin-2-yl)thio)-2-phenylacetamide ((R)-6)

[0118] Pale yellow solid, yield 31.7%. 1H NMR (400MHz, DMSO) δ7.99(s,1H),7.61–7.51(m,2H),7.43(s,1H),7.38(dd,J=8.2,6.3 Hz,2H),7.37–7.28(m,1H),5.69(s,1H),2.86(q,J=7.6Hz,2H),1.23(t,J=7.7Hz,3H).

[0119] (S)-2-((6-chloro-3,5-dicyano-4-ethylpyridin-2-yl)thio)-2-phenylacetamide ((S)-6)

[0120] Pale yellow solid, yield 30.1%. 1 H NMR (400MHz, DMSO) δ7.99 (s, 1H), 7.61–7.54 (m, 2H), 7.45–7.28 (m, 4H), 5.69 (s, 1H), 2.86 (q, J = 7.6Hz, 2H), 1.23 (t, J = 7.7Hz, 3H).

[0121] Example 2: Preparation of intermediate 7a

[0122] To a solution of compound 6 (200 mg, 0.56 mmol) and potassium carbonate (232 mg, 1.68 mmol) in CH2Cl2 (10 mL) was added tert-butyl piperidine-4-carbamate (224 mg, 1.12 mmol), stirred at room temperature for 4 hours, and then washed with 100 mL of water. The organic phase was then separated, dried over anhydrous sodium sulfate, and concentrated to obtain a crude product. The latter was purified by silica gel column chromatography to obtain 7a. The synthesis methods of compounds 7b, 12a, 12b, 16, 20, (R)-20, (S)-20, 24, 37a-c, 41 and 45 were the same as the above operation steps.

[0123] Tert-butyl (1-6 ((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)carbamate (7a)

[0124] White solid, yield 68.5%. 1 H NMR (400MHz, DMSO) δ7.91(s,1H),7.52(d,J=7.4Hz,2H),7.41–7.33(m,4H),6.94(d,J=7.7Hz,1H),5.52(s,1H),4.4 5(d,J=13.3Hz,2H),3.61(s,2H),2.76(q,J=7.5Hz,2H),1.88(d,J=12.9Hz,2H),1.41(s,9H),1.21(t,J=7.5Hz,3H).

[0125] Tert-butyl (1-6 ((2-amino-2-oxo-1-phenylethyl) thio) -3,5-dicyano-4-ethylpyridin-2-yl) piperidin-4-yl) (methyl) carbamate (7b)

[0126] White solid, yield 69.5%, 1 H NMR (400MHz, DMSO) δ7.91(s,1H),7.52(d,J=7.4Hz,2H),7.42–7.34(m,4H),6.94(d,J=7.7Hz,1H),5.52(s,1H),4.4 5(d,J=13.3Hz,2H),3.61(s,2H),2.76(q,J=7.5Hz,2H),1.88(d,J=12.9Hz,2H),1.41(s,9H),1.21(t,J=7.5Hz,3H).

[0127] Example 3: Preparation of Intermediate 8a

[0128] 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-ammonium salt (8a)

[0129] Compound 7a (292 mg, 0.56 mmol) was added to saturated hydrochloric acid ethyl acetate (10 mL), stirred at room temperature for 12 hours, filtered and purified by recrystallization of the filter cake with ethyl acetate (20 mL) to obtain 8a. The synthesis methods of compounds 8b, 13a, 13b, 17, 21, (R)-21, (S)-21, 25, 38a-c, 42 and 46 conformed to the operation steps. White solid, yield 85.8%. 1 H NMR (400MHz, DMSO) δ8.22(s,3H),8.01(s,1H),7.54(d,J=7.4Hz,2H),7.41–7.32(m,4H),5.58(s,1H),4.59(d,J=13. 7Hz,2H),3.32–3.20(m,2H),2.77(q,J=7.6Hz,2H),2.09(d,J=12.6Hz,2H),1.65–1.57(m,2H),1.22(t,J=7.6Hz,3H).

[0130] 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-methylpiperidin-4-ammonium salt (8b)

[0131] White solid, yield 90.4%. 1H NMR (400MHz, DMSO) δ9.03(s,2H),8.01(s,1H),7.53(d,J=7.4Hz,2H),7.39(dd,J=14.7,7.3Hz,4H),5.57(s,1H),4.63(d,J=13.6Hz,2H),3 .20(t,J=12.2Hz,3H),2.77(q,J=7.6Hz,2H),2.57(t,J=5.3Hz,3H),2.17(d,J=12.6Hz,2H),1.60(t,J=12.7Hz,2H),1.21(t,J=7.6Hz,3H).

[0132] Example 4: Preparation of Intermediates 9a-f

[0133] O-(tetrahydro-2H-pyran-2-yl)hydroxylamine (1 g, 8.54 mmol), the corresponding cyclic anhydride (8.54 mmol) and triethylamine (2.6 g, 25.61 mmol) were added to 15 mL CH2Cl2 and stirred at room temperature for 3 hours. The organic phase was then washed with a saturated NaCl solution (30 ml), the organic phase was separated and dried over anhydrous Na2SO4, and concentrated to obtain a crude product, which was purified by silica gel column chromatography to obtain compounds 9a-f.

[0134] 4-Oxo-4-(((tetrahydro-2H-pyran-2-yl)oxy)amino)butanoic acid (9a)

[0135] White solid, yield 53%. 1 H NMR (400MHz, DMSO) δ4.79 (d, J=3.3Hz, 1H), 3.95–3.87 (m, 1H), 3.50 (dd, J=10.2, 5.7Hz, 1H),2.37(t,J=7.0Hz,2H),2.20(t,J=7.1Hz,2H),1.72–1.57(m,3H),1.57–1.44(m,3H).

[0136] 5-Oxo-5-(((tetrahydro-2H-pyran-2-yl)oxy)amino)pentanoic acid (9b)

[0137] White solid, yield 45.7%. 1 H NMR(400MHz,DMSO)δ4.81(d,J=3.7Hz,1H),3.95–3.89(m,1H),3.50(dd,J=10.1,5.5Hz,1H ),2.19(t,J=7.4Hz,2H),2.02(t,J=7.4Hz,2H),1.74–1.60(m,5H),1.52(d,J=11.6Hz,3H).

[0138] 6-Oxo-6-(((tetrahydro-2H-pyran-2-yl)oxy)amino)hexanoic acid (9c)

[0139] White solid, yield 48.3%. 1 H NMR (400MHz, DMSO) δ10.92(s,1H),4.81(d,J=3.2Hz,1H),3.96–3.88(m,1H),3.50(dd,J=10 .3,5.3Hz,1H),2.22–2.15(m,2H),2.03–1.95(m,2H),1.71–1.58(m,3H),1.56–1.43(m,7H).

[0140] 7-Oxo-7-(((tetrahydro-2H-pyran-2-yl)oxy)amino)heptanoic acid (9d)

[0141] Colorless oily liquid, yield 48.1%. 1 H NMR (400MHz, DMSO) δ12.01(s,1H),10.90(s,1H),4.79(d,J=3.3Hz,1H),3.95–3.87(m,1H),3.50(dd,J=10.1,5. 5Hz,1H),2.18(t,J=7.4Hz,2H),1.97(t,J=7.1Hz,2H),1.71–1.59(m,3H),1.53–1.39(m,7H),1.24–1.17(m,2H).

[0142] 8-Oxo-8-(((tetrahydro-2H-pyran-2-yl)oxy)amino)octanoic acid (9e)

[0143] White solid, yield 46%. 1 H NMR (400MHz, DMSO) δ10.90 (s, 1H), 4.80 (d, J = 3.2Hz, 1H), 3.96–3.88 (m, 1H), 3.53–3.45 (m, 1H), 2.28 (t,J=7.4Hz,2H),1.96(t,J=7.3Hz,2H),1.68–1.60(m,3H),1.54–1.39(m,7H),1.24(h,J=3.9Hz,4H).

[0144] 9-Oxo-9-(((tetrahydro-2H-pyran-2-yl)oxy)amino)decanoic acid (9f)

[0145] White solid, yield 64.3%. 1H NMR (400MHz, DMSO) δ11.97(s,1H),10.90(s,1H),4.80(d,J=3.3Hz,1H),3.95–3.89(m,1H),3.49(s,1H),2 .19(t,J=7.4Hz,2H),1.96(d,J=7.4Hz,2H),1.69–1.60(m,3H),1.52–1.45(m,7H),1.24(d,J=4.0Hz,6H).

[0146] Example 5: Preparation of Intermediates 9g-9u

[0147] 4-((Tetrahydro-2H-pyran-2-yl)carbamoyl)benzoic acid (9 g)

[0148] The corresponding carboxylic acid (5.31mmol), EDCI (1.23g, 6.4mmol), DMAP (843mg, 6.9mmol), and pyranolamine (622mg, 5.31mmol) were added to 15mL CH2Cl2 and reacted at room temperature for 8 hours. After the reaction, the reaction solution was washed with 1M hydrochloric acid aqueous solution (100mL) and saturated sodium chloride aqueous solution (100mL) in sequence. The organic phase was then dried over anhydrous Na2SO4 and concentrated. 9g was obtained by purification by silica gel column chromatography. The synthesis method of compound 9h-u was the same as the operation steps. Colorless oily liquid, yield 70.8%. 1 H NMR (400MHz, DMSO) δ11.81(s,1H),8.02(d,J=8.1Hz,2H),7.87(d,J=8.2Hz,2H),5.03(d, J=3.1Hz,1H),4.06(t,J=9.5Hz,1H),3.56–3.51(m,1H),1.73(s,3H),1.59–1.51(m,3H).

[0149] 4-((4-(((Tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)benzyl)oxy)benzoic acid (9h)

[0150] White solid, yield 90.2%. 1H NMR (400MHz, DMSO) δ12.67(s,1H),11.67(s,1H),7.93–7.87(m,2H),7.79(d,J=8.0Hz,2H),7.55(d,J=8.1Hz,2H),7.14– 7.07(m,2H),5.26(s,2H),5.00(d,J=3.0Hz,1H),4.11–4.01(m,1H),3.57–3.48(m,1H),1.72(s,3H),1.57–1.51(m,3H).

[0151] 4-Oxo-4-((4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)phenyl)amino)butanoic acid (9i)

[0152] White solid, yield 44.5%. 1 H NMR (400MHz, DMSO) δ11.52(s,1H),10.25(s,1H),7.87–7.53(m,4H),4.98(d,J=3.1Hz,1H),4.08–4.02(m ,1H),3.51(d,J=11.4Hz,1H),2.58(t,J=6.9Hz,2H),2.52(s,2H),1.82–1.63(m,3H),1.57–1.49(m,3H).

[0153] 5-Oxo-5-((4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)phenyl)amino)pentanoic acid (9j)

[0154] White solid, yield 27.9%. 1 H NMR (400MHz, DMSO) δ11.54(s,1H),10.17(s,1H),7.95–7.53(m,4H),4.98(d,J=3.1Hz,1H),4.09–4.01(m,1H),3.52(dd ,J=9.8,5.4Hz,1H),2.38(t,J=7.4Hz,2H),2.28–2.22(m,2H),1.85–1.78(m,2H),1.74–1.67(m,3H),1.59–1.51(m,3H).

[0155] 6-Oxo-6-((4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)phenyl)amino)hexanoic acid (9k)

[0156] Pale yellow solid, yield 54%. 1H NMR (400MHz, DMSO) δ11.50(s,1H),10.15(s,1H),7.87–7.57(m,4H),4.98(d,J=3.2Hz,1H),4.12–4.00( m,1H),3.57–3.44(m,1H),2.34(t,J=7.1Hz,2H),2.22(t,J=7.3Hz,2H),1.71(s,3H),1.60–1.48(m,7H).

[0157] 4-Oxo-4-((4-(((tetrahydro-2H-pyran-2-yl)carbamoyl)benzyl)amino)butanoic acid (91)

[0158] White solid, yield 54.2%. 1 H NMR (400MHz, DMSO) δ11.52(s,1H),10.22(s,1H),7.72(d,J=8.7Hz,2H),7.66(d,J=8.8Hz,2H),4.97(d,J=3.2Hz,1H),4.06 (s,1H),3.70(t,J=6.5Hz,2H),3.51(d,J=11.4Hz,1H),2.53(s,2H),2.07–2.01(m,2H),1.71(s,3H),1.54(d,J=5.7Hz,3H).

[0159] 5-Oxo-5-((4-(((tetrahydro-2H-pyran-2-yl)carbamoyl)benzyl)amino)pentanoic acid (9m)

[0160] White solid, yield 48.1%. 1 H NMR (400MHz, DMSO) δ11.63(s,1H),8.45(t,J=6.0Hz,1H),7.72(d,J=8.1Hz,2H),7.32(d,J=8.0Hz,2H),4.99(d,J=3.1Hz,1 H),4.30(d,J=5.9Hz,2H),4.07(d,J=8.6Hz,1H),3.59(s,1H),2.24–2.17(m,4H),1.77–1.69(m,5H),1.54(d,J=5.7Hz,3H).

[0161] 4-Oxo-4-((4-(3-oxo-3-(((tetrahydro-2H-pyran-2-yl)oxy)amino)propyl-1-en-1-yl)phenyl)amino)butanoic acid (9n)

[0162] White solid, yield 38.0%. 1H NMR (400MHz, DMSO) δ12.15(s,1H),11.18(s,1H),10.15(s,1H),7.63(d,J=8.3Hz,2H),7.51(d,J=8.3Hz,2H),7.42(d,J=15.6Hz,1H),6.39(d,J =15.9Hz,1H),4.90(s,1H),3.96(s,1H),3.54(dd,J=11.1,4.8Hz,1H),2.58(t,J=6.6Hz,2H),2.53(d,J=5.8Hz,2H),1.69(s,3H),1.54(s,3H).

[0163] 4-Formyl-N-((tetrahydro-2H-pyran-2-yl)oxy)benzamide (9o)

[0164] White solid, yield 57.2%. 1 H NMR (400MHz, DMSO) δ11.88(s,1H),10.09(s,1H),8.04–7.94(m,4H),5.04(d,J=2.8Hz,1H ),4.10–4.03(m,1H),3.54(dd,J=11.3,4.6Hz,1H),1.79–1.71(m,3H),1.61–1.53(m,3H).

[0165] (E)-3-(4-Formylphenyl)-N-((tetrahydro-2H-pyran-2-yl)oxy)acrylamide (9p)

[0166] White solid, yield 39.1%. 1 H NMR (400MHz, DMSO) δ11.37(s,1H),10.02(s,1H),7.95(d,J=8.0Hz,2H),7.81(d,J=8.0Hz,2H),7.58(d,J=15.9Hz ,1H),6.67(d,J=15.9Hz,1H),4.93(s,1H),3.97(s,1H),3.55(dd,J=11.1,5.5Hz,1H),1.70(s,3H),1.55(s,3H).

[0167] 4-Formyl-N-(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)benzyl)benzamide (9q)

[0168] White solid, yield 31.6%. 1H NMR (400MHz, DMSO) δ11.61(s,1H),10.09(s,1H),9.33(t,J=5.9Hz,1H),8.08(d,J=8.2Hz,2H),8.02(d,J=8.2Hz,1H),7 .73(d,J=8.2Hz,2H),7.42(d,J=8.0Hz,2H),4.98(s,1H),4.55(d,J=6.0Hz,2H),4.06(s,1H),1.72(s,2H),1.55(s,2H).

[0169] 4-Formyl-N-(4-(3-oxo-3-(((tetrahydro-2H-pyran-2-yl)oxy)amino)propyl-1-en-1yl)benzyl)benzamide (9r)

[0170] White solid, yield 47.7%. 1 H NMR (400MHz, DMSO) δ11.24(s,1H),10.09(s,1H),9.24(t,J=6.0Hz,1H),8.09(d,J=8.2Hz ,1H),8.02(d,J=8.1Hz,1H),7.95(d,J=8.3Hz,1H),7.74(d,J=8.3Hz,1H),7.55(d,J=7.7H z,2H),7.48(d,J=15.5Hz,1H),7.37(dd,J=8.0,5.1Hz,2H),6.48(d,J=15.8Hz,1H),4.91 (s,1H),4.52(t,J=6.8Hz,2H),3.96(s,1H),3.55(s,1H),1.73–1.66(m,3H),1.54(s,3H).

[0171] (E)-3-(4-((4-Formylphenoxy)methyl)phenyl)-N-((tetrahydro-2H-pyran-2-yl)oxy)acrylamide (9s)

[0172] White solid, yield 52.8%. 1 H NMR (400MHz, DMSO) δ11.27(s,1H),9.87(s,1H),7.92–7.84(m,2H),7.62(d,J=7.9Hz,2H),7.55–7.46(m,3H),7.26–7.18(m,2H), 6.53(d,J=15.8Hz,1H),5.27(s,2H),4.92(s,1H),3.98(d,J=9.7Hz,1H),3.54(d,J=11.5Hz,1H),1.75–1.67(m,3H),1.54(s,3H).

[0173] 7-(4-Formylphenoxy)-N-((tetrahydro-2H-pyran-2-yl)oxy)heptylamide (9t)

[0174] White solid, yield 62.9%. 1 H NMR (400MHz, DMSO) δ10.92(s,1H),9.86(s,1H),7.91–7.81(m,2H),7.15–7.07(m,2H),4.81(d,J=3.2Hz,1H),4.07(t,J=6.4Hz,2H),3.95–3.88(m,1 H),3.52–3.45(m,1H),2.01–1.97(m,2H),1.73(p,J=6.7Hz,3H),1.68–1.6 0(m,3H),1.51(q,J=7.6Hz,6H),1.43–1.37(m,2H),1.31(q,J=4.5Hz,2H).

[0175] 2-Chloro-N-((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (9u)

[0176] White solid, yield 29.4%. 1 H NMR (400MHz, DMSO) δ11.46 (s, 1H), 8.66 (s, 2H), 4.95 (d, J = 3.3Hz, 1H), 4.08–3.97 (m, 1H), 1.75–1.65 (m, 3H), 1.57–1.50 (m, 3H).

