Ezh2 / hdac dual-targeted inhibitors and methods of making and medical uses thereof

Abstract

The application discloses an EZH2 / HDAC dual-target inhibitor and a preparation method and medical use thereof. The application relates to a compound with a structure as shown in general formula (I)-(V) or a pharmaceutically acceptable salt thereof, wherein R is selected from C5-C9 alkyl, a carbonyl alkyl chain, benzyl, a pyrimidine ring and an unsaturated five-membered heterocyclic ring. The application provides a novel EZH2 / HDAC dual-target inhibitor. The disclosed compound can effectively target EZH2 and HDAC, and can be used as an effective therapeutic agent for blood tumors and other cancers.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of pharmaceutical chemistry, in particular to the preparation and medical use of EZH2 / HDAC dual-targeted inhibitors. BACKGROUND

[0002] Currently, most of the approved drugs based on "single target single drug" are increasingly poor in treating complex, heterogeneous, and multi-gene cancer diseases. In this case, two or more drugs are used in preclinical and clinical settings to combine different targets, however, drug combination therapy is often denied due to poor drug interactions, unpredictable pharmacokinetics (PK) and safety profiles, and poor patient compliance. Recently, dual-targeted drugs have attracted high attention from medical researchers. Compared with single-targeted drugs, dual-targeted drugs can inhibit two pathways involved in disease progression and produce synergistic or additive effects.

[0003] Histone deacetylases (HDACs), also known as lysine deacetylases (KDACs), are important for removing acetyl groups from acetylated lysine residues of different protein substrates including non-histone and histone proteins. Overexpression of HDACs is associated with a series of human diseases, including some cancers and central nervous system (CNS) diseases. Therefore, HDACs represent a potential target for cancer treatment. Histone deacetylase (HDAC) inhibitors are powerful epigenetic modulators with great therapeutic potential and pleiotropic effects at the cellular and systemic levels.

[0004] Polycomb Repressive Complex 2 (PRC2) consists of three essential core subunits: Enhancer of Zeste Homolog (EZH2), Embryonic Ectoderm Development (EED), and Suppressor of Zeste Homolog (SUZ12). PRC2 transfers methyl S-adenosyl-l-methionine (SAM) from a cofactor to the epsilon-amino group of histone 3 lysine 27 (H3K27) through each catalytic cycle. EZH2 is the enzymatic subunit of PRC2. EZH2 has been found to be frequently overexpressed in prostate, breast, kidney, lung, myeloma, and lymphoma. As an epigenetic regulator, EZH2 has been identified as a driver of lymphoma development. Tazemetostat is the first selective inhibitor of EZH2-WT and mutants, which has been approved by FDA for the treatment of epithelioid sarcoma. In addition, other molecules also show good EZH2 inhibition activity, including GSK126, CPI-1205. C24, a selective inhibitor of EZH2, has high potency (IC 50 = 12 nM) for EZH2, and C24 has more than 200 times selectivity for EZH2 (IC 50 > 2.5 μM) over the highly homologous H3K27 methyltransferase EZH1.

[0005] Abnormal epigenetic modifications, coupled with genetic mutations and tumor suppressor silencing, are critical for the development and maintenance biology of hematological malignancies. HDAC and EZH2 inhibitors have been shown to be important epigenetic modulators, and there are many studies on their application in human hematological malignancies. At the same time, many researchers combine HDAC and EZH2 inhibitors to improve the efficacy of treating various blood cancers. It is shown that simultaneous inhibition of EZH2 and HDAC has a high degree of synergy in human hematological malignancies. Based on this information, the rational combination of EZH2 and HDAC inhibitors is a potential therapeutic strategy for the treatment of hematological malignancies. However, combination therapy is usually accompanied by poor patient compliance, unpredictable pharmacokinetic (PK) characteristics and drug interactions. The strategy to overcome these shortcomings is to develop dual inhibitors of HDAC and EZH2. SUMMARY

[0006] The purpose of the application is to develop a new type of EZH2 / HDAC dual-target inhibitor for the treatment of hematological tumors and other diseases related to EZH2 and HDAC.

[0007] Technical solution: A new type of EZH2 / HDAC dual-target inhibitor, comprising a compound with the following structure:

[0008]

[0009] wherein R is selected from -(CH2) n1 -, -CO(CH2) n2 -, benzyl, pyrimidine ring, unsaturated five-membered heterocyclic ring; n1 is any integer from 4 to 9, and n2 is any integer from 3 to 6.

[0010] As a preferred embodiment of the present application, when the dual-target inhibitor has the structure of general formula (I), R is selected from -(CH2) n1 -, -CO(CH2) n2 -, pyrimidine ring, n1 is 7 or 8, and n2 is 5 or 6; when the dual-target inhibitor has the structure of general formula (II), R is selected from -CO(CH2) n2 -, and n2 is 5 or 6; when the dual-target inhibitor has the structure of general formula (III), R is selected from -(CH2) n1 -, and n1 is 5, 6 or 7; when the dual-target inhibitor has the structure of general formula (V), R is selected from a pyrimidine ring.

[0011] The compound of the present application is selected from any one of the following:

[0012]

[0013] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxyamino)-4-oxobutyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0014]

[0015] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(5- (hydroxyamino)-5-oxopentyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0016]

[0017] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(6- (hydroxyamino)-6-oxohexyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0018]

[0019] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(7- (hydroxyamino)-7-oxoheptyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0020]

[0021] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(8- (hydroxyamino)-8-oxooctyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0022]

[0023] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(9- (hydroxyamino)-9-oxononyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0024]

[0025] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxycarbamoyl)benzyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0026]

[0027] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxycarbamoyl)benzyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0028]

[0029] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxycarbamoyl)benzyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0030]

[0031] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxycarbamoyl)benzyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0032]

[0033] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxycarbamoyl)benzyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0034]

[0035] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxycarbamoyl)benzyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide

[0036]

[0037] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(6-(hydroxyamino)- 6-oxohexanoyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4-carboxamide

[0038]

[0039] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(7-(hydroxyamino)- 7-oxoheptyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4-carboxamide

[0040]

[0041] N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(8-(hydroxyamino)- 8-oxooctanoyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4-carboxamide

[0042]

[0043] Nl-(4-(4-(((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)carbamoyl)-l- isopropyl-lH-indazol-6-yl)phenyl)-N8-hydroxyoctanediamide

[0044]

[0045] Nl-(4-(4-(((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)carbamoyl)-l- isopropyl-lH-indazol-6-yl)phenyl)-N7-hydroxyheptanediamide

[0046]

[0047] tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)carbamoyl)- 5-((5-(hydroxyamino)-5-oxopentyl)amino)-4-methylphenyl)pyridin-2-yl)piperazine- 1-carboxylate

[0048]

[0049] tert-Butyl 4-(5-(3-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)carbamoyl)-5-((7-(hydroxyamino)-7-oxoheptyl)amino)-4-methylphenyl)pyridin-3-yl)piperazine-1-carboxylate

[0050]

[0051] tert-Butyl 4-(5-(3-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)carbamoyl)-5-((7-(hydroxyamino)-7-oxoheptyl)amino)-4-methylphenyl)pyridin-3-yl)piperazine-1-carboxylate

[0052]

[0053] tert-Butyl 4-(5-(3-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)carbamoyl)-5-((7-(hydroxyamino)-7-oxoheptyl)amino)-4-methylphenyl)pyridin-3-yl)piperazine-1-carboxylate

[0054]

[0055] tert-Butyl 4-(5-(3-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)carbamoyl)-5-((7-(hydroxyamino)-7-oxoheptyl)amino)-4-methylphenyl)pyridin-3-yl)piperazine-1-carboxylate

[0056]

[0057] tert-Butyl 4-(5-(3-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)carbamoyl)-5-((7-(hydroxyamino)-7-oxoheptyl)amino)-4-methylphenyl)pyridin-3-yl)piperazine-1-carboxylate

[0058]

[0059] (R)-1-(sec-butyl)-N-(4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(9-(hydroxyamino)-9-oxononyl)piperazin-1-yl)pyridin-3-yl)-3-methyl-1H-indole-4-carboxamide

[0060]

[0061] (R)-1-(sec-butyl)-N-(4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(5- (hydroxycarbamoyl)pyrimidin-2-yl)piperazin-1-yl)pyridin-3-yl)-3-methyl-1H-indole-4- carboxamide

[0062]

[0063] N-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-3-(ethyl(tetrahydro-2H-pyran- 4-yl)amino)-5-(6-(4-(9-(hydroxylamino)-9-oxononyl)piperazin-1-yl)pyridin-3-yl)-2- methylbenzamide

[0064]

[0065] 2-(4-(5-(3-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)carbamoyl)-5-(ethyl(tetrahydro-2H-pyran-4-yl)amino)-4-methylphenyl)pyridin-2-yl)piperazin-1-yl)-N- hydroxypyrimidine-5-carboxamide as a preferred embodiment of the present application, the pharmaceutically acceptable salt of the compound of general formula I-V is selected from the acid addition salt of the compound of general formula I-V with an acid selected from the group consisting of hydrogen chloride, hydrogen bromide, sulfuric acid, carbonic acid, oxalic acid, citric acid, succinic acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid or ferulic acid.

[0066] A pharmaceutical composition, wherein the compound of general formula I-V or a pharmaceutically acceptable salt thereof according to the present application and a pharmaceutically acceptable carrier.

[0067] As a preferred embodiment of the present application, the pharmaceutical composition is prepared into any one of the dosage forms of tablets, capsules, powders, syrups, liquids, suspensions, freeze-dried powders or injections.

[0068] A preparation method of the EZH2 / HDAC dual-target inhibitor according to the present application:

[0069] Scheme 1: Synthesis of target compounds 10-24 a .