[0177] Example 6: Preparation of Intermediates 10a-10n

[0178] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -((Tetrahydro-2H-pyran-2-yl)oxy)succinamide (10a)

[0179] 9a (190 mg, 0.875 mmol) and NMM (110 mg, 1.09 mmol) were added to THF (10 mL), and isobutyl chlorocarbonate (120 mg, 0.875 mmol) was added, and the mixture was stirred at 0°C for 0.5 h. The resulting filtrate was added dropwise to CH2Cl2 (10 mL) containing 8a (200 mg, 0.437 mmol) and N-methylmorpholine (44 mg, 0.437 mmol) at 0°C. The mixture was stirred at room temperature for 6 hours, and then the solvent was removed by rotary evaporation. The obtained residue was washed with 100 mL of water and extracted with ethyl acetate (30 mL×3). The organic phase was washed with 1M aqueous hydrochloric acid solution (100 mL), saturated aqueous sodium bicarbonate solution (100 mL), and saturated aqueous sodium chloride solution (100 mL) in sequence. The organic phase was then dried over anhydrous Na2SO4 and concentrated. The compound 10a was purified by silica gel column chromatography, and the synthesis methods of compounds 10b-n, 29a-c, 39a-c and 43a were the same as the above steps. The compound 10a was a white solid with a yield of 44.1%. 1 H NMR (400MHz, DMSO) δ10.97 (s, 1H), 7.91 (d, J = 8.4Hz, 2H), 7.55–7.48 (m, 2H), 7.44–7 .30(m,4H),5.52(s,1H),4.79(s,1H),4.42(s,2H),3.91(s,2H),3.49(d,J=11.2Hz,3 H),2.76(q,J=7.4Hz,2H),2.32(t,J=7.7Hz,2H),2.24(d,J=7.2Hz,2H),1.88(d,J=1 2.7Hz,2H),1.64(s,3H),1.50(s,3H),1.43(t,J=10.3Hz,2H),1.20(t,J=7.6Hz,3H).

[0180] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -((Tetrahydro-2H-pyran-2-yl)oxy)glutaramide (10b)

[0181] White solid, yield 50.7%. 1H NMR (400MHz, DMSO) δ10.98 (s, 1H), 8.00 (s, 1H), 7.90 (d, J = 7.6Hz, 1H), 7.56–7.51 (m, 2H), 7. 41–7.30(m,4H),5.56(s,1H),4.82(s,1H),4.48–4.37(m,2H),3.91(d,J=11.5Hz,2H),3.55– 3.43(m,3H),2.76(q,J=7.6Hz,2H),2.08(t,J=7.5Hz,2H),2.00(t,J=7.5Hz,2H),1.89(d,J= 13.5Hz,2H),1.78–1.74(m,2H),1.69–1.61(m,3H),1.54–1.40(m,5H),1.20(t,J=7.5Hz,3H).

[0182] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 -((Tetrahydro-2H-pyran-2-yl)oxy)adipamide (10c)

[0183] White solid, yield 60.4%. 1 1H NMR (400MHz, DMSO) δ10.91(s,1H),7.92(s,1H),7.83(d,J=7.6Hz,1H),7.57–7.49(m,2H ),7.40–7.31(m,4H),5.53(s,1H),4.80(s,1H),4.43(s,2H),3.91(s,2H),3.49(d,J=11. 4Hz,1H),3.09(d,J=7.2Hz,2H),2.76(q,J=7.6Hz,2H),2.07(s,2H),1.99(d,J=6.5Hz,2H ),1.89(d,J=12.7Hz,2H),1.63(s,3H),1.53–1.40(m,7H),1.21(dd,J=10.9,7.4Hz,5H).

[0184] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 7 -((Tetrahydro-2H-pyran-2-yl)oxy)pimelanediamide (10d)

[0185] White solid, yield 66.2%. 1H NMR (400MHz, DMSO) δ10.91(s,1H),7.94(s,1H),7.83(d,J=7.6Hz,1H),7.55–7.49(m,2H),7. 42–7.30(m,4H),5.53(s,1H),4.79(t,J=3.0Hz,1H),4.49–4.37(m,2H),3.97–3.84(m,2H),3 .49(d,J=11.4Hz,1H),3.37(s,2H),2.75(q,J=7.5Hz,2H),2.05(t,J=7.4Hz,2H),1.97(d,J= 7.3Hz,2H),1.88(d,J=12.7Hz,2H),1.68–1.58(m,3H),1.53–1.39(m,9H),1.24–1.17(m,5H).

[0186] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 8 -((Tetrahydro-2H-pyran-2-yl)oxy)octanediamide (10e)

[0187] White solid, yield 31.1%. 1 H NMR(400MHz,DMSO)δ10.92(s,1H),7.94(s,1H),7.90–7.76(m,1H),7.55–7 .49(m,2H),7.48–7.17(m,4H),5.53(s,1H),4.84(s,1H),4.57–4.41(m,2H) ,4.00–3.87(m,2H),3.55–3.51(m,3H),2.76(d,J=7.8Hz,2H),2.34–2.26(m ,4H),2.05(t,J=7.4Hz,2H),1.86(s,2H),1.53–1.40(m,10H),1.25(m,7H).

[0188] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 9 -((Tetrahydro-2H-pyran-2-yl)oxy)octanediamide (10f)

[0189] White solid, yield 83%. 1H NMR (400MHz, DMSO) δ10.91(s,1H),7.94(s,1H),7.83(d,J=7.6Hz,1H),7.60–7.49(m,2H) ,7.46–7.29(m,4H),5.54(s,1H),4.79(s,1H),4.44(dd,J=13.1,8.9Hz,2H),3.96–3.86(m ,2H),3.49(d,J=11.0Hz,1H),2.76(q,J=7.6Hz,2H),2.06(t,J=7.4Hz,2H),1.97(d,J=7.4 Hz,2H),1.88(d,J=12.6Hz,2H),1.69–1.58(m,3H),1.56–1.38(m,9H),1.22–1.17(m,9H).

[0190] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -((Tetrahydro-2H-pyran-2-yl)oxy)terephthalamide (10 g)

[0191] White solid, yield 48.2%. 1 H NMR (400MHz, DMSO) δ11.79(s,1H),8.51(d,J=7.8Hz,1H),7.95(d,J=7.8Hz,2H),7.85(d ,J=8.2Hz,2H),7.52(d,J=7.5Hz,2H),7.44–7.30(m,4H),5.55(s,1H),5.02(d,J=3.3Hz, 1H),4.58(t,J=14.1Hz,2H),4.27–4.13(m,1H),4.08(d,J=7.7Hz,1H),3.54(q,J=6.6Hz, 3H),2.77(q,J=7.6Hz,2H),2.05–1.97(m,2H),1.77–1.53(m,8H),1.19(t,J=7.5Hz,3H).

[0192] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-4-((4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)phenyl)oxy)benzamide (10h)

[0193] White solid, yield 41.4%. 1H NMR (400MHz, DMSO) δ11.67 (s, 1H), 8.21 (d, J = 7.8Hz, 1H), 7.94 (s, 1H), 7.82 (dd ,J=21.7,8.3Hz,4H),7.53(t,J=7.8Hz,4H),7.47–7.25(m,4H),7.09(d,J=8.6Hz ,2H),5.54(s,1H),5.25(s,2H),5.00(s,1H),4.57(t,J=13.7Hz,2H),4.17(s,2H ),2.77(t,J=7.6Hz,2H),1.99(s,2H),1.69–1.49(m,8H),1.21(t,J=7.5Hz,3H).

[0194] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)phenyl)succinamide (10i)

[0195] White solid, yield 38.6%. 1 H NMR (400MHz, DMSO) δ11.56(s,1H),10.36(s,1H),8.04(s,1H),7.71(q,J=8.7Hz,4H),7.53(d,J =7.4Hz,2H),7.40–7.31(m,5H),5.57(s,1H),4.98(s,1H),4.43(d,J=13.0Hz,2H),3.91(s,2H) ,3.51(d,J=11.8Hz,2H),2.75(d,J=7.7Hz,2H),2.61(t,J=6.9Hz,2H),2.43(t,J=7.2Hz,2H),1 .89(d,J=12.5Hz,2H),1.71(s,2H),1.54(s,3H),1.45(t,J=12.6Hz,3H),1.22(t,J=7.6Hz,3H).

[0196] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)phenyl)glutaramide (10j)

[0197] White solid, yield 39.2%. 1H NMR (400MHz, DMSO) δ11.54(s,1H),10.32(s,1H),8.01(s,2H),7.71(q,J=8.7Hz,4H),7.52(d,J= 7.4Hz,2H),7.42–7.30(m,5H),5.57(s,1H),4.98(s,1H),4.41(d,J=13.0Hz,2H),4.05(d,J=7.6 Hz,1H),3.91(s,1H),3.51(d,J=11.7Hz,2H),2.75(q,J=7.7Hz,2H),2.61(t,J=6.9Hz,2H),2.41 (t,J=7.0Hz,2H),1.87(d,J=12.6Hz,2H),1.71(s,2H),1.66–1.30(m,8H),1.20(t,J=7.6Hz,3H).

[0198] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 -(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)phenyl)adipamide (10k)

[0199] White solid, yield 41.4%. 1 H NMR (400MHz, DMSO) δ11.60(s,1H),10.52(s,1H),8.13(s,1H),8.06(d,J=7.6Hz,1H),7.73(s,4 H),7.58–7.52(m,2H),7.40–7.29(m,4H),5.61(s,1H),4.98(s,1H),4.42(d,J=13.3Hz,2H),4.1 0–4.01(m,1H),3.91(s,1H),3.51(d,J=11.2Hz,3H),2.75(q,J=7.6Hz,2H),2.40(t,J=6.8Hz,2 H),2.18–2.10(m,2H),1.87(d,J=12.8Hz,2H),1.71(s,3H),1.55(s,9H),1.20(t,J=7.6Hz,3H).

[0200] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)benzyl)succinamide (101)

[0201] White solid, yield 42.5%. 1 H NMR (400MHz, DMSO) δ11.60(s,1H),8.46(t,J=6.0Hz,1H),8.00(s,1H),7.94(d,J=7.5Hz,1H),7.72(d, J=7.9Hz,2H),7.52(d,J=7.5Hz,2H),7.40–7.31(m,6H),5.56(s,1H),4.98(s,1H),4.40(d,J=10.8Hz, 2H),4.30(d,J=6.0Hz,2H),4.05(d,J=6.2Hz,1H),3.90(s,1H),3.54–3.43(m,2H),2.75(q,J=7.6Hz,2 H),2.43–2.33(m,4H),1.87(d,J=12.5Hz,2H),1.71(s,2H),1.62–1.35(m,6H),1.21(d,J=7.6Hz,3H).

[0202] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)benzyl)glutaramide (10m)

[0203] White solid, yield 41.3%. 1 H NMR (400MHz, DMSO) δ11.58(s,1H),8.42(t,J=6.0Hz,1H),7.98(s,1H),7.92(d,J=7.5Hz,1H),7.70(d, J=7.9Hz,2H),7.48(d,J=7.5Hz,2H),7.42–7.32(m,6H),5.57(s,1H),4.96(s,1H),4.40(d,J=10.8Hz, 2H),4.38(d,J=6.0Hz,2H),4.08(d,J=6.2Hz,1H),3.92(s,1H),3.52–3.41(m,2H),2.76(q,J=7.6Hz,2 H),2.41–2.31(m,4H),1.88(d,J=12.5Hz,2H),1.72(s,2H),1.64–1.31(m,8H),1.21(t,J=7.6Hz,3H).

[0204] (E)-N 1-(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(3-oxo-3-(((tetrahydro-2H-pyran-2-yl)oxy)amino)propyl-1-en-1-yl)phenyl)succinamide (10n)

[0205] White solid, yield 35.9%. 1 H NMR (400MHz, DMSO) δ11.22(s,1H),10.22(s,1H),7.99(d,J=8.2Hz,2H),7.65(d,J=8.3Hz,2H) ,7.59–7.45(m,4H),7.45–7.24(m,5H),6.40(d,J=15.8Hz,1H),5.55(s,1H),4.90(s,1H),4.42 (s,2H),3.93(s,2H),3.53(d,J=11.8Hz,2H),2.76(q,J=7.0Hz,2H),2.59(t,J=7.2Hz,2H),2. 43(t,J=7.2Hz,2H),1.93–1.85(m,2H),1.69(s,3H),1.58–1.39(m,5H),1.21(t,J=7.6Hz,3H).

[0206] Example 7: Preparation of intermediates 10o-10t

[0207] 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-((tetrahydro-2H-pyran-2-yl)oxy)benzamide (10o)

[0208] 8a (350 mg, 0.766 mmol) and 9o (287 mg, 1.15 mmol) were added to a mixed solvent of MeOH / CH2Cl2 (1:1, 10 mL). After reacting at room temperature for 9 hours, sodium triacetoxyborohydride (649 mg, 3.06 mmol) was added and stirred at room temperature for 3 hours. After the solvent was removed by rotary evaporation, the residue was washed with 50 mL of water and the mixture was extracted with ethyl acetate (30 mL×3). The separated organic phase was washed with 1 M hydrochloric acid aqueous solution (100 mL), saturated sodium bicarbonate aqueous solution (100 mL) and saturated sodium chloride aqueous solution (100 mL) in sequence. The organic phase was then dried over anhydrous Na2SO4 and concentrated. 10o was obtained by purification by silica gel column chromatography. The synthesis methods of compounds 10p-t, 18a, 22, (R)-22, (S)-22, 26, 29d and 43b were the same as the above operation steps. White solid, yield 41.2%. 1 H NMR (400MHz, DMSO) δ11.87(s,1H),9.58(d,J=6.5Hz,2H),8.04(s,1H),8.00(d,J=8.0Hz,4H),7.95(d,J= 8.0Hz,2H),7.51(d,J=7.5Hz,2H),7.36(m,4H),5.61(s,1H),5.03(d,J=2.8Hz,1H),4.65(d,J=13.4Hz,2 H),4.26(t,J=6.0Hz,2H),4.08(m,1H),3.53–3.42(m,2H),3.52–3.39(m,1H),3.22(d,J=13.0Hz,2H),2. 77(q,J=7.6Hz,2H),2.29(d,J=12.5Hz,2H),1.85–1.69(m,5H),1.58–1.53(m,3H),1.21(t,J=7.7Hz,3H).

[0209] (E)-3-(4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-((tetrahydro-2H-pyran-2-yl)oxy)acrylamide (10p)

[0210] White solid, yield 39.6%. 1H NMR (400MHz, DMSO) δ11.23 (s, 1H), 7.93 (d, J = 2.1Hz, 1H), 7.56–7.49 (m, 4H), 7.47–7.39 ( m,3H),7.39–7.28(m,4H),6.48(d,J=15.7Hz,1H),5.53(s,1H),4.91(s,1H),4.39(d,J=13 .1Hz,2H),3.96(s,1H),3.54(dd,J=10.0,5.5Hz,2H),2.75(q,J=7.3Hz,3H),1.94(d,J=1 2.5Hz,2H),1.70(s,3H),1.54(d,J=4.8Hz,3H),1.44–1.32(m,2H),1.20(t,J=7.6Hz,3H).

[0211] 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)benzyl)benzamide (10q)

[0212] White solid, yield 38.2%. 1 H NMR (400MHz, DMSO) δ11.20(s,1H),9.21(t,J=6.1Hz,1H),7.99(s,1H),7.98(d,J=8.0Hz,2H),7.80–7.70( m,4H),7.54–7.52(m,2H),7.41–7.27(m,6H),5.61(s,1H),4.88(s,1H),4.66(d,J=13.4Hz,2H),4.53(d,J =5.9Hz,2H),4.28(t,J=6.1Hz,2H),3.91(s,1H),3.36(m,1H),3.20(t,J=12.8Hz,2H),2.77(q,J=7.5Hz,2 H),2.30(d,J=12.4Hz,2H),1.77(dt,J=15.2,7.2Hz,2H),1.65(s,3H),1.54(s,3H),1.21(t,J=7.8Hz,3H)

[0213] (E)-4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-(4-(((tetrahydro-2H-pyran-2-yl)oxy)carbamoyl)propyl-1-en-1-yl)benzyl)benzamide (10r)

[0214] White solid, yield 35.1%. 1 H NMR (400MHz, DMSO) δ11.25(s,1H),9.07(t,J=6.1Hz,1H),7.95(s,1H),7.89(d,J=8.0Hz ,2H),7.62–7.43(m,7H),7.43–7.27(m,6H),6.48(d,J=15.9Hz,1H),5.54(s,1H),4.91( s,1H),4.47(dd,J=27.3,9.5Hz,4H),3.92(d,J=29.0Hz,4H),2.75(q,J=7.5Hz,2H),2.0 3–1.95(m,2H),1.70(s,3H),1.53(d,J=5.0Hz,3H),1.42(s,2H),1.20(t,J=7.6Hz,3H).

[0215] (E)-3-(4-((4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)phenoxy)methyl)phenyl)-N-(4-(((tetrahydro-2H-pyran-2-yl)oxy)acrylamide (10s)

[0216] White solid, yield 33.8%. 1 H NMR (400MHz, DMSO) δ11.29(s,1H),7.98(s,1H),7.60(d,J=7.9Hz,2H),7.51(dd,J=15.5,7.7Hz,5H) ,7.39(dd,J=8.2,6.2Hz,4H),7.37–7.30(m,2H),7.03(d,J=8.1Hz,2H),6.53(d,J=15.9Hz,1H),5.56 (s,1H),5.15(s,2H),4.92(s,1H),4.51(d,J=13.1Hz,2H),3.93(d,J=24.3Hz,2H),3.57–3.51(m,1H ),3.23(s,2H),2.76(d,J=7.6Hz,2H),2.08(s,2H),1.70(s,2H),1.54(s,5H),1.21(t,J=7.6Hz,3H).

[0217] 7-(4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)phenoxy)-N-(4-(((tetrahydro-2H-pyran-2-yl)oxy)benzamide (10t)

[0218] White solid, yield 32.1%. 1 H NMR(400MHz,DMSO)δ10.90(s,1H),7.93(s,1H),7.52(d,J=7.4Hz,2H),7.45– 7.22(m,6H),6.92(s,2H),5.54(s,1H),4.80(s,1H),4.47(s,2H),3.95(t,J= 6.5Hz,2H),3.48(s,2H),2.76(q,J=7.5Hz,2H),1.99(t,J=7.3Hz,2H),1.74– 1.59(m,5H),1.55–1.34(m,8H),1.31(d,J=7.4Hz,2H),1.21(t,J=7.6Hz,3H).