[0070]

[0071] aReagents and reaction conditions: (a) 2-iodopropane, K2CO3, CH3CN, reflux, 12 h, 75%; (b) tert-butyl 4-(5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)pyridin-2-yl)piperazine- 1-carboxylate, PdCl2(dppf)-CH2Cl2, K2CO3, DMF, reflux, N2, 4 h, 80%; (c) NaOH, MeOH / H2O, rt, 2 h, 95%; (d) 3-(aminomethyl)-4,6-dimethylpyridin-2(lH)-one, HATU, DIPEA, DCM, rt, 2 h, 84%; (e) CF3COOH, DCM, rt, 6 h, 85%; (f) for 10g-20g, various methyl bromoformates, Et3N, DMF, 90 °C, 12 h, 70-85%; for 21g-24g, various methyl monoformates, HATU, DIPEA, DCM, rt, 2 h, 84-94%; (g) NH2OK, CH3OH, rt, 3-5 h, 63-85%;

[0072] b For the linker R of the designed compounds, the left side is connected to a nitrogen atom and the right side is connected to a carbonyl group.

[0073] Scheme 2: Synthesis of target compounds 25, 26 a .

[0074]

[0075] a Reagents and reaction conditions: (a) tert-butyl (4-(4,4,5,5-tetramethyl-l,3,2- dioxaborolan-2-yl)phenyl)carbamate, PdCl2(dppf)-CH2Cl2, K2CO3, DMF, reflux, N2, 4 h, 84%; (b) NaOH, MeOH / H2O, rt, 2 h, 92%; (c) 3-(aminomethyl)-4,6-dimethylpyridin- 2(lH)-one, HATU, DIPEA, DCM, rt, 2 h, 88%; (d) CF3COOH, DCM, rt, 6 h, 79%; (e) various methyl monoformates, HATU, DIPEA, DCM, rt, 2 h, 84-90%; (f) NH2OK, CH3OH, rt, 3-5 h, 63-85%

[0076] b For the linker R of the designed compounds, the left side is connected to a nitrogen atom and the right side is connected to a carbonyl group.

[0077] Scheme 3: Synthesis of target compounds 27-32 a .

[0078]

[0079] a Reagents and reaction conditions: (a) 4-(5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2- yl)pyridin-2-yl)piperazine- 1 -carboxylic acid tert-butyl ester, PdCl2(dppf)-CH2Cl2, K2CO3, DMF, reflux, N2, 3 h, 70%; (b) NaOH, MeOH / H2O, rt, 2 h, 90%; (c) 3-(aminomethyl)-4,6-dimethylpyridin-2(lH)-one, HATU, DIPEA, DCM, rt, 2 h, 85%; (d) zinc powder, CH3COOH / H2O, 0 °C-rt, 12 h, 65%; (e) for 27-30, various methyl bromoformates, Cs2CO3, DMF, 90 °C, 1 h, 50-65%; for 31 and 32, various methyl monoformates, Et3N, DCM, rt, 1 h, 70-81%; (f) NH2OK, CH3OH, rt, 3-5 h, 60-82%; b For the linker R of the designed compounds, the left side is connected to a nitrogen atom and the right side is connected to a carbonyl group.

[0080] Scheme 4: Synthesis of target compounds 33, 34 a .

[0081]

[0082] a Reagents and reaction conditions: (a) 9-bromononanoic acid methyl ester or 2-chloropyrimidine-5-carboxylic acid methyl ester, Et3N, DMF, 90 °C, 12 h, 70-80%; (b) NH2OK, CH3OH, rt, 3-5 h, 64-80%;

[0083] b For the linker R of the designed compounds, the left side is connected to a nitrogen atom and the right side is connected to a carbonyl group.

[0084] Scheme 5: Synthesis of target compounds 35, 36 a .

[0085]

[0086] aReagents and reaction conditions: (a) 4-(5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2- yl)pyridin-2-yl)piperazine-l-carboxylic acid tert-butyl ester, PdCl2(dppf)-CH2Cl2, K2CO3, DMF, reflux, N2, 4 h, 80%; (b) NaOH, MeOH / H2O, rt, 2 h, 92%; (c) 3-(aminomethyl)-4,6-dimethylpyridin-2(lH)-one, HATU, DIPEA, DCM, rt, 2 h, 87%; (d) CF3COOH, DCM, rt, 6 h, 88%; (e) methyl 9-bromononanoate or methyl 2-chloropyrimidine-5-carboxylate, Et3N, DMF, 90 °C, 12 h, 70-85%; (f) NH2OK, CH3OH, rt, 3-5 h, 68-80%.

[0087] b For the designed compound, linker R is connected to the nitrogen atom on the left and to the carbonyl group on the right. BRIEF DESCRIPTION OF DRAWINGS

[0088] Figure 1 In vivo anti-tumor test. (A) Tumor weight on day 20 after treatment; (B) growth curve of transplanted MV4-11 xenograft in nude mice; (C) picture of dissected HMV4-11 tumor tissue; (D) body weight measurement. DETAILED DESCRIPTION

[0089] Example 1: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxyamino)-4-oxobutyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide (10):

[0090] Compound 10a (255.1 mg, 1.0 mmol) and 2-iodopropane (255.0 mg, 1.5 mmol) were dissolved in acetonitrile (10 mL) followed by the addition of potassium carbonate (276.4 mg, 2 mmol). The reaction was refluxed at 50 °C for 12 h and 10b was isolated by column. Then 10b (148.6 mg, 0.5 mmol) and tert-butyl 4-(5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)pyridin-2-yl)piperazine-l-carboxylate (467.16 mg, 0.6 mmol) were dissolved in DMF (10 mL), [l,l'-bis(diphenylphosphino)ferrocene]dichloropalladium(ll) and dichloromethane (27.3 mg, 0.05 mmol) and potassium carbonate (added 138.2 mg, 1 mmol) were added under nitrogen protection, the reaction was refluxed at 80 °C for 4 h to give compound 10c, which was hydrolyzed in CH3OH / H2O (5: 1) at room temperature for 2 h, then neutralized with dilute hydrochloric acid. Acid-amine condensation was carried out in the presence of DIPEA (12.9 mg, 0.1 mmol), HATU (570.5 mg, 1.5 mmol), DCM (10 mL), and then 10e was isolated by column. 10e (599.7 mg, 1 mmol) was deprotected in TFA / DCM (1:5) to give 10f, then 10f (499.6 mg, 1 mmol) was reacted with 4-bromobutyl methyl carbonate (362.1 mg, 2 mmol) in the presence of cesium carbonate at 90 °C for 12 h to give 10g. Then 10g was dissolved in NH2OK / CH3OH (10 mL) and stirred at room temperature for 4 h. When the reaction was complete, the solvent was evaporated under reduced pressure to give a residue, which was dissolved in water (15 mL). The mixture was neutralized by adding 1M HC1. The white precipitate was filtered and dried to give compound 10 with a yield of 34% as a cream-colored solid. 1 H NMR (600 MHz, DMSO-d6) δ 11.56 (s, 1H), 10.47 (s, 1H), 8.72 (s, 1H), 8.67 - 8.58 (m, 2H), 8.36 (s, 1H), 8.06 (d, J = 9.0 Hz, 2H), 7.85 (s, 1H), 6.95 (d, J = 8.9 Hz, 1H), 5.89 (s, 1H), 5.14 (p, J = 6.7 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.56 (s, 4H), 2.49 (s, 4H), 2.38 - 2.25 (m, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 2.02 (t, J = 7.4 Hz, 2H), 1.71 (p, J = 7.4 Hz, 2H), 1.49 (d, J = 6.6 Hz, 6H). 13C NMR (126 MHz, DMSO-d6) δ 169.5, 166.3, 163.6, 158.9, 150.2, 146.8, 143.3, 140.2, 136.8, 135.5, 133.2, 128.3, 125.0, 122.1, 120.8, 119.2, 108.9, 108.1, 107.3, 57.7, 52.8, 49.6, 45.1, 35.7, 30.7, 22.7, 22.6, 19.5, 18.7. HRMS: [M + Na] + calcd for C 32 H 40 N8NaO4, 437.1972, 623.3065, found 623.3066.

[0091] Example 2: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(5- (hydroxyamino)-5-oxopentyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide (11):

[0092] Compound 11 was prepared by replacing 4-bromobutanoic acid methyl ester in Example 1 with 5-bromopentanoic acid methyl ester, and following the same procedure as in Example 1. 1 H NMR (600 MHz, DMSO-d6) δ 11.56 (s, 1H), 10.47 (s, 1H), 8.72 (s, 1H), 8.67 - 8.58 (m, 2H), 8.36 (s, 1H), 8.06 (d, J = 9.0 Hz, 2H), 7.85 (s, 1H), 6.95 (d, J = 8.9 Hz, 1H), 5.89 (s, 1H), 5.14 (p, J = 6.7 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.56 (s, 4H), 2.49 (s, 4H), 2.38 - 2.25 (m, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 2.02 (t, J = 7.4 Hz, 2H), 1.71 (p, J = 7.4 Hz, 2H), 1.49 (d, J = 6.6 Hz, 6H). 13CNMR (126 MHz, DMSO-d6) δ 169.5, 166.3, 163.6, 158.9, 150.2, 146.8, 143.3, 140.2, 136.8, 135.5, 133.2, 128.3, 125.0, 122.1, 120.8, 119.2, 108.9, 108.1, 107.3, 57.7, 52.8, 49.6, 45.1, 35.7, 30.7, 22.7, 22.6, 19.5, 18.7. HRMS: [M + Na] + calcd for C 32 H 42 N8NaO4, 637.3221, found 637.3224.

[0093] Example 3: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(6-(hydroxyamino)-6-oxohexyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (12):

[0094] Compound 12 was prepared by replacing 4-bromobutanoic acid methyl ester in Example 1 with 6-bromohexanoic acid methyl ester, and following the same procedure as in Example 1. 1 H NMR (600 MHz, DMSO-d6) δ 10.41 (s, 1H), 8.65 (d, J = 2.5 Hz, 1H), 8.60 (t, J = 4.9 Hz, 1H), 8.35 (s, 1H), 8.06 (d, J = 5.8 Hz, 2H), 7.84 (s, 1H), 7.79 (s, 1H), 6.95 (d, J = 8.9 Hz, 1H), 5.90 (s, 1H), 5.13 (p, J = 6.6 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.55 (s, 4H), 2.47 (d, J = 5.1 Hz, 4H), 2.30 (t, J = 7.4 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.96 (t, J = 7.3 Hz, 2H), 1.53 (d, J = 7.5 Hz, 2H), 1.50 (d, J = 6.4 Hz, 6H), 1.46 (d, J = 7.7 Hz, 2H), 1.28 (d, J = 7.5 Hz, 2H). 13C NMR (151 MHz, DMSO-d6) δ 169.5, 166.3, 166.0, 163.6, 158.9, 157.5, 150.2, 146.8, 143.3, 140.2, 136.8, 135.5, 133.2, 128.3, 125.0, 122.1, 120.8, 119.2, 108.9, 108.0, 107.3, 58.4, 53.1, 49.6, 45.2, 35.7, 32.7, 27.0, 26.5, 25.6, 22.6, 19.4, 18.7. HRMS: [M+Na] + calcd for C 34 H 44 N8NaO4, 651.3378, found 651.3379.

[0095] Example 4: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(7-(hydroxyamino)-7-oxoheptyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (13):

[0096] The 4-bromobutylate in Example 1 was replaced with methyl 7-bromoheptanoate, and other procedures were the same as Example 1 to prepare compound 13, 1 H NMR (600 MHz, DMSO-d6) δ 10.40 (s, 1H), 8.65 (d, J = 2.6 Hz, 1H), 8.61 (t, J = 5.0 Hz, 1H), 8.36 (s, 1H), 8.09 - 8.04 (m, 2H), 7.84 (d, J = 1.2 Hz, 1H), 6.95 (d, J = 8.9 Hz, 1H), 5.89 (s, 1H), 5.14 (p, J = 6.6 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.55 (t, J = 5.1 Hz, 4H), 2.46 (t, J = 5.0 Hz, 4H), 2.30 (t, J = 7.4 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.95 (t, J = 7.4 Hz, 2H), 1.51 (d, J = 7.1 Hz, 2H), 1.49 (d, J = 6.6 Hz, 6H), 1.47 - 1.44 (m, 2H), 1.30 - 1.26 (m, 4H). 13C NMR (151 MHz, DMSO-d6) δ 169.5, 166.3, 166.0, 163.6, 158.9, 150.2, 146.8, 143.3, 140.2, 136.8, 135.5, 133.2, 128.3, 125.0, 122.1, 120.8, 119.2, 108.9, 108.0, 107.3, 58.4, 53.1, 49.6, 45.2, 35.7, 32.7, 29.0, 27.2, 26.7, 25.6, 22.6, 19.4, 18.7. HRMS: [M + Na] + calcd for C 35 H 46 N8NaO4, 665.3534, found 665.3538.

[0097] Example 5: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(8-(hydroxyamino)-8-oxooctyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (14):

[0098] Compound 14 was prepared by replacing 4-bromobutyl acid methyl ester in Example 1 with 8-bromooctanoic acid methyl ester, and following the same procedures as in Example 1. 1 H NMR (600 MHz, DMSO-d6) δ 10.41 (s, 1H), 8.66 (d, J = 2.5 Hz, 1H), 8.61 (s, 1H), 8.35 (s, 1H), 8.22 (s, 1H), 8.06 (d, J = 6.7 Hz, 2H), 7.96 (s, 1H), 7.84 (s, 1H), 6.96 (d, J = 8.9 Hz, 1H), 5.89 (s, 1H), 5.14 (p, J = 6.6 Hz, 1H), 4.38 (d, J = 4.9 Hz, 2H), 3.58 (q, J = 6.7, 5.4 Hz, 4H), 2.50 (s, 4H), 2.36 (q, J = 6.4, 5.9 Hz, 2H), 2.21 (s, 3H), 2.13 (s, 3H), 1.95 (t, J = 7.4 Hz, 2H), 1.49 (d, J = 6.6 Hz, 6H), 1.47 (d, J = 7.8 Hz, 4H), 1.32 - 1.25 (m, 5H). 13C NMR (151 MHz, DMSO-d6) δ 169.6, 166.3, 163.6, 162.8, 150.2, 146.8, 143.3, 140.2, 136.8, 135.5, 133.2, 130.1, 128.3, 125.1, 122.1, 120.8, 119.2, 108.9, 108.0, 107.3, 58.3, 52.9, 49.6, 45.0, 36.3, 35.7, 32.7, 31.2, 29.0, 27.3, 25.6, 22.6, 19.4, 18.7. HRMS: [M+Na]+calcd for C 36 H 40 N8NaO4, 671.3065, found 671.3067.

[0099] Example 6: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(9-(hydroxyamino)-9-oxononyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (15):

[0100] The 4-bromobutanoic acid methyl ester in Example 1 was replaced with 9-bromononanoic acid methyl ester, and other procedures were the same as Example 1 to prepare compound 15, 1 H NMR (600 MHz, DMSO-d6) δ 10.39 (s, 1H), 8.65 (d, J = 2.5 Hz, 1H), 8.61 (t, J = 5.0 Hz, 1H), 8.36 (s, 1H), 8.06 (d, J = 9.2 Hz, 2H), 7.84 (s, 1H), 6.96 (d, J = 8.9 Hz, 1H), 5.90 (s, 1H), 5.14 (q, J = 6.6 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.55 (t, J = 5.1 Hz, 4H), 2.47 (t, J = 5.2 Hz, 4H), 2.31 (t, J = 7.4 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.94 (t, J = 7.4 Hz, 2H), 1.49 (d, J = 6.6 Hz, 6H), 1.46 (d, J = 8.1 Hz, 4H), 1.28 (s, 8H). 13C NMR (151 MHz, DMSO-d6) δ 169.6, 166.3, 163.6, 158.9, 150.2, 146.8, 143.3, 140.2, 136.8, 135.5, 133.2, 128.3, 125.0, 122.1, 120.8, 119.2, 108.9, 108.0, 107.3, 58.4, 53.0, 49.6, 45.2, 35.7, 32.7, 29.4, 29.2, 29.0, 27.4, 26.7, 25.6, 22.6, 19.4, 18.7. HRMS: [M+Na]+calcd for C 37 H 50 N8NaO4 693.3847, found 693.3844.

[0101] Example 7: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(10- (hydroxyamino)-10-oxodecyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide (16):

[0102] The 4-bromobutyl methyl carbonate in Example 1 was replaced with 10-bromodecanoic acid methyl ester, and other operation steps were the same as Example 1 to prepare compound 16, 1 H NMR (600 MHz, DMSO-d6) δ 11.56 (s, 1H), 10.37 (s, 1H), 8.65 (d, J = 2.6 Hz, 1H), 8.60 (s, 1H), 8.35 (s, 1H), 8.28 (s, 2H), 8.06 (d, J = 8.2 Hz, 2H), 7.84 (s, 2H), 6.95 (d, J = 8.9 Hz, 1H), 5.90 (s, 1H), 5.15 - 5.12 (m, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.55 (s, 4H), 2.46 (t, J = 5.1 Hz, 4H), 2.31 (t, J = 7.4 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.94 (t, J = 7.2 Hz, 2H), 1.49 (d, J = 6.5 Hz, 6H), 1.47 (t, J = 7.2 Hz, 4H), 1.29 - 1.27 (m, 4H), 1.26 (d, J = 8.0 Hz, 6H). 13C NMR (151 MHz, DMSO-d6) δ 166.4, 163.6, 158.9, 150.3, 146.7, 143.4, 140.2, 136.8, 135.5, 133.2, 130.1, 128.3, 125.0, 122.0, 120.8, 119.2, 108.9, 108.1, 107.3, 58.4, 53.0, 49.6, 45.1, 35.7, 32.7, 30.3, 29.4, 29.2, 29.0, 27.4, 26.6, 25.6, 22.6, 19.4, 18.7. [M+Na]+calcd for C 38 H 52 N8NaO4, 707.4004, found 707.4006.

[0103] Example 8: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(4- (hydroxycarbamoyl)benzyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide (17):

[0104] Compound 17 was prepared by replacing 4-bromomethyl benzoate with 4- bromomethyl benzoate in Example 1, and following the procedures as described in Example 1. 1 H NMR (600 MHz, DMSO-d6) δ 8.65 (d, J = 2.5 Hz, 1H), 8.59 (t, J = 5.0 Hz, 1H), 8.38 (s, 2H), 8.35 (s, 1H), 8.05 (d, J = 7.1 Hz, 2H), 7.83 (d, J = 1.3 Hz, 1H), 7.74 (d, J = 7.9 Hz, 2H), 7.43 (d, J = 7.9 Hz, 2H), 6.95 (d, J = 8.9 Hz, 1H), 5.89 (s, 1H), 5.13 (p, J = 6.6 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.58 (d, J = 6.8 Hz, 11H), 2.22 (s, 3H), 2.13 (s, 3H), 1.49 (d, J = 6.5 Hz, 6H). 13C NMR (151 MHz, DMSO-d6) δ 166.3, 165.6, 163.6, 158.9, 150.2, 146.8, 143.3, 141.9, 140.2, 136.8, 135.5, 133.2, 132.1, 129.2, 128.3, 127.3, 125.1, 122.1, 120.8, 119.2, 108.9, 108.0, 107.3, 55.4, 52.8, 49.1, 45.2, 35.7, 22.6, 19.4, 18.7. [M+Na]+calcd for C 36 H 40 N8NaO4 671.3065, found 671.3067.