[0219] Example 8: Preparation of intermediate 10u

[0220] 8a (356 mg, 0.78 mmol), 9u (242 mg, 0.94 mmol) and K2CO3 (162 mg, 1.17 mmol) were added to acetonitrile (10 mL) and reacted at 80°C for 6 hours. The reaction solution was diluted with 50 mL of water and extracted with ethyl acetate (20 mL × 3). The separated organic phase was dried over anhydrous Na2SO4 and concentrated to obtain a crude product. The latter was purified by silica gel column chromatography to obtain 10u. The synthesis methods of compounds 10v, 14a, 14b, 18b, 22b, 29e, 39d, 43c and 47 were the same as the operation steps.

[0221] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-N-(4-(((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (10u)

[0222] White solid, yield 29.6%. 1 H NMR (400MHz, DMSO) δ11.51(s,1H),8.66(s,2H),7.97–7.89(m,2H),7.51(d,J=7.5Hz,2H),7.44–7.30(m,4H),5.53(s,1H),4.95(s,1H), 4.55(s,2H),4.18(s,1H),4.03(s,1H),2.76(q,J=7.3Hz,2H),2.03(d,J=13.3Hz,2H),1.71(s,3H),1.55(s,5H),1.21(t,J=7.6Hz,3H).

[0223] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)(methyl)amino)-N-(4-(((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (10u)

[0224] White solid, yield 31.1%. 1 H NMR (400MHz, DMSO) δ11.54(s,1H),8.71(s,2H),7.92(s,1H),7.52(d,J=7.3H z,2H),7.44–7.31(m,4H),5.55(s,1H),4.98(d,J=16.4Hz,2H),4.74(d,J=13. 5Hz,2H),4.04(s,1H),3.51(s,1H),3.27(d,J=12.0Hz,2H),3.02(s,3H),2.77 (q,J=7.3Hz,3H),1.75(d,J=27.5Hz,7H),1.55(s,3H),1.22(t,J=7.5Hz,3H).

[0225] Example 9: Preparation of target compound

[0226] Compound 10a (234 mg, 0.378 mmol) obtained in Example 6 was dissolved in saturated hydrochloric acid ethyl acetate (10 mL), stirred at room temperature for 12 h, filtered, and the filter cake was recrystallized and purified with EA or MeOH (20 mL) to obtain 11a. The synthesis methods of compounds 11b-v, 15a, 15b, 19a, 19b, 23a, (R)-23a, (S)-23a, 23b, 27, 30a-e, 40a-d, 44a-c and 48 were the same as the above operation steps.

[0227] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -Hydroxysuccinamide (11a)

[0228] White solid, yield 44.1%, mp 170-171°C. 1H NMR (400MHz, DMSO) δ10.30(s,1H),8.02–7.93(m,2H),7.53(d,J=7.4Hz,2H),7.42–7.30(m,4H),5.56(s,1H),4.48–4.37(m,2H),3.90(s,1H),3.4 1–3.30(m,2H),2.76(q,J=7.5Hz,2H),2.33(t,J=7.4Hz,2H),2.20(t,J= 7.4Hz,2H),1.92–1.83(m,2H),1.48–1.39(m,2H),1.18(t,J=7.5Hz,3H). 13 C NMR (101MHz, DMSO) δ174.32,174.06,170.81,170.04,169.99,164.87,164.78,164.40,158.46,136.66,129.12,128.67,128.63 ,116.43,115.15,93.78,87.59,53.64,47.47,46.93,45.68,32.04,30.55,29.62,29.24,27.58,14.01.HRMS-ESI(m / z)calc.for C 26 H 28 N7O4S[M–H] - 534.1929,found 534.1932.HPLC t R =6.19min (99.89% purity).

[0229] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (11b)

[0230] White solid, yield 55.8%, mp 171-173°C. 1H NMR(400MHz,DMSO)δ10.31(s,1H),8.01(s,1H),7.91(d,J=7.6Hz,1H),7.57–7.50 (m,2H),7.43–7.30(m,4H),5.56(s,1H),4.45–4.41(m,2H),3.91(d,J=8.6Hz,1H), 3.40–3.30(m,2H),2.76(q,J=7.5Hz,2H),2.08(t,J=7.4Hz,2H),2.01–1.94(m,2H ),1.91–1.85(m,2H),1.76–1.85(m,2H),1.51–1.40(m,2H),1.21(t,J=7.5Hz,3H). 13 C NMR (101MHz, DMSO) δ174.66,171.46,170.00,164.79,164.40,158.45,136.69,129.11,128.66,128.61,116.43,115.15,93.7 7,87.56,53.63,46.96,45.79,45.59,34.97,33.48,33.22,32.06,32.00,27.58,21.19,14.01,8.90.HRMS-ESI(m / z)calc.for C 27 H 30 N7O4S[MH] + 548.2085,found 548.2088.HPLC t R =6.65min (97.33% purity).

[0231] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 -Hydroxyadipamide (11c)

[0232] White solid, yield 34.5%, mp 173-175°C. 1H NMR (400MHz, DMSO) δ8.05(s,1H),7.94(d,J=7.6Hz,1H),7.54(d,J=7.4Hz,2H),7.43–7.30(m,4H),5.58(s,1H),4.43(d,J=14.0Hz,2H),3.91(s,1H) ,3.23–3.15(m,2H),2.76(q,J=7.6Hz,2H),2.07(d,J=7.6Hz,2H),1.98(d, J=9.2Hz,2H),1.88(d,J=12.7Hz,2H),1.48(s,6H),1.18(t,J=7.5Hz,3H). 13 C NMR (101MHz, DMSO) δ171.98,170.04,169.60,164.81,164.37,158.46,136.83,129.05,128.69,128.56,116.44,115.17,93 .77,87.54,53.59,47.04,46.95,45.54,35.87,32.71,32.06,28.84,27.57,25.70,25.52,14.00.HRMS-ESI(m / z)calc.for C 28 H 32 N7O4S[MH] + 562.2242,found 562.2246.HPLC t R =6.49min (98.83% purity).

[0233] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 7 -Hydroxypimelamide (11d)

[0234] White solid, yield 59.6%, mp 173-175°C. 1H NMR (400MHz, DMSO) δ10.47(s,1H),8.70(s,1H),8.14(s,1H),7.97(d,J=7.5Hz,1H) ,7.55(d,J=7.2Hz,2H),7.41–7.30(m,4H),5.62(s,1H),4.44(d,J=14.3,2H),3.90 (s,1H),3.33(d,J=13.4Hz,2H),2.75(q,J=7.5Hz,2H),2.07(t,J=7.3Hz,2H),1.95 (t,J=7.3Hz,2H),1.87(d,J=12.6Hz,2H),1.60–1.36(m,6H),1.21(q,J=7.1Hz,5H). 13 C NMR (101MHz, DMSO) δ171.94,170.05,169.57,164.81,164.38,158.45,136.78,129.07,128.69,128.58,116.44,115.16,93 .74,87.52,53.59,47.02,46.94,45.54,35.78,32.62,32.06,28.68,27.57,25.53,25.39,14.01.HRMS-ESI(m / z)calc.for C 29 H 35 N7O4S[M+Na] + 600.2375,found 600.2379.HPLC t R =12.41min (98.19% purity).

[0235] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 8 -Hydroxyoctanamide (11e)

[0236] Pale yellow solid, yield 35.2%, mp 177-179°C. 1H NMR (400MHz, DMSO) δ10.46 (s, 1H), 8.70 (s, 1H), 8.15 (s, 1H), 7.97 (d, J = 7.5Hz, 1H), 7 .55(d,J=7.3Hz,2H),7.35(dq,J=13.2,7.4Hz,4H),5.62(s,1H),4.51–4.38(m,2H),3 .90(s,1H),3.34(s,2H),2.75(q,J=7.5Hz,2H),2.06(q,J=6.9Hz,2H),1.94(t,J=7.2 Hz,2H),1.87(d,J=12.6Hz,2H),1.471.61–1.35(m,6H),1.23(dd,J=12.8,8.4Hz,7H). 13 C NMR (101MHz, DMSO) δ171.98,170.04,169.60,164.81,164.37,158.46,136.83,129.05,128.69,128.56,116.44,115.17,93 .77,87.54,53.59,47.04,46.95,45.54,35.87,32.71,32.06,28.84,27.57,25.70,25.52,14.00.HRMS-ESI(m / z)calc.for C 30 H 37 N7O4S[M+H] + 592.2706,found 592.2708.HPLC t R =6.89min (99.98% purity).

[0237] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 9 -Hydroxyazelazone (11f)

[0238] White solid, yield 48.2%, mp 177-179°C. 1H NMR (400MHz, DMSO) δ10.37(s,1H),7.98(s,1H),7.86(d,J=7.6Hz,1H),7.56–7.49(m,2H),7.43–7.29(m,4H),5.55(s,1H),4.47–4.42(m,2H) ),3.34(t,J=12.2Hz,2H),2.76(q,J=7.6Hz,2H),2.06(t,J=7.4Hz,2H),1.98–1.83(m,4H),1.55–1.35(m,6H),1.21(dd,J=14.7,7.1Hz,9H). 13 C NMR (101MHz, DMSO) δ171.97,169.99,169.57,164.78,164.40,158.48,136.67,129.12,128.67,116.43,115.14,93.79,87 .60,53.64,47.00,45.56,35.91,32.70,32.04,29.00,28.97,28.93,27.57,25.74,25.56,14.01.HRMS-ESI(m / z)calc.for C 31 H 39 N7O4S[M+Na] + 628.2682,found628.2681.HPLC t R =11.64min (99.9% purity).

[0239] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -Hydroxy-p-toluene diamide (11 g)

[0240] White solid, yield 67.6%, mp 214-216°C. 1H NMR (400MHz, DMSO) δ11.37(s,1H),8.50(d,J=7.7Hz,1H),7.98(s,1H),7.93(d,J =8.3Hz,2H),7.84(d,J=8.2Hz,2H),7.56–7.49(m,2H),7.44–7.29(m,4H),5.56( s,1H),4.65–4.52(m,2H),4.29–4.17(m,1H),3.34(q,J=4.3Hz,2H),2.77(q,J=7 .3Hz,2H),1.98(d,J=6.2Hz,2H),1.67(q,J=12.3Hz,2H),1.21(t,J=7.5Hz,3H)). 13 C NMR (101MHz, DMSO) δ169.98,165.47,164.87,164.44,163.90,158.51,137.20,136.62,135.48,129.15,128.68,128.65,127. 86,127.29,116.45,115.14,93.83,87.64,53.68,47.33,47.23,46.73,31.96,31.86,27.60,14.03.HRMS-ESI(m / z)calc.for C 30 H 29 N7O4S[M+Na] + 606.1905,found 606.1908.HPLC t R =6.73min (97.35% purity).

[0241] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-4-((4-(hydroxycarbamoyl)benzyl)oxy)benzamide (11h)

[0242] White solid, yield 53.0%, mp 202-204°C. 1H NMR (400MHz, DMSO) δ11.34(s,1H),8.33(d,J=7.6Hz,1H),8.19(s,1H),7.89(d,J=8.5Hz,2H) ,7.80(d,J=7.9Hz,2H),7.54(dd,J=16.1,7.7Hz,4H),7.43–7.30(m,4H),7.07(d,J=8.5Hz,2H ),5.65(s,1H),5.24(s,2H),4.64–4.52(m,2H),4.25–4.13(m,1H),3.30(d,J=13.0Hz,2H),2. 76(q,J=7.5Hz,2H),1.98–1.92(m,2H),1.66(dd,J=26.5,12.7Hz,2H),1.21(t,J=7.6Hz,3H). 13 C NMR(101MHz,DMSO)δ170.05,165.53,164.89,164.39,160.84,158.50,140 .34,136.80,132.73,129.71,129.06,128.70,128.57,127.87,127.57,12 7.49,116.46,115.18,114.72,93.78,87.59,69.20,60.24,53.62,47.49, 47.32,46.58,32.04,27.59,21.25,14.56,14.03.HRMS-ESI(m / z)calc.for C 37 H 34 N7O5S[MH] - 688.2348,found 688.2348.HPLC t R =4.86min (99.92% purity).

[0243] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(Hydroxycarbamoyl)phenyl)succinamide (11i)

[0244] White solid, yield 28.2%, mp 225-227°C. 1H NMR(400MHz,DMSO)δ11.12(s,1H),10.33(s,1H),8.93(s,1H),8.08–7.97(m,2H),7 .68(q,J=8.7Hz,3H),7.53(d,J=7.4Hz,2H),7.43–7.27(m,4H),5.57(s,1H),4.43( d,J=13.1Hz,2H),3.91(s,1H),2.75(q,J=7.5Hz,2H),2.61(t,J=7.1Hz,2H),2.43( t,J=7.2Hz,2H),1.88(d,J=12.7Hz,2H),1.54–1.37(m,2H),1.20(t,J=7.6Hz,3H). 13 C NMR (101MHz, DMSO) δ170.03,164.76,164.36,163.09,158.45,138.23,136.73,135.03,130.31,129.10,128.68,12 8.60,127.95,119.84,116.46,115.16,93.68,87.58,53.64,48.02,30.29,27.56,14.00.HRMS-ESI(m / z)calc.forC 33 H 33 N8O5S[MH] + 653.2300,found653.2302.HPLC t R =8.39min (99.06% purity).

[0245] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(Hydroxycarbamoyl)phenyl)glutaramide (11j)

[0246] White solid, yield 49.3%, mp 214-216°C. 1H NMR (400MHz, DMSO) δ10.25 (s, 1H), 8.02 (s, 1H), 7.94 (d, J = 7.6Hz, 1H), 7.70 (t, J = 6. 1Hz,4H),7.53(d,J=7.5Hz,2H),7.41–7.29(m,4H),5.56(s,1H),4.43(t,J=10.4Hz, 2H),3.92(d,J=10.4Hz,1H),3.40–3.28(m,2H),2.75(q,J=7.5Hz,2H),2.37(t,J=7. 4Hz,2H),2.15(t,J=7.4Hz,2H),1.92–1.79(m,4H),1.51–1.37(m,2H),1.20(s,3H). 13 C NMR (101MHz, DMSO) δ171.79,171.50,169.99,164.79,164.39,158.46,142.35,136.68,129.11,128.67,128.10,127.38,118 .72,116.43,115.15,93.77,87.57,53.63,46.97,45.60,36.20,35.12,32.08,27.58,21.64,14.01.HRMS-ESI(m / z)calc.for C 34 H 36 N8O5S[M+H] + 669.2612,found 669.2615.HPLC t R =10.52min(97.52%purity).

[0247] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 -(4-(Hydroxycarbamoyl)phenyl)adipamide (11k)

[0248] White solid, yield 33.1%, mp 211-213°C. 1H NMR (400MHz, DMSO) δ11.10(s,1H),10.19(s,1H),7.99–7.95(m,1H),7.90(d,J=7.6Hz,1H),7.7 4–7.64(m,4H),7.52(d,J=7.4Hz,2H),7.40–7.31(m,4H),5.54(s,1H),4.44(t,J=10.4Hz,2H),3 .97–3.89(m,1H),3.32(d,J=12.1Hz,2H),2.75(q,J=7.5Hz,2H),2.35(t,J=6.8Hz,2H),2.11(t ,J=6.8Hz,2H),1.97–1.81(m,2H),1.62–1.53(m,4H),1.47–1.37(m,2H),1.20(t,J=7.6Hz,3H). 13 CNMR(101MHz,DMSO)δ172.01,171.79,169.98,164.78,164.40,158.46,142.33,136.65,129.12,128.66,128.63,128.11,127.40,118 .72,116.43,115.14,93.79,87.59,53.66,46.97,45.60,36.73,35.75,32.08,27.58,25.48,25.18,14.01.HRMS-ESI(m / z)calc.forC 35 H 37 N8O5S[MH] - 681.2613,found681.2617.HPLC t R =6.4min (97.15% purity).

[0249] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(Hydroxycarbamoyl)phenyl)succinamide (11l)

[0250] White solid, yield 28.7%, mp 207-209°C. 1H NMR(400MHz,DMSO)δ11.18(s,1H),8.41(t,J=6.0Hz,1H),7.97–7.87(m,2H),7.7 0(d,J=7.9Hz,2H),7.51(d,J=7.4Hz,2H),7.42–7.29(m,6H),5.53(s,1H),4.47–4 .35(m,2H),4.30(d,J=5.9Hz,2H),3.95–3.87(m,1H),2.76(q,J=7.5Hz,2H),2.4 6–2.31(m,4H),1.88(d,J=12.7Hz,2H),1.51–1.36(m,2H),1.21(t,J=7.6Hz,3H). 13 C NMR (101MHz, DMSO) δ171.97,171.14,169.98,164.78,164.56,164.41,158.49,143.45,136.62,131.65,129.15,128.66,127.43,127 .33,116.43,115.14,93.81,87.63,53.67,46.93,45.64,42.22,32.07,31.98,31.24,31.15,27.58,14.01.HRMS-ESI(m / z)calc.for C 34 H 35 N8O5S[MH] - 667.2466,found 667.2467.HPLC t R =11.36min (99.30% purity).

[0251] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(Hydroxycarbamoyl)phenyl)glutaramide (11m)

[0252] White solid, yield 43.2%, mp 193-195°C. 1H NMR (400MHz, DMSO) δ11.19(s,1H),8.40(t,J=6.0Hz,1H),7.96(s,1H),7.88(d,J=7.6Hz,1H),7.70( d,J=8.1Hz,2H),7.52(d,J=7.1Hz,2H),7.42–7.26(m,6H),5.54(s,1H),4.48–4.37(m,2H),4.29(d,J =5.9Hz,2H),3.96–3.89(m,1H),3.37–3.31(m,2H),2.76(q,J=7.7Hz,2H),2.17(t,J=7.5Hz,2H),2. 10(t,J=7.5Hz,2H),1.94–1.83(m,2H),1.80–1.72(m,2H),1.48–1.40(m,2H),1.20(t,J=7.6Hz,3H). 13 CNMR(101MHz,DMSO)δ172.34,171.56,169.99,164.79,164.54,164.40,158.47,143.47,136.65,131.68,129.85,129.13,128.66,127.47,12 7.37,116.44,115.14,93.78,87.59,53.65,46.94,45.59,42.20,35.36,35.19,32.09,32.02,27.58,22.05,14.02.HRMS-ESI(m / z)calc.for C 35 H 37 N8O5S[MH] - 681.2613,found 681.2618.HPLC t R =6.21min (99.91% purity).