[0105] Example 9: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-((5-(hydroxycarbamoyl)furan-2-yl)methyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (18)

[0106] The compound 18 was prepared by replacing 4-bromobutyl acid methyl ester in Example 1 with 5-(chloromethyl)2-furancarboxylic acid methyl ester, and following the same procedure as in Example 1. 1 H NMR (600 MHz, DMSO-d6) δ 8.65 (d, J = 2.6 Hz, 1H), 8.60 (t, J = 5.0 Hz, 1H), 8.35 (s, 1H), 8.27 (s, 1H), 8.06 (d, J = 8.1 Hz, 2H), 7.83 (s, 1H), 7.00 (s, 1H), 6.95 (d, J = 8.9 Hz, 1H), 6.46 (d, J = 3.3 Hz, 1H), 5.89 (s, 1H), 5.13 (p, J = 6.6 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.57 (d, J = 4.7 Hz, 4H), 3.17 (s, 2H), 2.53 (s, 4H), 2.22 (s, 3H), 2.13 (s, 3H), 1.49 (d, J = 6.5 Hz, 6H). 13C NMR (151 MHz, DMSO-d6) δ 166.4, 163.7, 158.8, 154.3, 150.4, 146.7, 146.0, 143.4, 140.1, 136.9, 135.5, 133.2, 130.1, 128.3, 125.1, 122.0, 120.8, 119.2, 111.1, 109.0, 108.2, 107.4, 54.4, 52.5, 49.6, 49.1, 45.1, 22.6, 21.9, 19.4, 18.7. [M+Na]+calcd for C 34 H 38 N8NaO5, 661.2857, found 661.2861.

[0107] Example 10: Preparation of N1-(4-((4-(5-(4-(((4,6-dimethyl-2-oxo-1,2-dihydro- pyridin-3-yl)methyl)carbamoyl)-1-isopropyl-1H-indazol-6-yl)pyridin-2-yl)piperazin-1- yl)methyl)phenyl)-N8-hydroxyoctanediamide (19)

[0108] The 4-bromobutylate in Example 1 was replaced with 8-((4- (bromomethyl)phenyl)amino)-8-oxooctanoate, and other procedures were the same as Example 1 to prepare compound 19, 1 H NMR (500 MHz, DMSO-d6) δ 11.58 (s, 1H), 10.20 (s, 1H), 8.72 (s, 1H), 8.70-8.57 (m, 2H), 8.35 (s, 1H), 8.07 (s, 2H), 7.84 (s, 1H), 7.63 (d, J = 8.0 Hz, 2H), 7.42-7.23 (m, 3H), 6.97 (d, J = 8.9 Hz, 1H), 5.90 (s, 1H), 5.13 (p, J = 6.7 Hz, 1H), 4.38 (d, J = 4.9 Hz, 2H), 3.59-3.50 (m, 4H), 3.16 (d, J = 5.0 Hz, 4H), 2.32 (t, J = 7.5 Hz, 2H), 2.21 (s, 3H), 2.13 (s, 3H), 1.95 (t, J = 7.3 Hz, 2H), 1.56 (t, J = 7.2 Hz, 2H), 1.48 (d, J = 6.6 Hz, 6H), 1.29 (s, 6H), 0.84 (t, J = 6.8 Hz, 2H). 13C NMR (151 MHz, DMSO-d6) δ 171.8, 169.6, 166.3, 163.6, 150.3, 147.6, 146.7, 143.4, 140.2, 136.9, 135.5, 133.2, 130.1, 128.3, 124.8, 124.7, 122.1, 120.8, 119.4, 119.2, 109.0, 108.1, 107.5, 49.6, 36.8, 35.7, 32.7, 31.7, 31.6, 30.3, 29.5, 28.9, 25.5, 25.5, 22.6, 19.5, 18.7, 14.4. [M+Na]+calcd for C 43 H 53 N9NaO5 798.4062, found 798.4064;

[0109] Example 11: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(5- (hydroxycarbamoyl)pyrimidin-2-yl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide (20):

[0110] Example 1, using 2-chloropyrimidine-5-carboxylic acid methyl ester instead of 4-bromobutyric acid methyl ester, to give compound 20, 1 H NMR (500 MHz, DMSO-d6) δ 11.55 (s, 1H), 11.27 (s, 1H), 9.04 (s, 1H), 8.76 (s, 1H), 8.69 (d, J = 2.6 Hz, 1H), 8.61 (t, J = 5.0 Hz, 1H), 8.35 (s, 1H), 8.11 (d, J = 8.7 Hz, 1H), 8.08 (s, 1H), 7.85 (s, 1H), 7.03 (d, J = 8.9 Hz, 1H), 5.90 (s, 1H), 5.14 (p, J = 6.7 Hz, 1H), 4.39 (d, J = 4.8 Hz, 2H), 3.98 (dt, J = 19.3, 5.2 Hz, 4H), 3.75 - 3.61 (m, 4H), 2.22 (s, 3H), 2.13 (s, 3H), 1.49 (d, J = 6.5 Hz, 6H). 13C NMR (151 MHz, DMSO-d6) δ 166.3, 163.6, 162.3, 161.9, 158.6, 157.7, 150.2, 146.8, 143.3, 140.2, 137.0, 135.4, 133.2, 128.3, 125.3, 122.1, 120.8, 119.2, 115.3, 109.0, 108.0, 107.5, 49.6, 44.8, 43.6, 35.7, 22.6, 19.4, 18.7. [M+Na]+calcd for C 33 H 36 N 10 NaO4 659.2813, found 659.2815.

[0111] Example 12: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(5- (hydroxyamino)-5-oxopentanoyl)piperazin-l-yl)pyridin-3-yl)-l-isopropyl-lH-indazole-4- carboxamide (21):

[0112] Compound 10f (499.6 mg, 1 mmol) was coupled with 5-methoxy-5-oxopentanoic acid (175.4 mg, 1.2 mmol) in the presence of DIPEA (12.9 mg, 0.1 mmol), HATU (570.5 mg, 1.5 mmol), DCM (10 mL) at room temperature for 2 hours, then 21g was isolated by column. 21g was dissolved in NH2OK / CH3OH (10 mL) and stirred at room temperature for 4 hours. When the reaction was completed, the solvent was evaporated under reduced pressure to obtain a residue, which was dissolved in water (15 mL). The mixture was neutralized by adding 1M hydrochloric acid. The white precipitate was filtered and dried to obtain compound 21 with a yield of 32%. 1 H NMR (600 MHz, DMSO-d6) δ 11.57 (s, 1H), 10.48 (s, 1H), 8.68 (d, J = 2.6 Hz, 1H), 8.61 (t, J = 5.0 Hz, 1H), 8.36 (s, 1H), 8.25 (s, 1H), 8.12 - 8.07 (m, 2H), 7.85 (s, 1H), 7.00 (d, J = 9.0 Hz, 1H), 5.90 (s, 1H), 5.14 (p, J = 6.6 Hz, 1H), 4.39 (d, J = 4.8 Hz, 2H), 3.63 - 3.56 (m, 8H), 2.36 (t, J = 7.6 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 2.03 (d, J = 7.2 Hz, 2H), 1.75 (p, J = 7.5 Hz, 2H), 1.49 (d, J = 6.6 Hz, 6H).13 C NMR (151 MHz, DMSO-d6) δ 170.9, 166.3, 163.6, 158.6, 150.2, 146.7, 143.3, 140.2, 136.9, 135.4, 133.2, 128.3, 125.3, 122.1, 120.8, 119.2, 109.0, 108.1, 107.5, 49.6, 45.3, 45.0, 44.9, 41.1, 35.7, 32.1, 30.2, 29.5, 22.6, 19.4, 18.7. [M+Na]+calcd for C 33 H 40 N8NaO5, 651.3014, found 651.3014.

[0113] Example 13: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(6-(hydroxyamino)-6-oxohexanoyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (22):

[0114] Compound 22 was prepared by replacing 5-methoxy-5-oxopentanoic acid in Example 12 with monomethyl adipate, and following the same procedure as Example 12. 1 H NMR (500 MHz, DMSO-d6) δ 10.37 (s, 1H), 8.68 (d, J = 2.5 Hz, 1H), 8.61 (d, J = 5.1 Hz, 1H), 8.36 (s, 1H), 8.23 (s, 1H), 8.10 (d, J = 8.7 Hz, 1H), 8.08 (s, 1H), 7.85 (s, 1H), 6.99 (d, J = 8.8 Hz, 1H), 5.90 (s, 1H), 5.19 - 5.01 (m, 1H), 4.39 (d, J = 4.8 Hz, 2H), 3.58 (dt, J = 16.2, 7.9 Hz, 8H), 2.35 (t, J = 7.4 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.96 (d, J = 8.3 Hz, 2H), 1.49 (d, J = 6.5 Hz, 6H), 1.32 - 1.25 (m, 4H). 13C NMR (126 MHz, DMSO-d6) δ 171.3, 166.3, 163.6, 158.6, 150.2, 146.8, 143.3, 140.2, 136.9, 135.4, 133.2, 128.3, 125.3, 122.1, 120.9, 119.2, 109.0, 108.0, 107.5, 49.6, 45.3, 45.0, 41.1, 35.7, 32.7, 29.0, 25.1, 22.6, 19.4, 18.7. [M+Na]+calcd for C 34 H 42 N8NaO5, 655.3170, found 655.3173.

[0115] Example 14: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(7-(hydroxyamino)-7-oxoheptyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (23):

[0116] The 5-methoxy-5-oxopentanoic acid in Example 12 was replaced with 7-methoxy-7- oxoheptanoic acid, and other operation steps were the same as Example 12 to prepare compound 23, 1 H NMR (600 MHz, DMSO-d6) δ 10.38 (s, 1H), 8.68 (d, J = 2.5 Hz, 1H), 8.61 (t, J = 5.0 Hz, 1H), 8.36 (s, 1H), 8.10 (dd, J = 8.8, 2.6 Hz, 1H), 8.08 (s, 1H), 7.85 (s, 1H), 6.99 (d, J = 8.9 Hz, 1H), 5.89 (s, 1H), 5.14 (p, J = 6.6 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.59 (td, J = 19.6, 18.2, 5.6 Hz, 8H), 2.35 (t, J = 7.5 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.95 (t, J = 7.4 Hz, 2H), 1.56 - 1.50 (m, 4H), 1.49 (d, J = 6.5 Hz, 6H), 1.27 (t, J = 7.3 Hz, 2H). 13C NMR (151 MHz, DMSO-d6) δ 171.2, 169.5, 166.3, 164.7, 163.6, 158.6, 157.6, 150.2, 146.8, 143.3, 140.2, 136.9, 135.4, 133.2, 128.3, 125.3, 122.1, 120.8, 119.2, 109.0, 108.0, 107.5, 49.6, 45.3, 45.0, 44.9, 41.1, 35.7, 32.7, 28.8, 25.5, 25.0, 22.6, 19.4, 18.7. [M+Na]+calcd for C 35 H 44 N8NaO5, 679.3327, found 679.3324.