[0253] (E)-N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(3-(Hydroxyamino)-3-oxopropyl-1-en-1-yl)phenyl)succinamide (11n)

[0254] White solid, yield 30.8%, mp 212-214℃. 1H NMR(400MHz,DMSO)δ10.71(s,1H),10.14(s,1H),8.01–7.90(m,2H),7.64(d,J=8.3Hz,2H),7 .51(t,J=8.9Hz,4H),7.36(dt,J=22.5,7.6Hz,5H),6.36(d,J=15.8Hz,1H),5.54(s,1H),4.47 –4.36(m,2H),3.95–3.88(m,1H),3.34(d,J=12.0Hz,2H),2.76(q,J=7.5Hz,2H),2.60(t,J=7. 1Hz,2H),2.43(t,J=7.1Hz,2H),1.94–1.82(m,2H),1.54–1.39(m,2H),1.21(t,J=7.6Hz,3H). 13 C NMR (101MHz, DMSO) δ171.18,171.03,170.00,164.77,164.40,163.52,158.46,140.94,138.48,136.62,129.82,129.14,128.66,119 .41,117.65,116.43,115.14,93.79,87.60,53.66,46.92,45.71,32.13,32.08,32.01,30.71,27.58,14.02.HRMS-ESI(m / z)calc.for C 35 H 35 N8O5S[MH] + 679.2457,found 679.2462.HPLC t R =6.44min (97.25% purity).

[0255] 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-hydroxybenzamide (11o)

[0256] White solid, yield 26%, mp 201-203°C. 1H NMR (400MHz, DMSO) δ10.76(s,1H),9.03(s,1H),7.95(s,1H),7.53(dd,J=7.8,5.8Hz,4H),7.45(dd,J=11.7,4.0Hz,3H),7.41–7.28(m,4H),6.46(d,J =15.8Hz,1H),5.55(s,1H),4.43(d,J=13.3Hz,2H),3.86(s,2H),3.27(s,2 H), 2.86 (s, 1H), 2.75 (q, J = 7.6Hz, 2H), 1.44 (s, 2H), 1.20 (t, J = 7.6Hz, 3H). 13 CNMR(101MHz,DMSO)δ170.05,164.94,164.40,164.05,158.62,136.67,135.58,133.57,130.72,130.55,129.94,129.15,128.70,128.6 4,127.56,116.32,115.00,94.43,88.17,54.16,53.64,47.05,46.39,46.27,45.90,28.41,27.59,13.98,8.93.HRMS-ESI(m / z)calc.for C 30 H 32 N7O3S[M+H] + 570.2287,found 570.2297.HPLC t R =9.58min (97.78% purity).

[0257] (E)-3-4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-hydroxyacrylamide (11p)

[0258] White solid, yield 29.3%, mp 199-201℃. 1H NMR (400MHz, CDCl3) δ9.63 (s, 2H), 8.11 (d, J = 3.6Hz, 1H), 7.75–7.66 (m, 4H), 7.59 (d, J = 7. 2Hz,2H),7.53(d,J=15.8Hz,1H),7.47–7.36(m,4H),6.59(dd,J=15.9,1.7Hz,1H),5.65(s, 1H),4.70(d,J=13.4Hz,2H),4.28(t,J=6.0Hz,2H),3.47(d,J=6.0Hz,1H),3.24(t,J=12.9H z,2H),2.81(q,J=7.6Hz,2H),2.39–2.30(m,2H),1.88–1.73(m,2H),1.26(t,J=7.7Hz,3H). 13 C NMR (101MHz, DMSO) δ170.05,164.92,164.41,162.92,158.59,137.97,136.60,135.86,133.65,131.13,129.18,128.70,128.13,120.50 ,116.34,115.01,94.36,88.13,60.24,54.01,53.62,47.10,46.36,46.25,28.45,27.60,21.25,14.56,14.01.HRMS-ESI(m / z)calc.for C 32 H 33 N7O3S[M+H] + 596.2438,found 596.2440.HPLC t R =8.11min (98.52% purity).

[0259] 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(hydroxycarbamoyl)benzyl)carbamoyl)benzyl)piperidinium 4-ammonium salt (11q)

[0260] White solid, yield 28%, mp 197-199°C. 1H NMR (400MHz, DMSO) δ11.20(s,1H),9.66(s,2H),9.24(t,J=6.0Hz,1H),8.06(s,1H),7.98(d,J=8 .0Hz,2H),7.73(dd,J=10.7,8.1Hz,4H),7.58–7.52(m,2H),7.44–7.31(m,6H),5.61(s,1H),4.6 6(d,J=13.4Hz,2H),4.53(d,J=5.9Hz,2H),4.28(t,J=6.1Hz,2H),3.45(m,1H),3.20(t,J=12.8H z,2H),2.77(q,J=7.5Hz,2H),2.30(d,J=12.4Hz,2H),1.85–1.75(m,2H),1.21(t,J=7.8Hz,3H). 13 CNMR(101MHz,DMSO)δ170.05,166.17,164.92,164.51,164.41,158.59,143 .32,136.62,135.76,134.92,131.78,130.53,129.89,129.18,128.70,128 .01,127.68,127.55,127.42,116.34,115.02,94.36,88.14,54.09,53.62, 46.99,46.37,46.26,42.89,28.46,27.61,14.02.HRMS-ESI(m / z)calc.for C 38 H 39 N8O4S[M+H] + 703.2816,found 703.2821.HPLC t R =4.04min (96.86% purity).

[0261] (E)-1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(3-hydroxyamino)-3-oxopropyl-1-en-1-yl)benzyl)carbamoyl)benzyl)piperidin-4-ammonium salt (11r)

[0262] White solid, yield 68.4%, mp 207-208℃. 1H NMR (400MHz, DMSO) δ10.86(s,1H),9.77(s,2H),9.26(t,J=5.9Hz,1H),8.13(s,1H),7.99(d,J=7.9Hz, 2H),7.77(d,J=8.0Hz,2H),7.54(dd,J=13.3,7.6Hz,4H),7.44–7.24(m,7H),6.48(d,J=15.8Hz,1H),5 .63(s,1H),4.66(d,J=13.2Hz,2H),4.51(d,J=5.9Hz,2H),4.28(t,J=6.0Hz,2H),3.25–3.16(m,2H),3 .11–3.16(m,1H),2.77(q,J=7.7Hz,2H),2.36–2.27(m,2H),1.83–1.70(m,2H),1.21(t,J=7.6Hz,3H). 13 CNMR(101MHz,DMSO)δ170.07,166.15,164.92,164.40,163.20,158.57,141. 58,138.45,136.65,135.74,134.96,133.90,130.54,129.16,128.70,128.66 ,128.25,127.98,127.96,119.19,116.34,115.03,94.34,88.11,54.07,53.6 0,46.97,46.38,46.26,42.92,28.43,27.61,14.01.HRMS-ESI(m / z)calc.for C 40 H 41 N8O4S[M+H] + 729.2975,found729.2979.HPLC t R =4.02min (97.99% purity).

[0263] (E)-1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(3-hydroxyamino)-3-oxopropyl-1-en-1-yl)benzyl)oxy)benzyl)piperidin-4-ammonium salt (11s)

[0264] White solid, yield 58.4%, mp 187-189℃. 1H NMR (400MHz, DMSO) δ10.83(s,1H),9.35(s,2H),8.05(s,1H),7.58(d,J=7.9Hz,2H),7.54(d,J=7 .7Hz,4H),7.50–7.46(m,2H),7.46–7.30(m,5H),7.08(d,J=8.5Hz,2H),6.49(d,J=15.8Hz,1H), 5.60(s,1H),5.18(s,2H),4.65(d,J=13.3Hz,2H),4.14(t,J=5.9Hz,2H),3.39(s,2H),3.19(t,J =13.0Hz,2H),2.77(q,J=7.6Hz,2H),2.25(s,2H),1.72(t,J=13.7Hz,2H),1.21(t,J=7.6Hz,3H). 13 CNMR(101MHz,DMSO)δ170.04,164.92,164.41,159.07,158.59,138.69,138.26,136.60,134.91,132.14,129.18,128.69,128.55,128.06,124 .67,119.80,116.33,115.44,115.00,94.37,88.13,69.27,53.77,53.63,46.98,46.35,46.26,28.46,27.60,14.01.HRMS-ESI(m / z)calc.for C 40 H 41 N8O4S[M+Na] + 724.2682,found 724.2676.HPLC t R =6.45min (97.29% purity).

[0265] 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((7-(hydroxyamino)-7-oxoheptyl)oxy)benzyl)piperidin-4-ammonium salt (11t)

[0266] White solid, yield 45.2%, mp 191-193℃. 1H NMR (400MHz, DMSO) δ10.38(s,1H),9.36–9.31(m,2H),8.05(s,1H),7.53(t,J=8.4Hz,4H),7.46–7.33(m ,4H),7.00(d,J=8.2Hz,2H),5.60(s,1H),4.65(d,J=13.3Hz,2H),4.14(t,J=6.0Hz,2H),3.98(t,J=6.5H z,2H),3.20(t,J=12.7Hz,2H),2.77(q,J=7.5Hz,2H),2.27(d,J=10.7Hz,2H),1.95(t,J=7.3Hz,2H),1. 78–1.66(m,5H),1.54–1.48(m,2H),1.43–1.38(m,2H),1.34–1.27(m,2H),1.20(dt,J=14.5,7.3Hz,5H). 13 C NMR (101MHz, DMSO) δ170.07,169.58,164.92,164.41,159.57,158.58,136.60,132.12,129.17,128.69,124.16,116.33,115.02,114.98,9 4.34,88.10,67.94,53.71,53.62,47.00,46.37,46.26,32.66,28.95,28.78,28.45,27.60,25.69,25.52,14.01.HRMS-ESI(m / z)calc.for C 36 H 43 N7O4S[M+H] + 670.3170,found670.3177.HPLC t R =7.85min (98.24% purity)

[0267] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-N-hydroxypyrimidine-5-carboxamide (11u)

[0268] White solid, yield 38.3%, mp 185-187℃. 1H NMR (400MHz, DMSO) δ8.66(s,2H),7.95(s,1H),7.90(d,J=7.8Hz,1H),7.52(d,J=7.4Hz,2H),7.41–7.31(m,4H),5.55(s,1H),4.58–4.49(m,2H) ,4.18(t,J=9.2Hz,1H),3.37(dd,J=11.2,6.5Hz,2H),2.77(q,J=7.6Hz,2H),2.03(d,J=12.8Hz,2H),1.65–1.55(m,2H),1.22(t,J=7.6Hz,3H). 13 C NMR (101MHz, DMSO) δ170.04,164.80,164.48,162.35,158.42,136.51,129.17,128.68,116.45,1 15.39,115.15,93.77,87.55,53.63,47.76,46.99,31.77,27.58,14.00.HRMS-ESI(m / z)calc.for C 28 H 29 N9O3S[M+Na] + 580.1811,found 580.1810.HPLC t R =5.74min (97.25% purity).

[0269] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)(methyl)amino)-N-hydroxypyrimidine-5-carboxamide (11v)

[0270] White solid, yield 36.1%, mp 185-187℃. 1 H NMR (400MHz, DMSO) δ11.20(s,1H),9.03(s,1H),8.73(s,2H),8.07(s,1H),7.55(d,J=7.4Hz,2H),7.39–7.30(m,4H),5.61(s, 1H), 5.00 (s, 1H), 4.76 (d, J = 13.2Hz, 2H), 3.01 (s, 3H), 2.77 (q, J = 8.0Hz, 2H), 1.81 (d, J = 22.1Hz, 4H), 1.22 (t, J = 7.6Hz, 3H). 13C NMR (101MHz, DMSO) δ170.02,164.80,164.40,161.99,158.37,157.48,136.73,129.11,128.66,116.51,1 15.17,114.92,93.84,87.63,53.64,52.34,47.62,29.49,29.06,27.59,14.01.HRMS-ESI(m / z)calc.for C 28 H 29 N9O3S[M+Na] + 597.1965,found 594.1967.HPLC t R =4.52min(95.29%purity)

[0271] Tert-butyl-4-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)piperidin-1-carbamate (12a)

[0272] White solid, yield 38.3%. 1 H NMR(400MHz,DMSO)δ7.89(s,1H),7.53–7.46(m,2H),7.44–7.28(m,4H),5.48(s,1H),4.69(s,1H),4.05 (s,2H),3.13(s,3H),2.78(q,J=7.7Hz,2H),1.67(d,J=9.4Hz,4H),1.41(s,9H),1.22(t,J=7.6Hz,3H).

[0273] Tert-butyl-4-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)methyl)(methyl)amino)piperidin-1-carbamate (12b)

[0274] White solid, yield 42.8%. 1 H NMR(400MHz,DMSO)δ7.89(s,1H),7.53–7.46(m,2H),7.44–7.28(m,4H),5.48(s,1H),4.69(s,1H),4.05 (s,2H),3.13(s,3H),2.78(q,J=7.7Hz,2H),1.67(d,J=9.4Hz,4H),1.41(s,9H),1.22(t,J=7.6Hz,3H).

[0275] 4-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)piperidin-1-ammonium salt (13a)

[0276] White solid, yield 56.2%. 1 H NMR (400MHz, DMSO) δ8.82(d,J=37.0Hz,2H),8.08(s,1H),7.53(d,J=7.5Hz,2H),7.46–7.34(m,4H),5.53(s,1H),4.89(s,1H),3. 17(s,3H),3.15–3.05(m,2H),2.80(t,J=7.6Hz,2H),2.09(dd,J=19.2,8.3Hz,2H),1.80(t,J=14.8Hz,2H),1.23(t,J=7.6Hz,3H).

[0277] 4-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)methyl)(methyl)amino)piperidin-1-ammonium salt (13b)

[0278] White solid, yield 41.7%. 1 H NMR (400MHz, DMSO) δ8.74(s,2H),8.04(s,1H),7.54(d,J=7.1Hz,2H),7.45–7.34(m,4H),5.48(s,1H),3.89(dd,J=14.0,8.3Hz,1H),3.63–3 .56(m,1H),3.27(d,J=12.3Hz,2H),2.83–2.72(m,3H),2.03(d,J=9.4Hz,1H),1.75–1.66(m,2H),1.56–1.42(m,2H),1.22(t,J=7.7Hz,3H).

[0279] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)piperidin-1-yl)-N-((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (14a)

[0280] White solid, yield 37.2%. 1H NMR (400MHz, DMSO) δ11.51(s,1H),8.70(s,2H),7.90(s,1H),7.51(d,J=7. 0Hz,2H),7.43–7.32(m,4H),5.52(s,1H),4.96(s,1H),4.87(d,J=12.6Hz,3 H),4.04(s,1H),3.53(d,J=11.4Hz,1H),3.12(s,4H),2.80(t,J=7.6Hz,2H) ,1.80–1.74(m,3H),1.73–1.69(m,2H),1.55(s,3H),1.23(t,J=7.6Hz,3H).

[0281] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)methyl)(methyl)amino)piperidin-1-yl)-N-((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (14b)

[0282] White solid, yield 45.3%. 1 H NMR (400MHz, DMSO) δ11.51 (s, 1H), 8.69 (s, 2H), 7.92 (s, 1H), 7.46 (d, J = 7.5Hz, 2H), 7.41–7. 32(m,4H),7.21(dd,J=8.4,6.3Hz,1H),5.52(d,J=4.4Hz,1H),4.96(d,J=3.4Hz,1H),4.76(t, J=15.8Hz,2H),4.03(s,1H),3.92(dd,J=14.1,7.8Hz,1H),3.55(dd,J=14.3,6.9Hz,2H),2.9 5–2.83(m,2H),2.77(d,J=7.7Hz,2H),2.10(s,1H),1.75–1.49(m,8H),1.21(t,J=7.6Hz,3H).

[0283] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)piperidin-1-yl)-N-hydroxypyrimidine-5-carboxamide (15a)

[0284] White solid, yield 37.2%, mp 172-173°C. 1H NMR (400MHz, DMSO) δ11.28(s,1H),9.06(s,1H),8.73(s,2H),8.24(s,1H),7.60–7.50(m,2H),7.45–7.30(m,4H),5 .66(s,1H),4.87(d,J=25.6Hz,3H),3.10(s,5H),2.77(d,J=7.6Hz,2H),1.72(d,J=34.2Hz,4H),1.27–1.18(m,3H). 13 C NMR (101MHz, DMSO) δ170.13,164.41,162.37,161.71,158.61,157.72,136.58,129.09,128.65,128.56,116.75,115 .30,114.85,93.61,87.03,56.28,53.48,43.05,42.96,33.60,29.00,28.75,27.62,14.00.HRMS-ESI(m / z)calc.for C 28 H 29 N9O3S[M+Na] + 594.2007,found594.2002.HPLC t R =4.67min (96.98% purity).

[0285] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)(methyl)amino)piperidin-1-yl)-N-hydroxypyrimidine-5-carboxamide (15b)

[0286] White solid, yield 32.0%, mp 173-175℃. 1 H NMR (400MHz, DMSO) δ8.71 (s, 2H), 8.15 (s, 1H), 7.50 (d, J = 7.5Hz, 2H), 7.36 (s, 1H), 7.28(t,J=7.5Hz,2H),7.18(t,J=7.3Hz,1H),5.61(s,1H),4.74(t,J=16.8Hz,2H), 3.93(dd,J=14.2,7.8Hz,1H),3.54(dd,J=17.9,8.3Hz,2H),3.38(s,2H),2.87(q,J =13.4Hz,3H),2.76(q,J=7.4Hz,2H),2.08(s,1H),1.68–1.56(m,2H),1.20(m,5H). 13C NMR (101MHz, DMSO) δ170.13,164.50,164.34,161.67,157.91,157.66,144.73,136.87,134.34,129.01,128.52,128.39,116 .90,115.33,114.62,93.22,86.12,56.76,53.48,43.79,41.14,35.49,29.15,29.01,27.63,13.97.HRMS-ESI(m / z)calc.for C 28 H 29 N9O3S[M+Na] + 608.2160,found 608.2155.HPLC t R =4.55min (95.28% purity).

[0287] Tert-butyl-4-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazine-1-carbamate (16)

[0288] White solid, yield 81.2%. 1 H NMR(400MHz,DMSO)δ7.90(s,1H),7.54–7.49(m,2H),7.45–7.29(m,5H),5.53(s,1H),3.89(t ,J=5.3Hz,4H),3.25–3.18(m,4H),2.77(q,J=7.6Hz,3H),1.39(s,9H),1.21(t,J=7.5Hz,3H).

[0289] 4-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazine-1-ammonium salt (17)

[0290] White solid, yield 90.2%. 1 H NMR (400MHz, DMSO) δ11.23(s,1H),7.87(s,1H),7.57(d,J=7.7Hz,2H),7.54–7.46(m,3H),7.43–7.27(m,6H),6.50(d,J=15.9Hz,1H),5.51(s, 1H),4.92(s,1H),3.97(s,1H),3.90(s,4H),3.57(s,2H),3.53(s,1H), 2.76(q,J=7.6Hz,2H),1.70(s,3H),1.55(s,3H),1.20(t,J=7.6Hz,3H).

[0291] (E)-3-(4-((4-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)methyl)phenyl)-N-((tetrahydro-2H-pyran-2-yl)oxy)acrylamide (18a)

[0292] White solid, yield 48.8%. 1 H NMR (400MHz, DMSO) δ11.23(s,1H),7.87(s,1H),7.57(d,J=7.7Hz,2H),7.54–7.46(m,3H),7.43–7.27(m,6H),6.50(d,J=15.9Hz,1H),5.51(s, 1H),4.92(s,1H),3.97(s,1H),3.90(s,4H),3.57(s,2H),3.53(s,1H), 2.76(q,J=7.6Hz,2H),1.70(s,3H),1.55(s,3H),1.20(t,J=7.6Hz,3H).

[0293] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)-N-((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (18b)

[0294] White solid, yield 50.2%. 1 H NMR (400MHz, DMSO) δ11.63(s,1H),8.77(s,2H),7.98(s,1H),7.54(d,J=7.5Hz,2H),7.40(dd,J=15.2,7.7Hz,4H),5.59 (s,1H),4.98(s,1H),4.03(s,6H),4.00(s,4H),2.79(q,J=7.6Hz,2H),1.72(s,3H),1.55(s,3H),1.23(t,J=7.6Hz,3H).

[0295] (E)-3-(4-((4-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)methyl)phenyl)-N-hydroxyacrylamide (19a)

[0296] White solid, yield 30.3%, mp 187-189°C. 1H NMR (400MHz, DMSO) δ10.81(s,1H),9.04(s,1H),7.93(s,1H),7.55(d,J=7.7Hz,2H),7.50(d,J=7.3Hz,2H),7.46(d,J=15.5Hz,1H) ,7.40–7.29(m,7H),6.48(d,J=15.8Hz,1H),5.54(s,1H),3.90(s,4H),3.56(s,2H),2.75(q,J=7.4Hz,2H),1.20(t,J=7.6Hz,3H). 13 C NMR (101MHz, DMSO) δ169.96,164.79,164.44,158.51,136.55,129.97,129.13,128.64,127.91,119.3 4,116.37,115.07,94.07,87.85,61.74,53.63,52.86,47.93,27.55,13.98.HRMS-ESI(m / z)calc.for C 31 H 31 N7O3S[M+Na] + 604.2058,found 604.2059.HPLC t R =5.16min (95.27% purity).

[0297] 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)-N-hydroxypyrimidine-5-carboxamide (19b)

[0298] White solid, yield 31.2%, mp 211-213℃. 1 H NMR (400MHz, DMSO) δ11.16(s,1H),8.75(s,2H),7.97(s,1H),7.54(d,J=7.1Hz,2H),7.45–7.34(m,4H),5.58( s,1H),4.04(dd,J=7.3,3.6Hz,4H),3.98(dd,J=7.3,3.6Hz,4H),2.79(q,J=7.5Hz,2H),1.23(t,J=7.6Hz,3H). 13C NMR (101MHz, DMSO) δ170.02,164.91,164.39,162.19,161.68,160.15,158.62,157.62,136.42,129.21,128.7 3,128.70,116.37,115.52,115.06,94.14,87.93,53.71,47.31,43.54,27.57,14.01.HRMS-ESI(m / z)calc.for C 26 H 25 N9O3S[M+Na] + 566.1659,found566.1655.HPLC t R =4.51min (96.89% purity).

[0299] Tert-butyl-4-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)-1-carbamate (20)

[0300] White solid, yield 77.8%. 1 H NMR (400MHz, DMSO) δ7.92(s,1H),7.55–7.48(m,2H),7.41–7.30(m,4H),6.93(t,J=5.9Hz,1H),5.53(s,1H),4.55(d,J=13.3Hz,2 H),3.11(t,J=12.3Hz,2H),2.86(t,J=6.1Hz,2H),2.75(q,J=7.5Hz,2H),1.76(d,J=12.8Hz,3H),1.39(s,9H),1.25–1.16(m,5H).

[0301] Tert-butyl-(R)-((1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)-1-carbamate ((R)-20)

[0302] White solid, yield 63.5%. 1H NMR (400MHz, DMSO) δ7.92(s,1H),7.52(dt,J=6.0,1.4Hz,2H),7.42–7.28(m,4H),6.94(q,J=5.7Hz,1H),5.54(s,1H),4.55(d,J=13.3Hz ,2H),3.10(d,J=10.5Hz,2H),2.85(q,J=6.2Hz,2H),2.75(q,J=7.5Hz,2H),1.80–1.68(m,3H),1.38(d,J=5.7Hz,9H),1.29–1.15(m,5H).

[0303] Tert-butyl-(S)-((1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)-1-carbamate ((S)-20)

[0304] White solid, yield 66.2%. 1 H NMR (400MHz, DMSO) δ7.92(s,1H),7.55–7.46(m,2H),7.42–7.28(m,4H),6.93(t,J=5.9Hz,1H),5.54(s,1H),4.55(d,J=13.2Hz,2H) ,3.11(t,J=12.6Hz,2H),2.85(q,J=6.3Hz,2H),2.75(q,J=7.6Hz,2H),1.76–1.70(m,2H),1.38(d,J=5.6Hz,9H),1.29–1.15(m,5H).

[0305] (1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methylammonium salt (21)

[0306] White solid, yield 88.2%. 1 H NMR (400MHz, DMSO) δ7.94(s,1H),7.81(s,3H),7.52(d,J=7.5Hz,2H),7.43–7.31(m,4H),5.54(s,1H),4.58(d,J=1 3.2Hz,2H),3.17(d,J=5.0Hz,2H),3.10(s,2H),2.76(d,J=8.2Hz,2H),1.88(t,J=13.2Hz,3H),1.35–1.17(m,5H).

[0307] (R)-(1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methylammonium salt ((R)-21)

[0308] White solid, yield 67.6%. 1 H NMR(400MHz,DMSO)δ8.08(s,4H),7.54(d,J=7.5Hz,2H),7.43–7.29(m,4H),5.57(s,1H),4.57(d,J=13.4Hz,2H), 3.14(q,J=13.7Hz,2H),2.76(q,J=6.0Hz,4H),1.88(d,J=12.5Hz,3H),1.43–1.28(m,2H),1.20(t,J=7.6Hz,3H).

[0309] (S)-(1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methylammonium salt ((S)-21)

[0310] White solid, yield 58.5%. 1 H NMR (400MHz, DMSO) δ8.11(s,4H),7.57–7.52(m,2H),7.41–7.32(m,4H),5.58(s,1H),4.57(d,J=13.5Hz,2H ),3.14(q,J=13.7Hz,2H),2.79–2.73(m,4H),1.91–1.86(m,3H),1.40–1.30(m,2H),1.20(t,J=7.6Hz,3H).

[0311] (E)-3-(4-((((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-(tetrahydro-2H-pyran-2-yl)oxy)acrylamide (22a)

[0312] White solid, yield 53.2%. 1H NMR (400MHz, DMSO) δ11.22 (s, 1H), 7.92 (s, 1H), 7.51 (d, J = 8.3Hz, 4H), 7.45–7. 34(m,5H),6.48(d,J=15.6Hz,1H),5.52(s,1H),4.91(s,1H),4.57(d,J=13.5Hz, 2H),3.96(s,1H),3.76(s,1H),3.53(d,J=11.4Hz,1H),3.17–3.03(m,2H),2.72 (d,J=7.6Hz,2H),1.85(s,2H),1.69(s,3H),1.54(s,3H),1.20(t,J=7.5Hz,3H).

[0313] (E)-3-(4-((((1-(6-(((R)-2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-(tetrahydro-2H-pyran-2-yl)oxy)acrylamide ((R)-22a)

[0314] White solid, yield 33.0%, 1 H NMR (400MHz, DMSO) δ11.22(s,1H),7.92(s,1H),7.58–7.48(m,5H),7.48–7.28(m ,8H),6.50(d,J=15.8Hz,1H),5.53(s,1H),4.91(s,1H),4.56(d,J=13.3Hz,2H), 3.96(s,1H),3.81(s,2H),3.61–3.47(m,2H),3.17–3.04(m,2H),2.75(q,J=7.5H z,2H),1.87(d,J=12.2Hz,2H),1.70(s,3H),1.54(s,3H),1.19(t,J=7.7Hz,3H).

[0315] (E)-3-(4-((((1-(6-(((S)-2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-(tetrahydro-2H-pyran-2-yl)oxy)acrylamide ((S)-22a)

[0316] White solid, yield 35.2%. 1H NMR (400MHz, DMSO) δ11.26(s,1H),7.95(s,1H),7.57(d,J=7.9Hz,2H),7.52(d,J=7.3H z,3H),7.47(d,J=7.3Hz,2H),7.41–7.32(m,4H),6.51(d,J=15.8Hz,1H),5.55(s,1H), 4.92(s,1H),4.46(d,J=13.1Hz,2H),3.90(s,4H),3.05(s,2H),2.76(q,J=7.5Hz,2H), 2.02(s,2H),1.72(d,J=10.7Hz,3H),1.55(s,3H),1.47(s,2H),1.20(t,J=7.4Hz,3H).

[0317] 2-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)-N-((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (22b)

[0318] White solid, yield 48.3%. 1 H NMR (400MHz, DMSO) δ11.49(s,1H),8.63(d,J=11.4Hz,2H),7.98(t,J=6.2Hz,1H ),7.92(s,1H),7.51(d,J=7.6Hz,2H),7.41–7.29(m,4H),5.53(s,1H),4.95(s,1 H),4.57(d,J=13.4Hz,2H),4.03(s,1H),3.28(s,2H),2.75(q,J=7.4Hz,3H),1. 95(s,1H),1.83(d,J=12.8Hz,2H),1.71(s,3H),1.54(s,3H),1.35–1.22(m,5H).

[0319] (E)-1-(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N-(4-(3-(hydroxyamino)3-oxopropane-1-en-1-yl)benzyl)methanamine salt (23a)

[0320] White solid, yield 33.2%, mp 203-205℃. 1H NMR(400MHz,DMSO)δ10.85(s,1H),9.36(s,2H),8.02(s,1H),7.63(s,4H),7.57–7.45( m,3H),7.43–7.28(m,4H),6.53(d,J=15.8Hz,1H),5.56(s,1H),4.56(d,J=13.3Hz,2H) ,4.17(t,J=5.6Hz,2H),3.14(q,J=13.0Hz,2H),2.84(d,J=6.6Hz,2H),2.75(q,J=7.6H z,2H),2.15(s,1H),1.92(d,J=12.8Hz,2H),1.37–1.24(m,2H),1.20(t,J=7.6Hz,3H). 13 C NMR (101MHz, DMSO) δ170.03,164.81,164.34,162.93,158.51,137.98,136.69,135.86,133.39,131.31,129.11,128.68,128.61,128.05 ,120.48,116.43,115.14,93.77,87.73,53.64,51.57,50.46,49.06,47.86,47.72,32.84,30.00,27.56,14.01.HRMS-ESI(m / z)calc.for C 33 H 35 N7O3S[M+H] + 610.2570,found 610.2552.HPLC t R =5.76min (97.46% purity).

[0321] (R,E)-1-(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N-(4-(3-(hydroxyamino)3-oxopropane-1-en-1-yl)benzyl)methanamine salt ((R)-23a)

[0322] White solid, yield 50.2%, mp 201-202℃. 1H NMR(400MHz,DMSO)δ10.78(s,1H),9.04(s,1H),7.94(s,1H),7.58(d,J=7.8Hz,2H), 7.53(d,J=1.9Hz,2H),7.51–7.42(m,3H),7.42–7.28(m,4H),6.48(d,J=15.8Hz,1H), 5.54(s,1H),4.56(d,J=13.2Hz,2H),3.99(s,2H),3.14(d,J=8.8Hz,2H),2.75(q,J= 7.6Hz,2H),2.67(s,2H),1.98(s,1H),1.89(d,J=13.2Hz,2H),1.21(q,J=7.1Hz,5H). 13 C NMR(101MHz,DMSO)δ170.03,164.83,164.30,162.96,158.50,137.93,136.77 ,135.84,133.40,131.36,131.31,129.07,128.73,128.68,128.56,127.99,1 20.53,116.42,115.14,93.77,87.73,56.49,53.64,51.48,50.41,47.90,47. 73,45.78,32.81,30.05,27.55,19.03,13.99,8.88.HRMS-ESI(m / z)calc.forC 33 H 35 N7O3S[M+H] + 610.2570,found 610.2572.HPLC t R =5.78min (98.02% purity).

[0323] (S,E)-1-(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N-(4-(3-(hydroxyamino)3-oxopropane-1-en-1-yl)benzyl)methanamine salt ((S)-23a)

[0324] White solid, yield 47.5%, mp 202-204℃. 1H NMR(400MHz,DMSO)δ10.86(s,1H),9.29(s,2H),8.00(s,1H),7.63(s,3H),7.53(dd, J=7.0,1.9Hz,2H),7.50(s,1H),7.41–7.30(m,4H),6.53(d,J=15.8Hz,1H),5.56(s, 1H),4.56(d,J=13.3Hz,2H),4.17(t,J=5.6Hz,2H),3.21–3.07(m,3H),2.85(d,J=6. 0Hz, 2H), 2.75 (q, J = 7.6Hz, 2H), 2.14 (s, 1H), 1.96–1.88 (m, 2H), 1.31–1.17 (m, 5H). 13 C NMR(101MHz,DMSO)δ170.03,164.83,164.31,162.95,158.50,137.95,136 .75,135.83,133.40,131.35,131.30,129.08,128.75,128.68,128.57,12 8.00,120.50,116.43,115.15,93.76,87.73,56.49,53.63,51.49,50.41, 47.89,47.73,32.81,30.06,30.03,27.55,14.00.HRMS-ESI(m / z)calc.for C 33 H 35 N7O3S[2M+H] + 1219.5122,found 1219.5129.HPLC t R =5.82min (99.24% purity).

[0325] 2-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)-N-hydroxypyrimidine-5-carboxamide (23b)

[0326] White solid, yield 35.1%, mp 178-180℃. 1H NMR (400MHz, DMSO) δ8.65(s,2H),8.07(s,1H),7.96(s,1H),7.52(dd,J=7.1,1.9Hz,2H),7.40–7.28(m,4H),5.55(s,1H),4.58(d,J =13.3Hz,2H),3.29(d,J=5.7Hz,2H),3.12(d,J=12.6Hz,2H),2.80–2.72(m,2H),2.00–1.92(m,1H),1.89–1.80(m,2H),1.21(m,5H). 13 C NMR (101MHz, DMSO) δ170.03,164.69,164.37,162.85,158.42,157.61,136.73,129.09,128.67,128.58,116.48,115 .20,93.58,87.50,53.61,49.06,48.10,46.36,45.90,35.53,30.25,30.12,27.56,14.00.HRMS-ESI(m / z)calc.for C 28 H 29 N9O3S[M+H] + 572.2155,found 572.2150.HPLC t R =11.25min (99.40% purity).

[0327] Tert-butyl (2-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)ethyl)carbamate (24)

[0328] White solid, yield 85.2%. 1 H NMR (400MHz, DMSO) δ7.87 (s, 1H), 7.57–7.48 (m, 2H), 7.45–7.29 (m, 4H), 6.96 (t, J = 6.1Hz, 1H), 5.59 (s, 1H), 3. 87–3.74(m,2H),3.40(s,3H),3.21(t,J=5.7Hz,2H),2.76(q,J=7.5Hz,2H),1.34(s,9H),1.23(d,J=7.5Hz,3H).

[0329] 2-((6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)ethyl-1-ammonium salt (25)

[0330] White solid, yield 91.2%. 1 H NMR(400MHz,DMSO)δ8.27(s,4H),7.65–7.57(m,2H),7.47–7.31(m,4H),4.09(s,2H) ), 3.43 (d, J = 1.1Hz, 3H), 3.09 (s, 2H), 2.78 (q, J = 7.5Hz, 2H), 1.21 (d, J = 7.5Hz, 3H).

[0331] (E)-3-(4-(((2-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)ethyl)amino)methyl)phenyl)-N-(tetrahydro-2H-pyran-2-yl)oxy)acrylamide (26)

[0332] White solid, yield 51.5%. 1 H NMR (400MHz, DMSO) δ11.23(s,1H),7.96(s,1H),7.50(d,J=7.0Hz,4H),7.44(s,1H),7.42–7.27(m,6H),6.47(d,J=15.9Hz,1H),5.52(s,1H),4.91(s ,1H),3.99–3.91(m,2H),3.85(s,1H),3.72(s,2H),3.53(d,J=11.3Hz,2H ),2.74(q,J=7.4Hz,2H),1.70(s,3H),1.54(s,3H),1.18(d,J=7.1Hz,3H).

[0333] (E)-2-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)-N-(4-(3-(hydroxyamino)-3-oxopropane-1-en-1-yl)benzyl)ethyl-1-ammonium salt (27)

[0334] White solid, yield 32.7%, mp 197-199℃. 1H NMR (400MHz, DMSO) δ10.84(s,1H),9.51(s,2H),9.09(s,1H),8.32(s,1H),7.71–7.58(m,6H),7.55–7.52(m,1H),7.48(d,J=15.8Hz,1H),7 .41–7.32(m,3H),6.52(d,J=15.9Hz,1H),5.80(s,1H),4.26–4.11(m,4H),3.28–3.20(m,2H),2.79(q,J=7.6Hz,2H),1.21(t,J=7.6Hz,3H). 13 C NMR (101MHz, DMSO) δ170.55,164.48,163.01,158.18,138.04,136.44,135.92,133.44,131.14,129.17,128.79,128.7 1,128.12,120.48,116.82,115.15,93.97,87.12,52.94,50.58,48.06,44.46,27.62,13.96.HRMS-ESI(m / z)calc.for C 28 H 29 N9O3S[M+H] + 570.2282,found 570.2284.HPLC t R =5.18min (96.49% purity).