[0117] Example 15: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(8-(hydroxyamino)-8-oxooctanoyl)piperazin-l-yl)pyridin-3-yl)-l- isopropyl-lH-indazole-4-carboxamide (24):

[0118] The procedure was same as Example 12 except that 5-methoxy-5-oxopentanoic acid was replaced by octanedioic acid monomethyl ester to give compound 24, 1 H NMR (500 MHz, DMSO-d6) δ 10.42 (s, 1H), 8.68 (d, J = 2.5 Hz, 1H), 8.61 (t, J = 4.9 Hz, 1H), 8.36 (s, 1H), 8.15 - 8.05 (m, 2H), 7.85 (s, 1H), 7.00 (d, J = 8.9 Hz, 1H), 5.90 (s, 1H), 5.14 (p, J = 6.7 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 3.59 (dq, J = 16.1, 5.8 Hz, 8H), 2.37 (t, J = 6.9 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.98 (t, J = 7.0 Hz, 2H), 1.50 (t, J = 7.6 Hz, 9H), 1.24 (s, 4H). 13CNMR (151 MHz, DMSO-d6) δ 171.3, 166.3, 163.6, 158.6, 150.2, 146.8, 143.3, 140.2, 136.9, 135.4, 133.2, 128.3, 125.3, 122.1, 120.8, 119.2, 109.0, 108.0, 107.5, 49.6, 45.3, 45.1, 45.0, 41.1, 35.7, 32.7, 29.0, 28.9, 25.6, 25.1, 22.6, 19.4, 18.7. [M+Na]+calcd for C 36 H 46 N8NaO5, 693.3483, found 693.3480.

[0119] Example 16: Preparation of N1-(4-(4-(((4,6-dimethyl-2-oxo-1,2-dihydro- pyridin-3-yl)methyl)carbamoyl)-1-isopropyl-1H-indazol-6-yl)phenyl)-N8- hydroxyoctanediamide (25):

[0120] Example 16: Preparation of N1-(4-(4-(((4,6-dimethyl-2-oxo-1,2-dihydro- pyridin-3-yl)methyl)carbamoyl)-1-isopropyl-1H-indazol-6-yl)phenyl)-N8- hydroxyoctanediamide (25): 1 H NMR (500 MHz, DMSO-d6) δ 11.53 (s, 1H), 10.39 (s, 1H), 10.09 (s, 1H), 8.68 (d, J = 9.1 Hz, 1H), 8.62 (t, J = 5.1 Hz, 1H), 8.36 (s, 1H), 8.07 (s, 1H), 7.85 (s, 1H), 7.81 (d, J = 8.4 Hz, 2H), 7.75 (d, J = 8.4 Hz, 2H), 5.89 (s, 1H), 5.15 (p, J = 6.6 Hz, 1H), 4.40 (s, 2H), 2.36 (s, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.96 (t, J = 7.4 Hz, 2H), 1.60 (p, J = 7.3 Hz, 2H), 1.50 (d, J = 6.5 Hz, 6H), 1.35 - 1.25 (m, 6H). 13C NMR (126 MHz, DMSO-d6) δ 171.9, 169.6, 166.4, 150.1, 140.1, 139.6, 137.7, 134.8, 133.2, 128.3, 128.1, 122.1, 121.0, 119.7, 109.7, 108.0, 49.6, 36.9, 35.8, 32.7, 28.9, 25.5, 25.5, 22.6, 19.4, 18.7. [M+Na]+calcd for C 33 H 40 N6NaO5, 623.2952, found 623.2957.

[0121] Example 17: Preparation of N1-(4-(4-(((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3- yl)methyl)carbamoyl)-1-isopropyl-1H-indazol-6-yl)phenyl)-N7-hydroxyheptanediamide (26):

[0122] Example 17: Preparation of N1-(4-(4-(((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3- yl)methyl)carbamoyl)-1-isopropyl-1H-indazol-6-yl)phenyl)-N7-hydroxyheptanediamide (26): 1 H NMR (500 MHz, DMSO-d6) δ 11.55 (s, 1H), 10.49 (s, 1H), 10.32 (s, 1H), 8.69 (s, 1H), 8.64 (t, J = 5.1 Hz, 1H), 8.35 (s, 1H), 8.07 (s, 1H), 7.85 (s, 1H), 7.83 - 7.76 (m, 4H), 5.89 (s, 1H), 5.15 (p, J = 6.6 Hz, 1H), 4.39 (d, J = 4.9 Hz, 2H), 2.36 (t, J = 7.3 Hz, 2H), 2.22 (s, 3H), 2.13 (s, 3H), 1.98 (t, J = 7.4 Hz, 2H), 1.60 (t, J = 7.5 Hz, 2H), 1.53 (d, J = 7.4 Hz, 2H), 1.49 (d, J = 6.5 Hz, 6H), 1.30 (t, J = 3.6 Hz, 2H). 13C NMR (151 MHz, DMSO-d6) δ 171.3, 169.0, 165.8, 163.0, 149.6, 142.7, 139.5, 139.1, 137.1, 134.1, 132.6, 127.7, 127.5, 121.5, 120.4, 119.1, 119.1, 109.1, 107.5, 49.0, 36.2, 35.1, 32.0, 28.1, 24.8, 24.7, 22.0, 18.9, 18.1. [M+Na]+calcd for C 32 H 38 N6NaO5, 609.2796, found 609.2799.

[0123] Example 18: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2- dihydropyridin-3-yl)methyl)carbamoyl)-5-((5-(hydroxyamino)-5-oxopentyl)amino)-4- methylphenyl)pyridin-2-yl)piperazine-l-carboxylate (27):

[0124] 27a (274.1 mg, 1 mmol) and tert-butyl 4-(5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2- yl)pyridin-2-yl)piperazine-1-carboxylate (584.0 mg, 1.5 mmol) were dissolved in DMF (10 mL), [l,l'-bis(diphenylphosphino)ferrocene]dichloropalladium(ll) and dichloromethane (54.6 mg, 0.01 mmol) and potassium carbonate (138.2 mg, 1 mmol) were added, under nitrogen protection, and the reaction was refluxed at 80 °C for 4 h to obtain compound 27b (365.0 mg, 0.8 mmol), which was hydrolyzed in CH3OH / H2O (5:1) at room temperature for 2 h, then neutralized with HC1 to obtain 27c. DIPEA (12.9 mg, 0.1 mmol), HATU (570.5 mg, 1.5 mmol), DCM (10 mL), then 27d (576.27 mg, 1 mmol) was isolated by silica gel column. 27d (576.27 mg, 1 mmol) was reduced to 27e (355.3 mg, 0.65 mmol) under the condition of zinc powder (327.0 mg 5 mmol), formic acid (1 mL), H2O (5 mL) at room temperature. 27e (546.7 mg, 1 mmol) was coupled with 5-bromopentanoic acid methyl ester (195.0 mg, 1 mmol) in the presence of cesium carbonate (325.8 mg, 1 mmol) to obtain 27f (396.4 mg, 0.6 mmol), which was then dissolved in NH2OK / CH3OH (10 mL) and stirred at room temperature for 4 h. When the reaction was completed, the solvent was evaporated under reduced pressure to obtain a residue, which was dissolved in water (15 mL). The mixture was neutralized by adding 1 M HC1. The white precipitate was filtered and dried to obtain compound 27 (364.0 mg, 0.55 mmol) with a yield of 33% as a light brown solid. 1 H NMR (600 MHz, DMSO-d6) δ 10.40 (s, 1H), 8.32 (d, J = 2.5 Hz, 1H), 8.00 (s, 1H), 7.73 (dd, J = 8.8, 2.6 Hz, 1H), 6.93 - 6.84 (m, 2H), 6.65 (s, 1H), 6.02 (s, 1H), 4.29 (d, J = 5.1 Hz, 2H), 3.91 (d, J = 7.2 Hz, 4H), 3.43 (s, 4H), 2.33 (s, 3H), 2.32 - 2.27 (m, 2H), 2.21 (s, 3H), 2.00 (s, 3H), 1.97 (d, J = 12.8 Hz, 2H), 1.57 - 1.50 (m, 4H), 1.42 (s, 9H). 13C NMR (151 MHz, DMSO-d6) δ 170.0, 169.4, 164.2, 162.6, 158.4, 154.5, 148.0, 145.4, 144.4, 139.2, 136.0, 135.3, 126.0, 122.1, 117.6, 113.1, 112.2, 109.7, 107.6, 79.5, 45.0, 44.0, 36.2, 32.3, 28.5, 28.0, 23.2, 20.0, 19.2, 14.0. [M+Na]+calcd for C 35 H 47 N7NaO6, 684.3480, found 684.3480.

[0125] Example 19: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2- dihydropyridin-3-yl)methyl)carbamoyl)-5-(6-(hydroxyamino)-6-oxohexyl)amino)-4- methylphenyl)pyridin-3-yl)piperazine-l-carboxylate (28):

[0126] Example 19: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2- dihydropyridin-3-yl)methyl)carbamoyl)-5-(6-(hydroxyamino)-6-oxohexyl)amino)-4- methylphenyl)pyridin-3-yl)piperazine-l-carboxylate (28): 1 H NMR (500 MHz, DMSO-d6) δ 10.53 (s, 1H), 8.31 (d, J = 2.5 Hz, 1H), 8.03 (d, J = 5.4 Hz, 1H), 7.73 (d, J = 2.4 Hz, 1H), 7.57 (d, J = 45.4 Hz, 2H), 6.90 (d, J = 13.5 Hz, 2H), 6.64 (s, 1H), 6.01 (s, 1H), 5.05 (s, 1H), 4.28 (d, J = 5.2 Hz, 2H), 3.88 (t, J = 8.0 Hz, 2H), 3.58 - 3.45 (m, 6H), 3.16 (s, 1H), 2.33 (s, 3H), 2.20 (s, 3H), 1.98 (d, J = 6.8 Hz, 3H), 1.96 (d, J = 7.4 Hz, 2H), 1.56 - 1.49 (m, 4H), 1.42 (s, 9H), 1.28 (d, J = 7.8 Hz, 2H). 13C NMR (126 MHz, DMSO-d6) δ 170.0, 169.4, 162.6, 158.4, 154.4, 147.8, 145.4, 144.4, 139.2, 136.0, 126.0, 122.2, 113.1, 109.6, 107.6, 79.5, 49.0, 45.0, 44.3, 36.2, 32.6, 28.5, 28.1, 26.5, 25.3, 20.0, 19.2, 14.1. [M+Na]+calcd for C36H49N7NaO6, 698.3637, found 698.3642.

[0127] Example 20: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3- yl)methyl)carbamoyl)-5-((7-(hydroxyamino)-7-oxoheptyl)amino)-4-methylphenyl)pyridin-3- yl)piperazine-l-carboxylate (29):

[0128] The 5-bromopentanoic acid methyl ester in Example 18 was replaced with 7-bromoheptanoic acid methyl ester, and other procedures were the same as Example 18 to prepare compound 29, 1 H NMR (600 MHz, DMSO-d6) δ 11.48 (s, 1H), 10.36 (s, 1H), 8.66 (s, 1H), 8.39 (d, J = 2.6 Hz, 1H), 8.03 (t, J = 5.1 Hz, 1H), 7.81 (dd, J = 8.8, 2.6 Hz, 1H), 6.91 (d, J = 8.8 Hz, 1H), 6.68 (dd, J = 18.9, 1.8 Hz, 2H), 5.86 (s, 1H), 4.92 (t, J = 5.6 Hz, 1H), 4.27 (d, J = 5.1 Hz, 2H), 3.55 - 3.49 (m, 4H), 3.44 (d, J = 5.3 Hz, 4H), 3.17 - 3.13 (m, 2H), 2.20 (s, 3H), 2.11 (s, 3H), 2.03 (s, 3H), 1.97 - 1.91 (m, 2H), 1.59 (t, J = 7.3 Hz, 2H), 1.50 (q, J = 7.6 Hz, 2H), 1.43 (s, 9H), 1.39 - 1.34 (m, 2H), 1.32 - 1.28 (m, 2H). 13C NMR (151 MHz, DMSO-d6) δ 170.0, 169.6, 163.5, 158.4, 154.4, 149.9, 147.7, 145.7, 143.2, 139.1, 136.2, 135.6, 126.5, 122.1, 118.2, 112.5, 107.9, 107.6, 107.3, 79.5, 45.0, 43.5, 35.4, 32.7, 29.0, 28.8, 28.5, 26.9, 25.6, 19.4, 18.7, 14.1. [M+Na]+calcd for C37H51N7NaO6, 712.3793, found 712.3793.

[0129] Example 21: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3- yl)methyl)carbamoyl)-5-((8-(hydroxyamino)-8-oxooctyl)amino)-4-methylphenyl)pyridin-2- yl)piperazine-l-carboxylate (30):

[0130] Example 21: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3- yl)methyl)carbamoyl)-5-((8-(hydroxyamino)-8-oxooctyl)amino)-4-methylphenyl)pyridin-2- yl)piperazine-l-carboxylate (30): 1 H NMR (600 MHz, DMSO-d6) δ 10.33 (s, 1H), 8.32 (d, J = 2.5 Hz, 1H), 8.24 (s, 1H), 7.99 (t, J = 5.2 Hz, 1H), 7.73 (dd, J = 8.8, 2.6 Hz, 1H), 6.93 - 6.83 (m, 2H), 6.65 (d, J = 1.9 Hz, 1H), 6.00 (s, 1H), 4.29 (d, J = 5.1 Hz, 2H), 3.90 (t, J = 7.9 Hz, 2H), 3.51 (dd, J = 6.8, 3.8 Hz, 4H), 3.45 - 3.41 (m, 4H), 3.17 (s, 1H), 2.33 (s, 3H), 2.21 (s, 3H), 2.00 (s, 3H), 1.92 (t, J = 7.4 Hz, 2H), 1.53 (q, J = 7.5 Hz, 2H), 1.50 - 1.45 (m, 2H), 1.43 (s, 7H), 1.38 - 1.24 (m, 6H). 13C NMR (151 MHz, DMSO-d6) δ 170.0, 169.6, 162.6, 158.4, 154.5, 148.0, 147.7, 145.4, 144.4, 139.2, 136.0, 135.3, 126.0, 122.1, 117.6, 113.1, 112.2, 109.6, 107.6, 79.5, 49.1, 45.0, 44.3, 36.3, 32.7, 28.9, 28.8, 28.5, 28.3, 26.8, 25.5, 20.0, 19.2, 14.0. [M+Na]+calcd for C 38 H 53 N7NaO6, 726.3950, found 726.3951.

[0131] Example 22: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3- yl)methyl)carbamoyl)-5-(5-(hydroxyamino)-5-oxopentanamido)-4-methylphenyl)pyridin-2- yl)piperazine-l-carboxylate (31):

[0132] Example 22: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3- yl)methyl)carbamoyl)-5-(5-(hydroxyamino)-5-oxopentanamido)-4-methylphenyl)pyridin-2- yl)piperazine-l-carboxylate (31): 1 H NMR (600 MHz, DMSO-d6) δ 10.46 (s, 1H), 9.50 (s, 1H), 8.40 (d, J = 2.6 Hz, 1H), 8.27 (t, J = 5.1 Hz, 1H), 8.24 (s, 2H), 7.83 (dd, J = 8.9, 2.6 Hz, 1H), 7.61 - 7.53 (m, 1H), 7.28 (d, J = 1.9 Hz, 1H), 6.93 (d, J = 8.9 Hz, 1H), 5.88 (s, 1H), 4.30 (d, J = 5.0 Hz, 2H), 3.53 (dd, J = 6.8, 3.9 Hz, 4H), 3.43 (t, J = 5.2 Hz, 4H), 2.37 (t, J = 7.4 Hz, 2H), 2.21 (s, 3H), 2.15 (s, 3H), 2.12 (s, 3H), 2.05 (t, J = 7.6 Hz, 2H), 1.83 (p, J = 7.5 Hz, 2H), 1.43 (s, 9H). 13C NMR (151 MHz, DMSO) δ 171.5, 169.1, 165.6, 163.5, 154.4, 150.2, 145.7, 143.3, 139.3, 137.9, 136.2, 134.9, 130.1, 128.9, 124.9, 124.1, 122.0, 108.1, 107.7, 79.5, 44.9, 35.6, 35.5, 35.4, 32.1, 29.5, 28.5, 21.9, 19.4, 18.7, 14.9. [M+Na]+calcd for C 35 H 45 N7NaO7, 698.3273, found 698.3275.

[0133] Example 23: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2- dihydropyridin-3-yl)methyl)carbamoyl)-5-(6-(hydroxyamino)-6-oxohexanamido)-4- methylphenyl)pyridin-2-yl)piperazine-l-carboxylate (32):

[0134] Example 23: Preparation of tert-butyl 4-(5-(3-((4,6-dimethyl-2-oxo-l,2- dihydropyridin-3-yl)methyl)carbamoyl)-5-(6-(hydroxyamino)-6-oxohexanamido)-4- methylphenyl)pyridin-2-yl)piperazine-l-carboxylate (32): 1 H NMR (600 MHz, DMSO-d6) δ 9.50 (s, 1H), 8.39 (d, J = 2.6 Hz, 2H), 8.27 (s, 1H), 7.82 (dd, J = 8.9, 2.6 Hz, 1H), 7.60 - 7.54 (m, 1H), 7.28 (d, J = 2.0 Hz, 1H), 6.93 (d, J = 8.9 Hz, 1H), 5.87 (s, 1H), 5.33 (t, J = 4.9 Hz, 1H), 4.29 (d, J = 5.0 Hz, 2H), 3.53 (dd, J = 6.7, 3.9 Hz, 8H), 2.36 (t, J = 6.8 Hz, 2H), 2.21 (s, 3H), 2.15 (s, 2H), 2.12 (s, 3H), 2.01 (dd, J = 9.8, 5.7 Hz, 2H), 1.57 (qd, J = 14.0, 12.8, 6.4 Hz, 4H), 1.43 (s, 9H). 13C NMR (151 MHz, DMSO-d6) δ 171.8, 169.1, 166.7, 163.5, 158.5, 154.4, 150.2, 145.7, 143.3, 139.4, 137.9, 136.2, 135.0, 130.1, 128.9, 124.9, 124.0, 122.0, 108.0, 107.7, 79.5, 44.9, 36.0, 35.6, 35.4, 30.3, 29.5, 28.5, 27.0, 22.6, 19.4, 18.7, 14.9, 14.4. [M+Na]+calcd for C 36 H 47 N7NaO7712.3429, found 712.3426.