[0335] Example 10: Preparation of target compounds 30a-e

[0336] 5-((2-((tert-Butoxycarbamoyl)amino)phenyl)amino)-5-oxopentanoic acid (28a)

[0337] The corresponding carboxylic acid (2.66mmol), EDCI (615mg, 3.2mmol), DMAP (422mg, 3.45mmol), and tert-butyl 2-aminophenylcarboxylate (555mg, 2.66mmol) were added to 15mL CH2Cl2 and reacted at room temperature for 8 hours. After the reaction, the reaction solution was washed with 1M hydrochloric acid aqueous solution (100mL) and saturated sodium chloride aqueous solution (100mL) in sequence. The organic phase was then dried over anhydrous Na2SO4 and concentrated. 28a was obtained by purification by silica gel column chromatography. The synthesis method of compounds 28d-e was the same as the operation steps. White solid, yield 25.8%. 1H NMR (400MHz, DMSO) δ12.10(s,1H),9.46(s,1H),8.32(s,1H),7.54(d,J=8.0Hz,1H),7.41(d,J=7.8Hz,1 H),7.19–7.05(m,2H),2.40(t,J=7.3Hz,2H),2.29(t,J=7.4Hz,2H),1.83(p,J=7.3Hz,2H),1.46(s,9H).

[0338] 4-((2-((tert-Butoxycarbamoyl)amino)phenyl)carbamoyl)benzoic acid (28b)

[0339] White solid, yield 78.1%. 1 H NMR (400MHz, DMSO) δ13.31 (s, 1H), 9.98 (s, 1H), 8.74 (s, 1H), 8.11–8.05 (m, 4H), 7.56 (t, J = 9.4Hz, 2H), 7.21–7.14 (m, 2H).

[0340] 2-(4-((2-((tert-Butoxycarbamoyl)amino)phenyl)carbamoyl)phenyl)acetic acid (28c)

[0341] White solid, yield 96.4%. 1 H NMR (400MHz, DMSO) δ9.83 (s, 1H), 8.69 (s, 1H), 7.90 (d, J = 8.2Hz, 2H), 7.57–7 .51(m,2H),7.46–7.40(m,2H),7.22–7.15(m,2H),3.69(s,2H),1.45(s,9H).

[0342] Tert-butyl (2-(4-formylbenzamido)phenyl)carbamate (28d)

[0343] Yellow solid, yield 43.9%. 1 H NMR (400MHz, DMSO) δ10.13(s,1H),10.01(s,1H),8.74(s,1H),8.15(d,J=8.0Hz ,2H),8.07(d,J=8.3Hz,2H),7.58–7.50(m,2H),7.25–7.11(m,2H),1.44(s,9H).

[0344] Tert-butyl (2-(2-chloropyrimidine-5-carboxamido)phenyl)carbamate (28e)

[0345] Yellow solid, yield 72.7%. 1H NMR (400MHz, DMSO) δ10.13(s,1H),10.01(s,1H),8.74(s,1H),8.15(d,J=8.0Hz ,2H),8.07(d,J=8.3Hz,2H),7.59–7.51(m,2H),7.26–7.15(m,2H),1.44(s,9H).

[0346] Tert-butyl (2-(5-((1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-5-oxopentanylamino)phenyl)carbamate (29a)

[0347] White solid, yield 84.3%. 1 H NMR(400MHz,DMSO)δ9.43(s,1H),8.38(s,1H),7.96–7.87(m,2H),7.58–7 .49(m,3H),7.47–7.29(m,5H),7.19–7.08(m,2H),5.53(s,1H),4.52–4.3 9(m,2H),3.94(s,1H),2.76(q,J=7.5Hz,2H),2.37(t,J=7.4Hz,2H),2.16 (t,J=7.5Hz,2H),1.94–1.80(m,4H),1.46(s,9H),1.22(t,J=6.4Hz,3H).

[0348] Tert-butyl (2-(4-((1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)carbamoyl)benzamido)phenyl)carbamate (29b)

[0349] White solid, yield 82.2%. 1 H NMR (400MHz, DMSO) δ10.01(s,1H),8.76(s,1H),8.58(d,J=7.6Hz,1H),8.05(q,J= 8.6Hz,5H),7.57–7.52(m,4H),7.39(t,J=7.3Hz,2H),7.34(dd,J=4.9,2.3Hz,2H), 7.24–7.14(m,2H),5.58(s,1H),4.60(t,J=13.5Hz,2H),4.23(s,1H),2.78(q,J=7. 6Hz,2H),2.03(s,2H),1.68(t,J=11.9Hz,2H),1.45(s,9H),1.23(t,J=7.6Hz,3H).

[0350] Tert-butyl (2-(4-(2-((1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-2-oxoethyl)benzamido)phenyl)carbamate (29c)

[0351] White solid, yield 75.5%. 1 H NMR (400MHz, DMSO) δ9.86 (s, 1H), 8.71 (s, 1H), 8.27 (d, J = 7.5Hz, 1H), 7.99–7.95 (m, 1H), 7 .91(d,J=8.0Hz,2H),7.58–7.49(m,4H),7.44(d,J=8.0Hz,2H),7.41–7.29(m,4H),7.25–7. 15(m,2H),5.55(s,1H),4.49–4.39(m,2H),3.95–3.89(m,1H),3.53(s,2H),3.38(d,J=11.2 Hz,2H),2.76(q,J=7.5Hz,2H),1.91(d,J=12.8Hz,2H),1.44(s,9H),1.21(t,J=7.5Hz,3H).

[0352] Tert-butyl (2-(4-(((1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)benzamido)phenyl)carbamate (29d)

[0353] White solid, yield 50.2%. 1 H NMR (400MHz, DMSO) δ9.83 (s, 1H), 8.71 (s, 1H), 7.93 (d, J = 6.4Hz, 3H), 7.61–7.51 (m, 6H), 7.41–7.30 (m, 4H), 7.23–7.15 (m, 2H), 5.54 (s ,1H),4.43(s,2H),3.88(s,2H),3.30(d,J=11.4Hz,2H),2.75(q,J=7.5Hz,3H),2.02–1.90(m,2H),1.45(s,11H),1.20(t,J=7.6Hz,3H).

[0354] Tert-butyl (2-(2-((1-(6-((2-amino-2-oxo-phenethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)pyrimidine-carboxamido)phenyl)carbamate (29e)

[0355] White solid, yield 46.0%. 1 H NMR (400MHz, DMSO) δ9.71 (s, 1H), 8.92 (s, 2H), 8.57 (s, 1H), 7.93 (s, 1H), 7.61 (d, J = 7. 8Hz,1H),7.54(d,J=7.1Hz,2H),7.48(dd,J=7.9,1.6Hz,1H),7.44–7.30(m,4H),7.21– 7.15(m,2H),5.57(s,1H),5.09–4.99(m,1H),4.77(d,J=13.3Hz,2H),3.30(s,2H),3.0 6(s,3H),2.78(q,J=7.5Hz,2H),1.93–1.76(m,4H),1.45(s,9H),1.23(t,J=7.6Hz,3H).

[0356] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(2-Aminophenyl)glutaramide (30a)

[0357] White solid, yield 74.2%, mp 174-176℃. 1 H NMR (400MHz, DMSO) δ9.09 (s, 1H), 7.89 (dd, J = 12.0, 4.8Hz, 2H), 7.58–7.43 (m, 3H), 7.41–7.2 8(m,4H),7.24–7.05(m,2H),6.95–6.85(m,1H),6.72(d,J=7.9,1H),6.54(t,J=7.5,1H),5.5 3(s,1H),4.46–4.41(m,2H),3.94(s,1H),3.37(d,J=12.6Hz,2H),2.77(q,J=7.3Hz,2H),2.3 4(t,J=7.4Hz,2H),2.30–2.17(m,2H),2.05–1.80(m,4H),1.45(s,2H),1.21(t,J=7.5Hz,3H). 13C NMR(101MHz,DMSO)δ171.62,171.51,171.27,169.98,164.79,164.41,158.5 0,155.11,142.37,136.64,129.13,128.66,126.18,125.82,123.98,121.63 ,116.60,116.42,116.32,115.11,93.85,87.63,53.69,46.96,45.62,35.62 ,35.32,32.07,28.47,27.57,24.13,22.03,13.99.HRMS-ESI(m / z)calc.for C 33 H 36 N8O3S[M+Na] + 647.2529,found 647.2529.HPLC t R =10.24min (99.93% purity).

[0358] N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(2-Aminophenyl)terephthalamide (30b)

[0359] White solid, yield 76.5%, mp 247-249°C. 1 H NMR (400MHz, DMSO) δ10.64(s,1H),8.66(d,J=7.6Hz,1H),8.32–7.99(m,6H),7.67–7.25(m,12H),5.63(s,1H),4.65–4.54(m,2H) ,4.26–4.18(m,1H),3.34(d,J=12.1Hz,2H),2.77(q,J=7.6Hz,2H),2.06–1.98(m,2H),1.80–1.60(m,2H),1.22(t,J=7.6Hz,3H). 13C NMR (101MHz, DMSO) δ170.05,165.45,165.38,164.91,164.40,158.53,137.68,136.79,136.41,129.08,128.71,128.58,128.52,127. 82,127.76,126.95,123.70,116.46,115.17,93.82,87.64,53.63,47.43,47.27,46.83,31.92,27.60,14.04.HRMS-ESI(m / z)calc.for C 36 H 34 N8O3S[M+Na] + 681.2372,found681.2374.HPLC t R =11.46min (99.31% purity).

[0360] 4-(2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-2-oxoethyl)-N-(2-aminophenyl)benzamide (30c)

[0361] White solid, yield 79.2%, mp 245-247℃. 1 H NMR (400MHz, DMSO) δ10.41(d,J=11.4Hz,1H),8.34(t,J=7.0Hz,1H),8.04(td,J=9.7,4.1H z,3H),7.53(d,J=7.4Hz,3H),7.44(d,J=8.0Hz,2H),7.40–7.27(m,7H),5.57(d,J=2.2Hz,1 H),4.45(t,J=10.5Hz,2H),3.92(d,J=9.9Hz,1H),3.37(dd,J=13.6,10.5Hz,2H),2.76(q, J=7.6Hz,2H),1.91(d,J=12.4Hz,2H),1.48(t,J=14.4Hz,2H),1.24–1.18(t,J=7.6Hz,3H). 13C NMR (101MHz, DMSO) δ170.00,169.46,166.02,164.80,164.40,158.48,141.21,136.70,132.32,129.39,129.11,128.67,128.48,127 .61,127.02,116.45,115.14,93.82,87.61,53.64,46.89,45.89,42.72,31.96,27.59,21.25,14.57,14.02.HRMS-ESI(m / z)calc.for C 36 H 34 N8O3S[M–H] - 671.2558,found 671.2559.HPLC t R =5.18min (98.52% purity).

[0362] 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-(2-aminophenyl)benzamide (30d)

[0363] White solid, yield 70.2%, mp 221-223℃. 1 H NMR (400MHz, DMSO) δ10.21(s,1H),9.82(s,2H),8.13(d,J=8.5Hz,3H),7.81(d,J=7 .9Hz,2H),7.56(d,J=7.4Hz,2H),7.47–7.27(m,5H),7.14(s,2H),6.98(s,1H),5.63 (s,1H),4.67(d,J=13.1Hz,2H),4.32(d,J=6.1Hz,2H),3.22(d,J=13.8Hz,2H),2.77 (q,J=7.6Hz,2H),2.33(d,J=11.7Hz,2H),1.88–1.75(m,2H),1.22(t,J=7.5Hz,3H). 13C NMR (101MHz, DMSO) δ170.06,165.43,164.93,164.40,158.60,136.66,136.10,134.89,130.46,129.17,128.71,128.64,127.54, 127.08,116.34,115.03,94.36,88.14,55.42,54.09,53.61,46.97,46.40,46.28,28.43,27.61,14.02.HRMS-ESI(m / z)calc.for C 36 H 36 N8O2S[M+Na] + 645.2755,found 645.2756.HPLC t R =7.35min (97.74% purity).

[0364] 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)(methyl)amino)-N-(2-aminophenyl)pyrimidine-5-carboxamide (30e)

[0365] White solid, yield 70.2%, mp 217-219℃. 1 H NMR(400MHz,DMSO)δ9.52(s,1H),8.93(s,2H),7.95(s,1H),7.57–7.50(m,2H ),7.40–7.31(m,4H),7.14(d,J=7.8Hz,1H),7.01–6.93(m,1H),6.77(d,J=7.9 Hz,1H),6.59(t,J=7.5Hz,1H),5.57(s,1H),4.95(s,2H),4.76(d,J=13.3Hz,2 H),3.05(s,3H),2.83–2.75(m,2H),1.91–1.77(m,4H),1.22(t,J=7.6Hz,3H). 13C NMR (101MHz, DMSO) δ170.01,164.79,164.43,163.23,162.03,158.42,143.80,136.60,129.16,128.64,127.38,127.06,123.26,116. 65,116.57,116.51,116.37,115.16,93.89,87.70,53.71,52.45,47.63,29.58,29.09,29.04,27.60,14.01.HRMS-ESI(m / z)calc.for C 34 H 34 N 10 O2S[MH] - 645.2489,found 645.2494.HPLC t R =5.21min (96.94% purity).

[0366] Example 11: Preparation of target compounds 40a-d, 44a-c, 48

[0367] Methyl 4-(N-methylmethanesulfonamido)benzoate (32)

[0368] Methanesulfonyl chloride (1.04 g, 9.08 mmol) was slowly added to CH2Cl2 (10 mL) containing 31 (1 g, 6.05 mmol) and triethylamine (1.22 g, 12.10 mmol) at 0°C and reacted at room temperature for 1.5 hours. The reaction solution was diluted with 50 mL CH2Cl2, and the organic phase was washed with 1 M aqueous hydrochloric acid solution (100 mL), saturated aqueous sodium bicarbonate solution (100 mL), and saturated aqueous sodium chloride solution (100 mL) in sequence. The organic phase was then dried over anhydrous Na2SO4 and concentrated. 32 was purified by silica gel column chromatography as a pale yellow solid with a yield of 83.5%. 1 H NMR (400MHz, DMSO) δ8.02–7.94(m,2H),7.59–7.51(m,2H),3.86(s,3H),3.30(s,3H),3.01(s,3H).

[0369] N-(4-(Hydroxymethyl)phenyl)-N-methylmethanesulfonamide (33)

[0370] DIBAL (8.22 mmol) was added to a THF solution of 32 (1 g, 4.11 mmol), and the mixture was stirred at -20°C for 6 hours. After the reaction was completed, 30 mL of 10% NaOH solution was used to quench the reaction. The mixture was then extracted with ethyl acetate (10 mL × 3), and the organic phase was concentrated to obtain a crude product, which was purified by column chromatography to obtain 33 as a gray solid with a yield of 68.9%. 1 H NMR (400MHz, DMSO) δ7.40–7.30(m,4H),5.22(t,J=5.7Hz,1H),4.49(d,J=5.7Hz,2H),3.22(s,3H),2.92(s,3H).

[0371] 4-(N-Methylmethylsulfonamido)benzyl-4-methylbenzenesulfonate (34)

[0372] Pale yellow solid, yield 45.8%. MS-ESI (m / z) calc.for C 16 H 19 N1O5S2[M+H] + 370.1.

[0373] N-(4-(((6-amino-3,5-dicyano-4-ethylpiperidin-2-yl)thio)methyl)phenyl)-N-methylmethanesulfonamide (35)

[0374] Brown-yellow solid, yield 71.6%. 1 H NMR (400MHz, DMSO) δ7.55–7.50(m,2H),7.35–7.30(m,2H),4.46(s,2H),3.21(s,3H),2.93(s,3H),2.69(q,J=7.8Hz,2H),1.18(t,J=7.6Hz,3H).

[0375] N-(4-(((6-Chloro-3,5-dicyano-4-ethylpiperidin-2-yl)thio)methyl)phenyl)-N-methylmethanesulfonamide (36)

[0376] Brown-yellow solid, yield 31.3%. 1 H NMR (400MHz, DMSO) δ7.51(d,J=7.2Hz,2H),7.42–7.33(m,2H),4.58(s,2H),3.21(s,3H),2.94(s,3H),2.88(q,J=8.2Hz,2H),1.25(t,J=8.6Hz,3H).

[0377] Tert-butyl (2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)ethyl)carbamate (37a)

[0378] Yellow solid, yield 79.7%. 1 H NMR (400MHz, DMSO) δ7.44(d,J=8.4Hz,2H),7.41–7.34(m,2H),6.94(t,J=6.0Hz,1H),4.52(s,2H),3.83(t,J=6.1H z,2H),3.39(s,3H),3.20(d,J=8.2Hz,5H),2.92(s,3H),2.77(q,J=7.6Hz,2H),1.31(s,9H),1.21(t,J=7.6Hz,3H).

[0379] Tert-butyl (3-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)propyl)carbamate (37b)

[0380] Yellow solid, yield 77.2%. 1 H NMR (400MHz, DMSO) δ7.45(d,J=8.2Hz,2H),7.37(dd,J=8.3,1.5Hz,2H),6.85(d,J=5.6Hz,1H),4.52(s,2H),3.73(t,J=7.4Hz ,2H),3.21(s,3H),2.94(dd,J=8.3,3.5Hz,5H),2.77(q,J=7.6Hz,2H),1.79–1.71(m,2H),1.33(s,9H),1.21(t,J=7.6Hz,3H).

[0381] tert-Butyl (2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)ethyl)(methyl)carbamate (37c)

[0382] Yellow solid, yield 74.0%. 1 H NMR (400MHz, DMSO) δ7.43(d,J=8.3Hz,1H),7.37(d,J=8.3Hz,1H),4.53(s,1H),3.91(d,J=25. 3Hz,1H),3.37(d,J=7.7Hz,3H),2.91(s,2H),1.38(s,2H),1.31(s,2H),1.22(d,J=6.8Hz,5H).

[0383] 2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)ethyl-1-ammonium salt (38a)

[0384] Pale yellow solid, yield 90.1%. 1 H NMR (400MHz, DMSO) δ8.01(s,3H),7.46(d,J=8.3Hz,2H),7.38(d,J=8.5Hz,2H),4.54(s,2H),3.99(t,J=6.4Hz,2 H), 3.45 (s, 3H), 3.22 (s, 3H), 3.10 (d, J = 7.1Hz, 2H), 2.94 (s, 3H), 2.80 (q, J = 7.6Hz, 3H), 1.23 (t, J = 7.5Hz, 3H).

[0385] 3-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)propyl-1-ammonium salt (38b)

[0386] Pale yellow solid, yield 84.1%. 1 H NMR (400MHz, DMSO) δ7.95 (s, 3H), 7.46 (d, J = 8.4Hz, 2H), 7.39 (d, J = 8.3Hz, 2H), 4.54 (s, 2H), 3.99 (t, J = 6.4Hz, 2 H),3.45(s,3H),3.23(s,3H),3.11(t,J=6.5Hz,2H),2.94(s,3H),2.80(q,J=7.6Hz,2H),1.23(t,J=7.6Hz,3H).

[0387] 2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)propyl-1-ammonium salt (38c)

[0388] Pale yellow solid, yield 85.0%. 1 H NMR (400MHz, DMSO) δ8.82(s,2H),7.50–7.43(m,2H),7.43–7.35(m,2H),4.56(s,2H),4.04(t,J=6.3Hz,2H),3.46(s,4H ),3.22(s,3H),3.17(q,J=6.8Hz,3H),2.94(s,3H),2.80(q,J=7.6Hz,2H),2.54(d,J=5.3Hz,3H),1.23(t,J=7.5Hz,3H).

[0389] N 1 -2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N 4 -((Tetrahydro-2H-pyran-2-yl)oxy)succinamide (39a)

[0390] Pale yellow solid, yield 31.8%. 1 H NMR (400MHz, DMSO) δ10.89(s,1H),7.94(t,J=5.9Hz,1H),7.45(d,J=8.5Hz,2H),7.37(d,J=8. 4Hz,2H),4.79(s,1H),4.51(s,2H),3.89(d,J=11.1Hz,1H),3.82(t,J=6.2Hz,2H),3.48(d,J= 11.6Hz,1H),3.38(s,3H),3.29(d,J=6.1Hz,1H),3.21(s,3H),2.92(s,3H),2.78(q,J=7.5Hz, 2H), 1.98 (dt, J = 20.0, 7.5Hz, 4H), 1.65 (p, J = 7.8Hz, 5H), 1.50 (s, 3H), 1.22 (t, J = 7.6Hz, 3H).

[0391] N 1 -2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N 5 -((Tetrahydro-2H-pyran-2-yl)oxy)glutaramide (39b)

[0392] Pale yellow solid, yield 44.2%. 1 H NMR (400MHz, DMSO) δ10.86 (s, 1H), 7.96 (t, J = 6.1Hz, 1H), 7.44 (d, J = 8.4Hz, 2H) ,7.37(d,J=8.3Hz,2H),4.77(s,1H),4.49(s,2H),3.81–3.74(m,2H),3.40(s,3H ),3.29(d,J=6.1Hz,3H),3.20(s,3H),2.91(s,3H),2.76(q,J=7.5Hz,2H),,1.98 (dt,J=20.0,7.5Hz,4H),1.71–1.64(m,5H),1.50(s,3H),1.22(t,J=7.6Hz,3H).

[0393] N1 -2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)propyl)-N 4 -((Tetrahydro-2H-pyran-2-yl)oxy)succinamide (39c)

[0394] Pale yellow solid, yield 39.5%. 1 H NMR (400MHz, DMSO) δ10.95(s,1H),7.89(s,1H),7.48–7.41(m,2H),7.41–7.34(m,2H),4. 78(s,1H),4.52(s,2H),3.90(s,1H),3.73(s,2H),3.48(s,2H),3.22(s,3H),3.05(d,J=6 .1Hz,2H),2.93(d,J=1.9Hz,3H),2.77(d,J=8.0Hz,2H),2.42(d,J=4.7Hz,2H),2.26(s,2 H),2.21(d,J=6.8Hz,2H),1.75(s,2H),1.62(s,3H),1.49(s,3H),1.21(t,J=7.7Hz,3H).

[0395] 2-((2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)(methyl)amino)ethyl)(methyl)amino)-N-((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (39d)

[0396] Pale yellow solid, yield 39.2%. 1 H NMR(400MHz,DMSO)δ11.49(s,1H),8.62(s,2H),7.44–7.32(m,4H),5.75(s, 1H),4.93(s,1H),4.47(s,2H),4.08(s,2H),4.02(s,1H),3.91(t,J=5.8Hz,2 H),3.52(d,J=11.4Hz,1H),3.35(s,3H),3.21(s,3H),3.08(s,3H),2.92(s,3 H),2.72(d,J=7.6Hz,10H),1.71(s,3H),1.54(s,3H),1.17(t,J=7.6Hz,3H).

[0397] N 1 -(2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N4 -Hydroxysuccinamide (40a)

[0398] Pale yellow solid, yield 21.2%, mp 152-154°C. 1 H NMR (400MHz, DMSO) δ10.39(s,1H),8.68(d,J=1.6Hz,1H),8.04(t,J=5.9Hz,1H), 7.45(d,J=8.2Hz,2H),7.38(d,J=8.3Hz,2H),4.52(s,2H),3.81(t,J=6.2Hz,2H) ,3.39(s,3H),3.30(d,J=6.1Hz,3H),3.22(s,3H),2.93(s,3H),2.78(q,J=7.5Hz ,2H),2.25(dd,J=8.8,6.6Hz,2H),2.15(t,J=7.4Hz,2H),1.22(t,J=7.5Hz,3H). 13 C NMR (101MHz, DMSO) δ171.93,168.79,164.72,164.54,157.63,141.30,135.82,129.88,126.83,116.99,115 .36,93.51,86.00,51.69,38.18,36.93,35.47,33.45,31.06,28.17,27.62,14.00.HRMS-ESI(m / z)calc.for C 34 H 34 N 10 O2S[M+Na] + 596.1726,found 596.1728.HPLC t R =3.98min (96.78% purity).

[0399] N 1 -(2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N 5 -Hydroxyglutaramide (40b)

[0400] Pale yellow solid, yield 30%, mp 117-119°C. 1H NMR (400MHz, DMSO) δ10.37(s,1H),8.68(s,1H),7.90(t,J=5.6Hz,1H),7.44(d,J=8.2 Hz,2H),7.37(d,J=8.1Hz,2H),4.52(s,2H),4.03(td,J=8.6,5.8Hz,1H),3.73(t,J=7 .5Hz,2H),3.22(s,3H),3.05(q,J=6.4Hz,3H),2.93(s,3H),2.77(q,J=7.4Hz,2H),2. 30–2.24(m,2H),2.16(t,J=7.4Hz,2H),1.76(t,J=7.5Hz,2H),1.22(t,J=7.9Hz,3H). 13 C NMR (101MHz, DMSO) δ171.56,168.89,164.80,164.59,157.44,141.32,135.81,129.87,126.82,116.93,115.38, 93.41,85.92,50.28,38.18,36.50,35.49,33.43,31.04,28.25,27.64,14.56,13.99.HRMS-ESI(m / z)calc.forC 26 H 33 N7O5S2[M+Na] + 610.1836,found586.1831.HPLC t R =4.09min (95.64% purity).

[0401] N 1 -(3-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)propyl)-N 4 -Hydroxysuccinamide (40c)

[0402] Pale yellow solid, yield 39.7%, mp 110-112°C. 1H NMR (400MHz, DMSO) δ10.41(s,1H),8.70(s,1H),8.04(t,J=5.9Hz,1H),7.45(d,J= 8.2Hz,2H),7.38(d,J=8.3Hz,2H),4.52(s,2H),3.82(t,J=6.2Hz,2H),3.39(s,3H) ,3.31(q,J=6.1Hz,3H),3.22(s,3H),2.93(s,3H),2.78(q,J=7.5Hz,2H),2.01(t, J=7.6Hz,2H),1.92(t,J=7.4Hz,2H),1.66(p,J=7.5Hz,2H),1.22(t,J=7.5Hz,3H). 13 C NMR (101MHz, DMSO) δ172.49,169.17,164.75,164.53,157.66,141.31,135.80,129.90,126.81,116.97,115.34, 93.52,86.00,51.71,38.19,36.85,35.53,35.19,33.50,32.22,27.61,21.81,13.98.HRMS-ESI(m / z)calc.forC 26 H 33 N7O5S2[MH] - 586.1887,found586.1883.HPLC t R =7.85min (98.24% purity).

[0403] 2-((2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)(methyl)amino)-N-hydroxypyrimidine-5-carboxamide (40d)

[0404] Orange-yellow solid, yield 27%, mp 155-157°C. 1 H NMR (400MHz, DMSO) δ11.05(s,1H),9.00(s,1H),8.61(s,2H),7.42(d,J=8.2Hz,2H),7.36(d,J=8.2Hz,2H),4.48(s,2H),4.12–4.0 5(m,2H),3.91(t,J=5.9Hz,2H),3.35(s,3H),3.32(s,3H),3.07(s,3H),2.93(s,3H),2.72(q,J=7.7Hz,2H),1.17(t,J=7.6Hz,3H). 13C NMR (101MHz, DMSO) δ172.49,169.17,164.75,164.53,157.66,141.31,135.80,129.90,126.81,116.97,115.34 ,93.52,86.00,51.71,38.19,36.85,35.53,35.19,33.50,32.22,27.61,21.81,13.98.HRMS-ESI(m / z)calc.for C 27 H 31 N9O4S2[MH] - 608.1845,found 608.1640.HPLC t R =9.90min (98.58% purity).

[0405] Tert-butyl (1-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)piperidin-4-yl)carbamate (41)

[0406] White solid, yield 73.5%. 1 H NMR (400MHz, DMSO) δ7.44(d,J=8.3Hz,2H),7.37(d,J=8.3Hz,2H),6.90(d,J=7.8Hz,1H),4.50(s,2H),4.39(d,J=13.5Hz,2H),3.60(s,1H),3.2 7(s,1H),3.21(s,3H),2.92(s,3H),2.76(q,J=7.9Hz,2H),1.85(d,J=12.7Hz,2H),1.44(d,J=12.6Hz,2H),1.39(s,9H),1.21(t,J=7.6Hz,3H).

[0407] 1-((3,5-Dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)piperidin-4-ammonium salt (42)

[0408] White solid, yield 80.8%. 1H NMR (400MHz, DMSO) δ8.12(s,3H),7.45(d,J=8.4Hz,2H),7.38(d,J=8.4Hz,2H),4.53(d,J=6.8Hz,4H),3.24(d,J=1 7.7Hz,5H),2.94(s,3H),2.78(q,J=7.6Hz,2H),2.06(d,J=12.6Hz,2H),1.67–1.54(m,2H),1.22(t,J=7.6Hz,3H).

[0409] N 1 -(1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)piperidin-4-yl)-N 5 -((Tetrahydro-2H-pyran-2-yl)oxy)glutaramide (43a)

[0410] White solid, yield 37%. 1 H NMR (400MHz, DMSO) δ10.91(s,1H),7.81(d,J=7.6Hz,1H),7.44(d,J=8.3Hz,2H),7.37(d,J=8.4Hz,2H),4.8 0(s,1H),4.50(s,2H),4.37(d,J=13.4Hz,2H),3.91(d,J=9.6Hz,2H),3.49(d,J=11.6Hz,1H),3.37(d,J=12 .4Hz,2H),3.21(s,3H),3.17(d,J=5.2Hz,1H),2.93(s,3H),2.77(q,J=7.6Hz,2H),2.06(t,J=7.5Hz,2H),1 .99(t,J=7.4Hz,2H),1.87(d,J=12.8Hz,2H),1.77–1.60(m,5H),1.54–1.37(m,5H),1.22(t,J=7.6Hz,3H).

[0411] (E)-3-(4-(((1-(3,5-dicyano-4-ethyl-6-((N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-((tetrahydro-2H-pyran-2-yl)oxy)acrylamide (43b)

[0412] White solid, yield 45%. 1H NMR (400MHz, DMSO) δ11.21(s,1H),7.52(t,J=7.6Hz,3H),7.44(d,J=8.6Hz,2H),7.41(d,J =7.9Hz,2H),7.37(d,J=8.4Hz,2H),6.48(d,J=15.8Hz,1H),4.91(s,1H),4.51(s,2H),4.36 (d,J=13.3Hz,2H),3.97(s,1H),3.77(s,2H),3.21(s,3H),2.92(s,3H),2.77(d,J=7.5Hz, 2H),1.95(s,2H),1.70(s,3H),1.55(s,3H),1.37(d,J=10.5Hz,2H),1.22(t,J=7.7Hz,3H).

[0413] 2-((1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)piperidin-4-yl)amino)-N-((tetrahydro-2H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (43c)

[0414] White solid, yield 40.9%. 1 H NMR (400MHz, DMSO) δ11.51(s,1H),8.64(s,2H),7.91(d,J=7.9Hz,1H),7.45(d,J=8.5Hz,2H),7.41–7.33(m,2H),4.95(s,1H),4.48(d,J=22.6Hz,4H ),4.18(s,1H),3.20(s,3H),2.92(s,3H),2.78(q,J=7.6Hz,2H),1.99(d,J =12.3Hz,2H),1.71(s,3H),1.56(d,J=12.7Hz,5H),1.22(t,J=7.6Hz,3H).

[0415] N 1 -(1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (44a)

[0416] White solid, yield 23.5%, mp 149-151°C. 1H NMR (400MHz, DMSO) δ10.41(s,1H),8.73–8.66(m,1H),7.91(d,J=7.7Hz,1H),7.44(d,J=8.2Hz,2H ),7.37(d,J=8.3Hz,2H),4.50(s,2H),4.38(d,J=13.4Hz,2H),3.91(s,1H),3.21(s,3H),3.17(d,J =5.2Hz,1H),2.93(s,3H),2.77(q,J=7.6Hz,2H),2.06(t,J=7.5Hz,2H),1.96(dt,J=14.7,7.4Hz, 3H), 1.86 (d, J = 12.7Hz, 2H), 1.70 (p, J = 7.7Hz, 2H), 1.44 (q, J = 11.4Hz, 2H), 1.21 (t, J = 7.6Hz, 3H). 13 C NMR (101MHz, DMSO) δ171.42,169.22,165.05,164.49,158.59,141.25,136.09,129.79,126.83,116.44,115.18,94.38,87. 83,60.23,46.93,45.50,38.18,35.46,35.23,33.53,33.24,32.18,31.92,31.72,27.60,21.92,21.03,14.59,14.03.HPLC t R =9.90min (98.58% purity).

[0417] (E)-(1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (44b)

[0418] White solid, yield 27.2%, mp 164-166°C. 1H NMR (400MHz, DMSO) δ10.81 (s, 1H), 9.07 (s, 1H), 7.55 (d, J = 7.8Hz, 2H), 7.50–7. 41(m,5H),7.38(d,J=8.1Hz,2H),6.48(d,J=15.7Hz,1H),4.51(s,2H),4.43(d,J =13.2Hz,2H),3.90(s,1H),3.29(d,J=11.9Hz,3H),3.21(s,3H),2.92(s,3H),2. 77(q,J=7.5Hz,2H),2.04(d,J=12.1Hz,2H),1.49(s,2H),1.21(t,J=7.6Hz,3H). 13 C NMR(101MHz,DMSO)δ165.04,164.52,163.17,158.44,141.25,138.45,136.14,129.81,127.91,126.83,1 16.47,115.19,94.33,87.74,53.23,46.47,38.20,35.48,33.49,27.60,14.02.HRMS-ESI(m / z)calc.for C 33 H 37 N7O4S2[M+H] + 660.2368,found 660.2371.HPLC t R =5.47min (96.01% purity).

[0419] (E)-3-(4-(((1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-hydroxyacrylamide (44c)

[0420] White solid, yield 29%, mp 137-139°C. 1H NMR (400MHz, DMSO) δ11.07(s,1H),9.00(s,1H),8.63(s,2H),7.82(d,J=7.8Hz ,1H),7.45(d,J=8.1Hz,2H),7.37(d,J=8.1Hz,2H),4.51(s,2H),4.46(s,2H), 4.17(s,1H),3.44(d,J=7.1Hz,2H),3.20(s,3H),2.92(s,3H),2.78(q,J=7.7H z, 2H), 1.99 (d, J = 12.3Hz, 2H), 1.57 (d, J = 11.7Hz, 2H), 1.22 (t, J = 7.5Hz, 3H). 13 C NMR (101MHz, DMSO) δ165.07,164.55,162.54,159.83,158.50,141.22,136.12,129.77,126.85,116.48,115.20,94 .37,87.76,64.00,55.39,47.58,46.98,38.17,35.42,33.51,31.71,27.61,14.69,14.03.HRMS-ESI(m / z)calc.for C 28 H 31 N9O4S2[MH] - 620.1840,found 620.1838.HPLC t R =2.27min (99.37% purity).

[0421] Tert-butyl 4-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)piperazine-1-carbamate (45)

[0422] White solid, yield 81.1%. 1 H NMR (400MHz, DMSO) δ7.46(d,J=8.2Hz,2H),7.38(d,J=8.2Hz,2H),4.53(s,2H),3.86(t,J=5.2Hz,4H ),3.46(s,3H),3.22(s,3H),2.93(s,3H),2.77(q,J=7.5Hz,2H),1.42(s,9H),1.22(t,J=7.5Hz,3H).

[0423] 4-(3,5-Dicyano-4-ethyl-6-((4-(N-methylmethanesulfonamido)benzyl)thio)pyridin-2-yl)piperazine-1-ammonium salt (46)

[0424] White solid, yield 86.1%. 1 H NMR (400MHz, DMSO) δ9.42(s,2H),7.46(d,J=8.0Hz,2H),7.38(d,J=8.2Hz,2H),4.54(s,2H) ,4.05(q,J=6.5Hz,4H),3.23(s,7H),2.94(s,3H),2.84–2.75(m,2H),1.24(d,J=7.2Hz,3H).

[0425] 2-(4-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)benzyl)thio)pyridin-2-yl)piperazin-1-yl)-N-((tetrahydro-2-H-pyran-2-yl)oxy)pyrimidine-5-carboxamide (47)

[0426] White solid, yield 49.2%. 1 H NMR (400MHz, DMSO) δ12.05(s,1H),11.56(s,1H),9.05(s,2H),7.47(d,J=8 .3Hz,2H),7.38(d,J=8.4Hz,2H),4.96(s,1H),4.55(s,2H),4.02–3.99(m,4 H),3.96(dd,J=7.1,3.5Hz,4H),3.55(s,2H),3.21(s,3H),2.92(s,3H),2. 80(q,J=7.8Hz,2H),1.74–1.71(m,3H),1.55(s,3H),1.23(t,J=7.6Hz,3H).