[0135] Example 24: Preparation of (R)-1-(sec-butyl)-N-(4,6-dimethyl-2-oxo-1,2- dihydro-pyridin-3-yl)methyl)-6-(6-(4-(9-(hydroxyamino)-9-oxo-nonanoyl)piperazin-1- yl)pyridin-3-yl)-3-methyl-1H-indole-4-carboxamide (33):

[0136] GSK126 (526.7 mg, 1 mmol) was coupled with 9-bromononanoic acid methyl ester (251.2 mg, 1 mmol) in the presence of cesium carbonate (325.8 mg, 1 mmol) to give 33a (592.0 mg 0.85 mmol). Then 33a (592.0 mg 0.85 mmol) was dissolved in NH2OK / CH3OH (10 mL) and stirred at room temperature for 4 hours. When the reaction was complete, the solvent was evaporated under reduced pressure to give a residue which was dissolved in water (15 mL). The mixture was neutralized by the addition of 1M HC1. The white precipitate was filtered and dried to give compound 33 (384.1 mg, 0.55 mmol) with a yield of 55% as a cream-colored solid. 1H NMR (500 MHz, DMSO-d6) δ 11.48 (s, 1H), 10.40 (s, 1H), 8.66 (s, 1H), 8.51 (d, J = 2.5 Hz, 1H), 8.14 (t, J = 5.1 Hz, 1H), 7.93 (d, J = 8.9 Hz, 1H), 7.73 (s, 1H), 7.25 (s, 1H), 7.18 (s, 1H), 6.93 (d, J = 8.8 Hz, 1H), 5.87 (s, 1H), 4.60 (q, J = 6.9 Hz, 1H), 4.35 (d, J = 5.1 Hz, 2H), 3.67 - 3.48 (m, 4H), 3.02 (s, 4H), 2.24 (s, 3H), 2.16 (s, 3H), 2.12 (s, 3H), 1.95 (t, J = 7.4 Hz, 2H), 1.85 - 1.76 (m, 2H), 1.56 - 1.45 (m, 4H), 1.41 (d, J = 6.7 Hz, 3H), 1.32 - 1.24 (m, 10H), 0.73 (t, J = 7.3 Hz, 3H). 13 C NMR (151 MHz, DMSO-d6) δ 169.0, 168.6, 162.9, 149.2, 145.3, 142.5, 137.6, 135.8, 130.5, 129.5, 124.3, 122.8, 121.6, 115.9, 109.5, 107.6, 107.3, 106.9, 51.4, 34.9, 32.1, 29.3, 28.9, 28.8, 28.6, 28.5, 28.5, 28.4, 24.9, 21.9, 20.7, 18.8, 18.0, 13.8, 11.5, 10.6. [M+Na]+calcd for C 34 H 38 N8NaO5, 661.2857, found 661.2861.

[0137] Example 25: Preparation of (R)-1-(sec-butyl)-N-(4,6-dimethyl-2-oxo-1,2- dihydropyridin-3-yl)methyl)-6-(6-(4-(5-(hydroxycarbamoyl)pyrimidin-2-yl)piperazin- 1-yl)pyridin-3-yl)-3-methyl-1H-indole-4-carboxamide (34):

[0138] Using the same procedure as in Example 24, but replacing 9-bromononanoic acid methyl ester in Example 24 with 2-chloropyrimidine-5-carboxylic acid methyl ester, compound 34 was prepared. 1H NMR (500 MHz, DMSO-d6) δ 11.41 (d, J = 102.3 Hz, 2H), 9.07 (s, 1H), 8.77 (s, 2H), 8.54 (s, 1H), 8.19 (d, J = 5.3 Hz, 1H), 7.97 (dd, J = 9.0, 2.6 Hz, 1H), 7.75 (s, 1H), 7.27 (s, 1H), 7.19 (s, 1H), 6.99 (d, J = 8.9 Hz, 1H), 5.88 (s, 1H), 4.61 (q, J = 7.0 Hz, 1H), 4.35 (d, J = 5.1 Hz, 2H), 3.95 (d, J = 5.7 Hz, 4H), 3.66 (t, J = 5.3 Hz, 4H), 3.38 (s, 3H), 2.24 (s, 3H), 2.16 (s, 3H), 2.12 (s, 3H), 1.86 - 1.73 (m, 2H), 1.40 (d, J = 6.6 Hz, 3H), 0.72 (t, J = 7.3 Hz, 3H). 13 C NMR (126 MHz, DMSO-d6) δ 169.2, 163.6, 161.9, 158.2, 157.7, 149.9, 146.0, 143.2, 138.2, 136.6, 131.1, 130.1, 126.8, 124.9, 123.4, 122.2, 116.5, 115.3, 110.2, 108.2, 107.9, 107.7, 52.0, 45.0, 43.6, 35.5, 30.0, 21.4, 19.5, 18.7, 12.2, 11.2. [M+Na]+calcd for C 36 H 41 N9NaO4, 686.3174, found 686.3176.

[0139] Example 26: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-3- (ethyl(tetrahydro-2H-pyran-4-yl)amino)-5-(6-(4-(9-(hydroxyamino)-9- oxononyl)piperazin-l-yl)pyridin-3-yl)-2-methylbenzamide (35):

[0140] Example 26: Preparation of N-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3-yl)methyl)-3- (ethyl(tetrahydro-2H-pyran-4-yl)amino)-5-(6-(4-(9-(hydroxyamino)-9- oxononyl)piperazin-l-yl)pyridin-3-yl)-2-methylbenzamide (35): 1H NMR (600 MHz, DMSO-d6) δ 11.50 (s, 1H), 10.45 (s, 1H), 8.77 - 8.66 (m, 1H), 8.39 (d, J = 2.5 Hz, 1H), 8.18 (t, J = 5.0 Hz, 1H), 7.81 (dd, J = 8.8, 2.6 Hz, 1H), 7.36 (d, J = 1.9 Hz, 1H), 7.17 (d, J = 1.8 Hz, 1H), 6.90 (d, J = 8.9 Hz, 1H), 5.87 (s, 1H), 4.29 (d, J = 4.9 Hz, 2H), 3.82 (dt, J = 11.7, 3.2 Hz, 2H), 3.54 (s, 4H), 3.24 (td, J = 11.7, 2.0 Hz, 2H), 3.07 (q, J = 7.0 Hz, 2H), 3.04 - 2.97 (m, 1H), 2.65 - 2.55 (m, 2H), 2.39 (p, J = 1.9 Hz, 2H), 2.21 (d, J = 8.3 Hz, 6H), 2.11 (s, 3H), 1.94 (t, J = 7.4 Hz, 2H), 1.69 - 1.59 (m, 2H), 1.50 (ddq, J = 25.2, 14.2, 7.2, 5.7 Hz, 6H), 1.39 - 1.12 (m, 9H), 0.82 (t, J = 7.0 Hz, 3H). 13 C NMR (151 MHz, DMSO-d6) δ 169.6, 169.6, 163.5, 158.6, 150.1, 149.4, 145.8, 143.2, 140.1, 136.2, 135.0, 132.3, 125.2, 122.6, 122.1, 120.4, 107.9, 107.5, 66.8, 58.2, 52.7, 44.9, 41.6, 35.3, 32.7, 30.8, 29.3, 29.2, 29.0, 27.3, 25.6, 19.4, 18.7, 15.0, 13.1. [M+Na]+calcd for C 41 H 60 N7O5, 730.4650, found 730.4650.

[0141] Example 27: Preparation of 2-(4-(5-(3-((4,6-dimethyl-2-oxo-l,2-dihydropyridin-3- yl)methyl)carbamoyl)-5-(ethyl(tetrahydro-2H-pyran-4-yl)amino)-4-methylphenyl)pyridin- 2-yl)piperazin-l-yl)-N-hydroxyprimidine-5-carboxamide (36):

[0142] Compound 36 was prepared by replacing 10b in Example 1 with 35a, replacing 4-bromobutyric acid methyl ester with 2-chloropyrimidine-5-carboxylic acid methyl ester, and following the same procedure as in Example 1. 1 H NMR (500 MHz, DMSO-d6) δ 11.19 (s, 1H), 8.74 (s, 2H), 8.43 (d, J = 2.5 Hz, 1H), 8.17 (t, J = 5.1 Hz, 1H), 7.89 - 7.82 (m, 1H), 7.37 (s, 1H), 7.20 (s, 1H), 6.96 (d, J = 8.9 Hz, 1H), 5.87 (s, 1H), 4.30 (d, J = 4.9 Hz, 2H), 3.98 - 3.90 (m, 4H), 3.86 - 3.79 (m, 2H), 3.66 (t, J = 5.2 Hz, 4H), 3.25 (s, 2H), 3.07 (t, J = 7.2 Hz, 2H), 3.04 - 2.98 (m, 1H), 2.22 (d, J = 8.6 Hz, 6H), 2.12 (s, 3H), 1.67 (d, J = 12.4 Hz, 2H), 1.52 (dt, J = 12.0, 5.8 Hz, 2H), 0.83 (t, J = 7.0 Hz, 3H). 13 C NMR (126 MHz, DMSO-d6) δ 169.6, 163.5, 161.9, 158.5, 157.6, 150.0, 149.5, 145.9, 143.3, 140.1, 136.3, 135.0, 132.4, 125.4, 122.6, 122.1, 120.4, 107.9, 107.6, 66.8, 58.3, 44.9, 43.6, 41.7, 35.4, 30.8, 19.4, 18.7, 15.0, 13.2. [M+Na]+calcd for C 37 H 45 N9NaO5, 718.3436, found 718.3434.

[0143] Example 28: In vitro HDAC1 and HDAC6 inhibitory activity assay

[0144] HDAC1 (#.AB101661) and HDAC6 (#.AB42632) enzymes were purchased from Abeam. Reaction mixtures contained 25 mM Tris (pH 8.0), 1 mM MgCl2, 0.1 mg / mL BSA, 137 mM NaCl, 2.7 mM KCl, HDACs (HDAC1, 10 ng per well; HDAC6, 20 ng per well) 40 microliters in a total volume. Test compounds (3-fold dilutions, 6 concentrations) were diluted in 10% DMSO, 5 μL of dilution was added, the purified recombinant HDAC was pre-incubated for 5 minutes at room temperature, then the substrate was added. Finally, the enzyme substrate was added, the plate was incubated for 30 minutes at 37°C, with a final volume of 50 μL. The reaction was quenched with 50 μL of HDAC Assay Developer for 30 minutes at room temperature. The assay was performed by quantifying the amount of fluorescent product in the solution after the enzymatic reaction. The fluorescence was then analyzed on a multimode plate reader (Envison 2015) with an excitation wavelength of 350-360 nm and an emission wavelength of 450-460 nm. IC 50 values were calculated using Prism GraphPad software using non-linear regression with normalized dose response fitting. All experiments were performed independently at least 3 times.

[0145] Table 1. HDAC inhibitory activity of compounds 10-36 a

[0146]

[0147]

[0148]

[0149]

[0150] a IC 50 values. Data are expressed as mean ± SD of three independent experiments.