[0427] 2-(4-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)benzyl)thio)pyridin-2-yl)piperazin-1-yl)-N-hydroxypyrimidine-5-carboxamide (48)

[0428] White solid, yield 33%, mp 170-171°C. 1 H NMR (400MHz, DMSO) δ8.73(s,2H),7.47(d,J=8.4Hz,2H),7.38(d,J=8.5Hz,2H),4.55(s,2H),4.00(dd,J=7.2,3 .5Hz,4H),3.95(dd,J=7.1,3.6Hz,4H),3.21(s,3H),2.92(s,3H),2.80(q,J=7.6Hz,2H),1.23(t,J=7.6Hz,3H). 13CNMR(101MHz,DMSO)δ165.14,164.47,161.72,158.70,157.58,141.27,136.09,129.79,126.81,116.41, 115.57,115.12,94.78,88.00,47.19,43.34,38.19,35.54,33.62,27.58,14.01.HRMS-ESI(m / z)calc.for C 27 H 29 N9O4S2[MH] - 606.1688, found 606.1683.HPLCt R =9.16min (99.65% purity).

[0429] Experimental Example 1: Inhibitory activity of target compounds on HDACs

[0430] 1. Materials:

[0431] HDACs enzyme (HeLa cell nuclear extract); Boc-Lys(Ac)-AMC substrate; trypsin; EDTA; LBH-589 (10 mM, dissolved in dimethyl sulfoxide); 96-well all-black ELISA plate; glycerol; ultrapure water

[0432] 2. Methods:

[0433] Buffer preparation:

[0434] The formula is 15 mM Tris-HCl (pH=8.0), 250 μM EDTA, 250 mM NaCl, and 10% glycerol.

[0435] Preparation of Trypsin solution:

[0436] The formula is 10 mg / mL trypsin, Buffer, 2 μM LBH589.

[0437] Preparation of substrate solution:

[0438] Dissolve the substrate in DMSO to prepare a 30 mM stock solution, then dilute to 300 μM with buffer.

[0439] The content of DMSO is about 1%.

[0440] Dilution of enzyme solution:

[0441] Dilute the enzyme solution with buffer at a ratio of 1:80.

[0442] Preparation of compound solution:

[0443] Dilute the compounds (test compounds and positive control drug SAHA) to 5× final concentration with buffer

[0444] Preparation and determination of 100% and blank:

[0445] 50μL buffer was mixed with 10μL enzyme solution. After 5 minutes, 40μL substrate was added and reacted at 37℃ for 0.5 hour. Then 100μL Trypsin solution was added to terminate the reaction. After reacting at 37℃ for 20 minutes, the fluorescence intensity was measured at (390nm / 460nm) to obtain 100% absorption. After 40μL substrate was added to 60μL buffer and reacted at 37℃ for 0.5 hour, 100μL Trypsin solution was added and reacted at 37℃ for 20 minutes to obtain blank absorption.

[0446] Determination of the inhibition of HDACs activity by compounds:

[0447] 50 μL of drug-containing buffer was mixed with 10 μL of enzyme solution and pre-incubated for 5 minutes. After adding 40 μL of substrate, the reaction was allowed to proceed at 37°C for 0.5 hours. Then, 100 μL of Trypsin solution was added to terminate the reaction. After reacting at 37°C for 20 minutes, the fluorescence intensity was measured at (390 nm / 460 nm). The inhibition rate and IC were calculated using GraphPad Prism software and the formula 50 Value. (Table 1)

[0448] Table 1. HDACs inhibitory activity of target compounds

[0449]

[0450] a A,IC 50 Greater than 5 μM; B, IC 50 Between 1μM and 5μM; C, IC 50 Between 0.2μM and 1μM; D, IC 50 Less than 0.2μM

[0451] b Not tested

[0452] Experimental Example 2: Inhibitory activity of target compounds on DNMT1

[0453] 1. Materials:

[0454] MT1hypo-SFRP1-Luc cells; 96-well all-black ELISA plate; RPMI1640 medium; 10% fetal bovine serum (Hyclone, USA); PBS buffer; Luciferase assay reagent, positive control drugs SAHA and GSK5032.

[0455] 2. Methods:

[0456] Inhibition of DNMT1 can restore the expression of the hypermethylated SFRP1 gene. Because the luciferase gene is inserted into SFRP1, its protein expression content can be determined by detecting the fluorescence intensity. MT1hypo-SFRP1-Luc cells (5400 per well) were seeded in a black well plate and cultured in a constant temperature incubator (37°C, 5% carbon dioxide) for 16 hours. Then, different concentrations of compound dilutions prepared from the corresponding cell culture medium were added, and after addition, they were continued to be cultured in a constant temperature incubator for 72 hours. Finally, luciferase detection reagent was added to each well and the corresponding fluorescence intensity was detected with an enzyme reader. The inhibitory activity was expressed as fold change (fluorescence intensity of the compound group / fluorescence intensity of the blank control group) (Table 2).

[0457] Table 2. DNMT1 inhibitory activity of target compounds

[0458]

[0459] a A, fold change is less than 100; B, fold change is between 100 and 200; C, fold change is between 200 and 300; D, fold change is greater than 300

[0460] Experimental Example 3: Antiproliferative Activity Experiment of Target Compound

[0461] 1. Materials:

[0462] MV4-11 (human acute myeloid leukemia cells), KG-1 (human acute myeloid leukemia cells), THP-1 (human monocytic leukemia cells), Jurkat (human T lymphocyte leukemia cells), K562 (human chronic myeloid leukemia cells), MC38 (mouse colon cancer cells), 10% fetal bovine serum (Hyclone, USA), 2.5 g·L -1 Trypsin (Gibco, USA), CCK-8, modified RPMI1640 medium, DMEM medium (Hyclone, USA), positive control drugs SAHA and GSK5032, 96-well plates.

[0463] 2. Methods:

[0464] Culture cells routinely and collect logarithmically growing cells for experiments; dilute logarithmically growing solid tumor cells to 4×103 pc·mL -1 (Hematologic tumor cells were diluted to 1×10 4 pc·mL -1 ) were inoculated into a 96-well plate (100 μL was added to each well), and the wells without cells were used as blank wells, and then placed in a constant temperature incubator (37°C, 5% carbon dioxide) for 8 hours; the target compound solution and positive drug solution prepared with culture medium were added, and the wells without drugs were used as 100% wells. After culturing in a constant temperature incubator (37°C, 5% carbon dioxide) for 72 hours, 20 μL CCK-8 was added, and the absorbance value of each well was measured at a wavelength of 450 nm using an enzyme reader after three hours, and the inhibition rate and IC 50 Values ​​(Table 3).

[0465] Table 3. Antiproliferative activity of compounds

[0466]

[0467] a A,IC 50 Greater than 20 μM; B, IC 50 Between 4μM and 20μM; C, IC 50 Between 0.4μM and 4μM; D, IC 50 Less than 0.4μM

[0468] Experimental Example 4: Therapeutic effect of the compound on human acute myeloid leukemia

[0469] 1. Materials:

[0470] Cell line: Human acute myeloid leukemia cells (MV4-11).

[0471] Experimental animals: Six-week-old BALB / c female nude mice were purchased from Beijing Weitonglihua Laboratory Animal Technology Co., Ltd.

[0472] 2. Methods:

[0473] About 2×10 7 MV4-11 cells were resuspended in 50 μL serum-free high-glucose medium and 50 μL Matrigel and implanted subcutaneously in the right axillary region of BALB / c nude mice. 3 At 14:00, oral administration was started for 28 consecutive days. The treatment group dose was 30 mg / kg / d, the combined group dose was 30 mg / kg / d + 30 mg / kg / d, and the control group was given an equal amount of normal saline. The tumor size and mouse body weight were measured every day during the treatment period. Tumor volume calculation formula: V (mm 3 )=Length(mm)×Width 2(mm) / 2; Tumor growth inhibition value (TGI) calculation formula: TGI=(1-average tumor volume of treatment group / average tumor volume of control group)×100%.

[0474] 3. Experimental results:

[0475] Test results see Figure 1 The tumor inhibition rate (TGI) of the target compound (R)-23a was as high as 98%, and its tumor inhibition effect was significantly better than that of the combined treatment group of the positive control drug GSK5032 and SAHA.

[0476] Experimental Example 5: Therapeutic effect of compounds on mouse colon cancer

[0477] 1. Materials:

[0478] Cell line: Mouse colon carcinoma cell line (MC38).

[0479] Experimental animals: Six-week-old BALB / c female nude mice were purchased from Beijing Weitonglihua Laboratory Animal Technology Co., Ltd.

[0480] 2. Methods:

[0481] About 1.2×10 6 MC38 cells were resuspended in 100 μL serum-free high-glucose medium and implanted subcutaneously in the right axillary region of BALB / c nude mice. 3 At 14:00, oral administration was started for 14 consecutive days. The treatment group dose was 30 mg / kg / d, the combined group dose was 30 mg / kg / d + 30 mg / kg / d, and the control group was given an equal amount of saline. The tumor size and mouse body weight were measured every day during the treatment period. Tumor volume calculation formula: V (mm 3 )=Length(mm)×Width 2 (mm) / 2; Tumor growth inhibition value (TGI) calculation formula: TGI=(1-average tumor volume of treatment group / average tumor volume of control group)×100%.

[0482] 3. Experimental results:

[0483] Test results see Figure 2 The tumor inhibition rate (TGI) of the target compound (R)-23a in the mouse colon cancer model was 82%, and its tumor inhibition effect was better than that of the positive control drug GSK5032 and its combined treatment group with SAHA.

Claims

1. A DNMT1-HDAC dual-target inhibitor, wherein the structural formula of the inhibitor is as shown in formula (I) or a pharmaceutically acceptable salt of formula (I): in, X is an aromatic heterocycle or one of the following linking groups: A is one of the following structures: R1 is hydroxyl or 2-aminophenyl; R2 is one of the following structures R3 is a hydrogen atom and is one of the following structures:

2. The DNMT1-HDAC dual-target inhibitor according to claim 1, characterized in that The inhibitor is selected from one of the following compounds: N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -Hydroxysuccinamide (11a); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (11b); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 - Hydroxyadipoic acid amide (11c); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 7 -Hydroxypimelamide (11d); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 8 -Hydroxyoctanamide (11e); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 9 -Hydroxyazelazone (11f); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 - Hydroxy-p-toluene diamide (11 g); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-4-((4-(hydroxycarbamoyl)benzyl)oxy)benzamide (11h); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(Hydroxycarbamoyl)phenyl)succinamide (11i); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(Hydroxycarbamoyl)phenyl)glutaramide (11j); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 6 -(4-(Hydroxycarbamoyl)phenyl)adipamide (11k); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(Hydroxycarbamoyl)phenyl)succinamide (111); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(4-(Hydroxycarbamoyl)phenyl)glutaramide (11m); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(4-(3-(Hydroxyamino)-3-oxopropyl-1-en-1-yl)phenyl)succinamide (11n); 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-hydroxybenzamide (11o); (E)-3-4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-hydroxyacrylamide (11p); 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(hydroxycarbamoyl)phenyl)carbamoyl)phenyl)piperidinium 4-ammonium salt (11q); (E)-1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(3-hydroxyamino)-3-oxopropyl-1-en-1-yl)benzyl)carbamoyl)benzyl)piperidin-4-ammonium salt (11r); (E)-1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((4-(3-hydroxyamino)-3-oxopropyl-1-en-1-yl)benzyl)oxy)benzyl)piperidin-4-ammonium salt (11s); 1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)-N-(4-((7-(hydroxyamino)-7-oxoheptyl)oxy)benzyl)piperidin-4-ammonium salt (11t); 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-N-hydroxypyrimidine-5-carboxamide (11u); 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)(methyl)amino)-N-hydroxypyrimidine-5-carboxamide (11v); 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)amino)piperidin-1-yl)-N-hydroxypyrimidine-5-carboxamide (15a) 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)(methyl)(methyl)amino)piperidin-1-yl)-N-hydroxypyrimidine-5-carboxamide (15b); (E)-3-(4-((4-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)methyl)phenyl)-N-hydroxyacrylamide (19a); 2-(4-((6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)-N-hydroxypyrimidine-5-carboxamide (19b); (E)-3-(4-((((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-hydroxyacrylamide (23a); (R,E)-3-(4-((((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-hydroxyacrylamide ((R)-23a); (S,E)-3-(4-((((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)methyl)phenyl)-N-hydroxyacrylamide ((S)-23a); 2-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)methyl)amino)-N-hydroxypyrimidine-5-carboxamide (23b); (E)-3-(4-((4-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperazin-1-yl)methyl)phenyl)-N-hydroxyacrylamide (27); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 5 -(2-aminophenyl)glutaramide (30a); N 1 -(1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)-N 4 -(2-aminophenyl)terephthalamide (30b); 4-(2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)-2-oxoethyl)-N-(2-aminophenyl)benzamide (30c); 4-(((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)amino)methyl)-N-(2-aminophenyl)benzamide (30d); 2-((1-(6-((2-amino-2-oxo-1-phenylethyl)thio)-3,5-dicyano-4-ethylpyridin-2-yl)piperidin-4-yl)(methyl)amino)-N-(2-aminophenyl)pyrimidine-5-carboxamide (30e); N 1 -(2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N 4 -Hydroxysuccinamide (40a); N 1 -(2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)-N 5 -Hydroxyglutaramide (40b); N 1 -(3-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)propyl)-N 4 -Hydroxysuccinamide (40c); 2-((2-((3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)(methyl)amino)ethyl)(methyl)amino)-N-hydroxypyrimidine-5-carboxamide (40d); N 1 -(1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (44a); (E)-(1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)-N 5 -Hydroxyglutaramide (44b); (E)-3-(4-(((1-(3,5-dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperidin-4-yl)amino)methyl)phenyl)-N-hydroxyacrylamide (44c); 2-(4-(3,5-Dicyano-4-ethyl-6-((4-(N-methylmethylsulfonylamino)phenyl)thio)pyridin-2-yl)piperazin-1-yl)-N-hydroxypyrimidine-5-carboxamide (48).

3. The method for preparing the DNMT1-HDAC dual-target inhibitor according to claim 2, wherein the method comprises any one of the following methods: The preparation methods of compounds 11a-11V are as follows: The preparation methods of compounds 15a, 15b, 19a and 19b are as follows: The preparation methods of compounds 23a, (R)-23a, (S)-23a, 23b, 27, and 30a-e are as follows: The preparation method of compound 40a-d is as follows: The preparation methods of compounds 44a-c and 48 are as follows:

4. The preparation method according to claim 3, characterized in that In the preparation method of compounds 11a-11V, The reagents and conditions are as follows: (a) p-toluenesulfonyl chloride, N,N-diisopropylethylamine, 4-dimethylaminopyridine, acetonitrile, 80°C, 6 hours; (b) n-propionaldehyde, N-methylmorpholine, anhydrous ethanol, room temperature, 30 hours; (c) N,N-diisopropylethylamine, N,N-dimethylformamide, 85°C, 5 hours; (d) isoamyl nitrite, cupric chloride, acetonitrile, 80°C, 2.5 hours; (e) aminopiperidine derivatives, carbon 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (g) 1) 9a-n, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 9o-t, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 3) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours.

5. The preparation method according to claim 3, characterized in that: In the preparation method of compounds 15a, 15b, 19a and 19b, The reagents and conditions are as follows: (a) aminopiperidine derivative, potassium carbonate, dichloromethane, 4 hours; (b) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (c) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (d) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (e) piperazine-1-tert-butylcarboxylate, potassium carbonate, dichloromethane, 4 hours; (f) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (g) 1) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 2) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours.

6. The preparation method according to claim 3, characterized in that: In the preparation methods of compounds 23a, (R)-23a, (S)-23a, 23b, 27, 30a-e, The reagents and conditions are as follows: (a) piperidine-4-aminomethyl tert-butyl carbamate, potassium carbonate, dichloromethane, 4 hours; (b) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (c) 1) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 2) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (d) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (e) tert-butyl (2-(methylamino)ethyl)carbamate, potassium carbonate, dichloromethane, 4 hours; (f) 4M hydrochloric acid / ethyl acetate ethyl acetate, room temperature, 12 hours; (g) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (i) 1) 28a-c, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 28d, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 3) 28e, potassium carbonate, acetonitrile, 80°C, 6 hours; (j) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours.

7. The preparation method according to claim 3, characterized in that: In the preparation method of compound 40a-d, The reagents and conditions are as follows: (a) methanesulfonyl chloride, triethylamine, dichloromethane, room temperature, 4 hours; (b) diisobutylaluminum hydride, tetrahydrofuran, -20°C, 6 hours; (c) p-toluenesulfonyl chloride, triethylamine, DMAP, acetonitrile, 80°C, 4 hours; (d) 4, triethylamine, DMF, 85°C, 5 hours; (e) isoamyl nitrite, cupric chloride, acetonitrile, 80°C, 2.5 hours; (f) 2-methylamino derivative, potassium carbonate, dichloromethane, 4 hours; (g) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (h) 1) 9a or 9b, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (i) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours.

8. The preparation method according to claim 3, characterized in that: In the preparation method of compounds 44a-c and 48, Reagents and conditions are as follows: (a) tert-butyl 4-carbamate, potassium carbonate, dichloromethane, 4 hours; (b) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (c) 1) 9b, isobutyl chloroformate, N-methylmorpholine, dichloromethane / tetrahydrofuran (1:1), room temperature, 6 hours; 2) 9p, sodium triacetoxyborohydride, acetic acid, room temperature, 12 hours; 3) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (d) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (e) piperazine-1-tert-butylcarboxylate, potassium carbonate, dichloromethane, 4 hours; (f) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours; (g) 9u, potassium carbonate, acetonitrile, 80°C, 6 hours; (h) 4M hydrochloric acid / ethyl acetate, room temperature, 12 hours.

9. A pharmaceutical composition comprising a therapeutically effective amount of the DNMT1 and HDAC dual-target inhibitor according to claim 1 and a pharmaceutically acceptable excipient.

10. Use of the DNMT1-HDAC dual-target inhibitor according to claim 1 in the preparation of a drug for treating tumor diseases related to the target; The tumor is lung cancer, liver cancer, non-small cell lung cancer, colon cancer, skin cancer, pancreatic cancer, ovarian cancer, breast cancer, bladder cancer, lymphoma, esophageal cancer, gastrointestinal cancer, nasopharyngeal cancer, leukemia or myeloma.

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