[0151] We evaluated the HDAC1 and HDAC6 inhibitory activity of the target compounds (Table 1). Table 1 shows that the inhibitory activity of HDAC1 and HDAC6 generally increases with increasing linker length, but if the linker length is too long, the inhibitory activity of HDAC1 and HDAC6 decreases, for example, compounds 15 (HDAC1 IC 50 = 0.70 μM, HDAC6 IC 50 = 0.39 μM) and 16 (HDAC1 IC 50 = 6.02 μM, HDAC6 IC 50= 3.32 mM). We also evaluated the inhibitory activity of compounds with unsaturated chains as linkers (17-20), compound 20 showed the best potency, HDAC1 IC 50 = 0.12 mM and HDAC6 IC 50 = 0.70 mM. Changing the linker attachment position resulted in compounds 27-32, which showed moderate inhibitory activity against HDAC1 and HDAC6. Combining the kinetic results against the recombinant PRC2 complex and the antiproliferative results (Table 2 and Table 3), we concluded that the linkers of compounds 15 and 20 were more suitable to maintain potency against HDAC1 and HDAC6, therefore we attached the linkers to other pharmacophores of EZH2 (33-36), the pyrimidine linker showed better potency against HDAC1 and the alkane linker showed better potency against HDAC6.

[0152] Example 29: Investigation of the binding capacity of representative compounds against the recombinant PRC2 complex

[0153] Binding affinity was evaluated using a Biacore T200 instrument (GE Healthcare) at 25 °C. Briefly, compounds were diluted to the indicated concentrations with running buffer (10 mM HEPES supplemented with 5% DMSO). pH screening for immobilization of the recombinant PRC2 complex (Active Motif, Cat# 31387) was performed using 10 mM acetate buffers at pH values 4.0, 4.5, 5.0 and 5.5. pH 5.0 was found to be the most suitable condition for PRC2 immobilization. Approximately 12,000 response units (RU) of recombinant PRC2 were immobilized on a CM5 sensor chip by standard amine coupling procedures. Compounds were injected at concentrations of 0.5, 0.25, 0.125, 0.0625, 0.03125, 0.015625 and 0.0078125 mM at a flow rate of 30 pL / min into the PRC2 immobilized flow cell for 90 seconds and allowed to run buffer as dissociation blank for 90 seconds. Steady-state K D values were calculated using Biacore T200 evaluation software.

[0154] Table 2. Kinetic results of representative compounds against the recombinant PRC2 complex a

[0155]

[0156] a Values were calculated by Biacore T200 evaluation software. k a is the association rate constant, k d is the dissociation rate constant. K D = k d / k a

[0157] We selected a few representative compounds to evaluate the binding affinity of the PRC2 complex (Table 2). Compound 20 showed the best binding affinity (K D = 0.039 mM). After we changed the pharmacophore of EZH2, we obtained compounds 33-36, which did not show better binding affinity than 20.

[0158] Example 30: Anti-proliferative activity of compounds on different hematological and solid tumor cells

[0159] MV4-11, HL-60, SU-DHL-10, MDA-MB-231, HCT116 cells were seeded in 96-well plates and then treated with vehicle alone or with test compounds for 96 hours. 20 pL CCK8 solution (#C0005, TOPSCIENCE was added and the mixture was incubated at 37°C for 2-5 hours. Next, the absorbance of each well was measured at 450 nm using a plate reader (BioTek). Cell viability was calculated as follows: Cell viability (%) = [(As-Ab) / (Ac-Ab)]x100. (As represents the absorbance of experimental wells, Ab represents the absorbance of blank wells, and Ac represents the absorbance control well). In this study, the experiment was repeated at least 3 times, and IC 50 values were plotted by GraphPad Prism 6.

[0160] Table 3. Anti-proliferative test in vitro a

[0161]

[0162]

[0163] a IC50, data represent the mean value of three independent experiments ± SD.

[0164] b The combination index (CI) was calculated according to the Chou-Talalay method. CI > 1 indicates antagonism, CI = 1 indicates additive effect, and CI < 1 indicates synergy.

[0165] To further investigate the anti-proliferative effects of these dual-targeting inhibitors, we selected three hematological cancer cell lines: acute myeloid leukemia cells HL-60, MV4-11, diffuse large B-cell lymphoma cells SU-DHL-10; two solid tumor cell lines: colon cancer cells HCT-116, triple-negative breast cancer cells MDA-MB-231 (Table 3). The combination therapy of SAHA and GSK126, Tazemetostat, C24 showed synergistic effects in hematological cancer cells at a ratio of 1:1, respectively. In contrast, the combination therapy did not show obvious synergistic effects in treating solid tumor cells, which is consistent with our aim of using HDAC inhibitors in combination with EZH2 inhibitors to treat hematological malignancies. As shown in Table 3, the combination index (CI) in MV4-11 was the smallest among the three hematological malignancy cell lines, while the combination therapy of SAHA and GSK126 was the most effective (CI = 0.38). In general, our dual-targeting compounds showed the best inhibitory activity in MV4-11 cells among the three hematological cancer cell lines. The most effective compound 20 showed the best inhibitory activity (IC 50 = 0.17 μM) in MV4-11 cells, which was superior to the combination therapy of SAHA and GSK126 (IC 50 = 0.40 μM) in MV4-11 cells.

[0166] Example 31: Evaluation of anti-tumor activity of compounds in vivo.

[0167] Specific pathogen-free grade female BALB / c nude mice (6 weeks after birth) were purchased from Jiangsu Jizhuangkang Biotechnology Co., Ltd. MV4-11 cells (3 x 10 7 cells) were resuspended in PBS and inoculated subcutaneously in the right flank. When the average volume of tumor xenografts increased to 100 mm 3 , the mice were randomly divided into 6 groups (6 mice per group): control group, SAHA (100 mg / kg), GSK126 (100 mg / kg), SAHA + GSK126 (100 mg / kg + 100 mg / kg), 20 (50 mg / kg), 20 (100 mg / kg), intraperitoneal administration for 20 consecutive days. Body weight was monitored every 2 days. Tumor was measured every 2 days with a vernier caliper, and the tumor volume was calculated according to the following formula: volume (mm3) = length (mm) x width (mm) 2X 0.5. The compounds were dissolved in a mixture consisting of 5% DMSO, 5% PEG400, 5% castor oil and 85% (20% 2-hydroxypropyl-beta-cyclodextrin in saline). After treatment, mice were sacrificed, tumor tissues were dissected and weighed, and TGI was calculated according to the following formula: TGI = (average tumor weight of control group - average tumor weight of treatment group) / average tumor weight of control group. One-way ANOVA and Dunnett's multiple comparison test were used to determine statistical significance of tumor weight between treatment and control groups: (*) p < 0.05, and (**) p < 0.01.

[0168] A nude mouse MV4-11 xenograft model was established to evaluate the in vivo anti-tumor efficacy of compound 20. Mice were divided into six groups (six mice per group) according to drug dose and type: control, SAHA (100 mg / kg), GSK126 (100 mg / kg), SAHA + GSK126 (100 mg / kg + 100 mg / kg), 20 (50 mg / kg), 20 (100 mg / kg), intraperitoneal administration for 20 consecutive days. Tumor growth inhibition (TGI) was calculated at the end of treatment. Compared with the control group, the tumor growth of the five treatment groups was significantly inhibited by 52.7%, 68.5%, 76.2%, 65.7% and 82.0% (tumor growth inhibition rate), respectively. 20 reduced tumor growth in the MV4-11 xenograft model. Although the tumor inhibition effect in the 50 mg / kg group was not better than that of the positive drug SAHA, the 100 mg / kg treatment group was better than the positive drugs SAHA and GSK126, and slightly better than SAHA and GSK126 (82.0% vs. 76.2%). The final tumor weight and tumor growth curve are shown in Figures 1 and 2, respectively. Figure 1 A, B, and compared with each positive drug alone, showed improved anti-tumor efficacy in vivo.

Claims

1. A dual-target inhibitor targeting EZH2 / HDAC, characterized in that, Selected from the following compounds or their pharmaceutically acceptable salts: N-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-6-(6-(4-(5-(hydroxycarbamoyl)pyrimidin-2-yl)piperazin-1-yl)pyridin-3-yl)-1-isopropyl-1H-indazole-4-carboxamide.

2. The EZH2 / HDAC dual-target inhibitor according to claim 1, characterized in that, The pharmaceutically acceptable salt of the compound is selected from the acid addition salt formed by the compound of claim 1 with an acid selected from: hydrogen chloride, hydrogen bromide, sulfuric acid, carbonic acid, oxalic acid, citric acid, succinic acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, or ferulic acid.

3. The method for preparing the EZH2 / HDAC dual-target inhibitor according to claim 1, characterized in that, The reaction route is as follows: R b ： a. Reagents and reaction conditions: (a) 2-Iodopropane, K2CO3, CH3CN, reflux, 12h; (b) tert-butyl-4-(5-(4,4,5,5-tetramethyl-1,3,2-dioxoboronyl-2-yl)pyridin-2-yl)piperazine-1-carboxylate, PdCl2(dppf)·CH2Cl2, K2CO3, DMF, reflux, N2, 4h; (c) NaOH, MeOH / H2O, rt, 2h; (d) 3-(aminomethyl)-4,6-dimethylpyridin-2(1H)-one, HATU, DIPEA, DCM, rt, 2h; (e) CF3COOH, DCM, rt, 6h; (f) methyl 2-chloropyrimidin-5-carboxylate, Et3N, DMF, 90℃, 12h; (g) NH2OK, CH3OH, rt, 3-5h; For the linker R of the designed compound, the left side is connected to the nitrogen atom and the right side is connected to the carbonyl group.

4. A pharmaceutical composition comprising the compound of any one of claims 1 to 2 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

5. The pharmaceutical composition according to claim 4, characterized in that... The pharmaceutical composition is prepared into any one of the following dosage forms: tablets, capsules, powders, syrups, liquids, suspensions, lyophilized powder for injection, or injections.

6. Use of the compound of any one of claims 1 to 2 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating hematologic malignancies or other cancers.

7. The use of the compound of any one of claims 1 to 2 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for the treatment or prevention of EZH2 and HDAC-mediated diseases.

8. The application according to claim 7, characterized in that... The use of the compound of any one of claims 1 to 2 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for the treatment or prevention of EZH2 and HDAC-mediated hematologic malignancies.

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