A plk1 inhibitor and preparation method and application thereof

Abstract

The present application relates to the technical field of medicine, in particular to a kind of PLK1 inhibitor and its preparation method and application, the PLK1 inhibitor is compound with formula (I) or its pharmaceutically acceptable salt, the inhibitor structure novelty, the inhibition of PLK1 has effectiveness and specificity, has extensive and good proliferation inhibiting capacity to the tumor cells of multiple sources such as human lung cancer, liver cancer, bladder cancer and breast cancer.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and in particular to a PLK1 inhibitor, its preparation method, and its application. Background Technology

[0002] Polo-like kinases (PLKs) are a family of serine / tyrosine kinase proteins widely distributed in eukaryotic cells, playing important roles in multiple stages of the cell cycle. To date, the PLK protein family includes five members: PLK1, PLK2, PLK3, PLK4, and PLK5. These kinases have a conserved enzyme active domain (KD) at the N-terminus and a highly conserved non-catalytic active domain (PBD) at the C-terminus, primarily regulating interactions between proteins and substrates or regulatory factors. Among these, PLK1 has been the most extensively studied. Throughout the cell cycle, PLK1 participates in regulating various processes, such as checkpoint recovery, timing of mitotic entry, centrosome maturation and bipolar spindle assembly, stability of microtubule-kinetochordate attachment, and proper chromosome segregation in anaphase. PLK1 is typically expressed at low levels during interphase, peaking at G2 / M phase, and then gradually increasing. After mitosis, PLK1 is significantly degraded, resulting in a rapid decline in protein levels.

[0003] Cell cycle dysregulation is a prominent feature of many cancers. Upregulation of PLK1 expression in malignant cells leads to multiple defects in mitosis and cell division, as well as enhanced chromosomal instability, which typically manifests as high tumor grade and poor patient prognosis. PLK1 promotes cancer progression not only through cell cycle dysregulation but also through metabolic reprogramming, such as increasing the flow of the pentose phosphate pathway (PPP) and driving glucose into macromolecular synthesis pathways. Furthermore, downregulation of PLK1 expression generally results in reduced proliferation of various cancer cells. Therefore, PLK1 is considered a potent proto-oncogene and a potential drug target for cancer therapy.

[0004] To date, at least 10 PLK1 inhibitors (PLK1i) have entered clinical trials, but none have been approved for marketing. For example, PLK1 KD inhibitors (such as BI2536, GSK461364, NMS-128693, and Volasertib) face challenges due to their KD having similar catalytic mechanisms, common substrates (ATP), and similar protein folding characteristics to other protein kinases, making broad selectivity a major issue. Furthermore, studies have reported that PLK2 and PLK3 are effective tumor inhibitors, but the broad selectivity of PLK1i may lead to reduced efficacy and varying degrees of dose-limiting toxicity, restricting their application.

[0005] To avoid the broad selectivity of PLK1 inhibitors, many researchers have focused on the unique C-terminal polo-box domain (PBD) of PLK1. PLHSpT (phosphopeptide) and its derivatives have been identified as PLK1 PBD-specific inhibitors, primarily inhibiting PLK1 by specifically binding to the PBD and blocking its activated substrate recruitment and subcellular localization functions. However, these peptide inhibitors have serious drawbacks, such as poor cell permeability and protein hydrolysis instability, resulting in low bioavailability. To overcome this limitation, some small molecule compounds such as thymosin, poloxipan, and thymosin have been identified as PLK1-PBD inhibitors. However, the structural diversity and moderate selectivity of PLK1-PBD inhibitors greatly restrict structure-based design. Therefore, developing effective therapeutic PLK1 inhibitors remains an urgent need. Thus, developing PLK1 inhibitors with novel structures and better activity is of great significance. Summary of the Invention

[0006] The purpose of this invention is to provide a PLK1 inhibitor, its preparation method, and its application, addressing the aforementioned problems. This inhibitor has a novel structure, exhibits effectiveness and specificity in inhibiting PLK1, and demonstrates broad and effective inhibitory capabilities against the proliferation of tumor cells from various sources, including human lung cancer, liver cancer, bladder cancer, and breast cancer.

[0007] This invention provides a PLK1 inhibitor, wherein the PLK1 inhibitor is a compound having the structure of formula (I) or a pharmaceutically acceptable salt thereof.

[0008]

[0009] Among them, R1 is selected from alkyl, R2 is selected from -H, methyl, methoxy, fluorine, or trifluoromethyl.

[0010] The present invention also provides a method for preparing the above-mentioned compound, comprising the following steps:

[0011] (1) Under nitrogen protection, potassium salt of 4-sulfon-1,8-naphthalenedicarboxylic anhydride reacts with the corresponding amine in the first solvent to give intermediate product 1;

[0012] The structural formula of the potassium salt of 4-sulfonyl-1,8-naphthalenedicarboxylic anhydride is:

[0013]

[0014] The structural formula of intermediate product 1 is:

[0015]

[0016] Where R1 represents the base, The corresponding amines are N,N-dimethylethylenediamine, n-butylamine, 1-(2-aminoethyl)pyrrolidine, 1-(2-aminoethyl)piperidine, N-(2-aminoethyl)morpholine, and 3-dimethylamino-1-propylamine;

[0017] (2) Under nitrogen protection, intermediate 1 reacts with thionyl chloride in a second solvent as a catalyst to obtain intermediate 2; the structural formula of intermediate 2 is:

[0018]

[0019] (3) Intermediate product 2 and the corresponding raw material are dissolved in dichloromethane, and triethylamine is added under nitrogen protection to react and obtain the PLK1 inhibitor. The corresponding raw material is selected according to the different substituents and the different substituent positions. The general formula of the corresponding raw material is R2-benzylamine, wherein R2 is -H, methyl, methoxy, fluorine or trifluoromethyl.

[0020] In the above preparation method, in step (1), the molar ratio of the potassium 4-sulfonyl-1,8-naphthalenedicarboxylic anhydride to the corresponding amine is 1:2, and the solid-liquid ratio of the potassium 4-sulfonyl-1,8-naphthalenedicarboxylic anhydride to the first solvent is 1:40.

[0021] In the above preparation method, in step (2), the mass-volume ratio of intermediate product 1 to thionyl chloride is 1:15, and the solid-liquid ratio of intermediate product 1 to the second solvent is 1:30.

[0022] In the above preparation method, in step (3), the molar ratio of the intermediate product 2 to the corresponding raw material is 1:1.5-3, and the solid-liquid ratio of the intermediate product 2 to dichloromethane is 1:20.

[0023] In the above preparation method, the first solvent and the second solvent are selected from one or more combinations of ethanol, methanol, dichloromethane and N,N-dimethylformamide, and the reaction temperature in steps (1) and (2) is from 40°C to the reflux temperature of the first solvent.

[0024] Preferably, the preparation method further includes step (4), which is a method for purifying the product obtained in step (3): the product obtained in step (3) is purified by silica gel column chromatography, eluted with an eluent composed of dichloromethane and methanol in a volume ratio of 20-50:1, and recrystallized with dichloromethane and methanol in a volume ratio of 5:1 to obtain the purified target compound.

[0025] The present invention also provides the application of the PLK1 inhibitor in the preparation of antitumor drugs.

[0026] The present invention also provides the use of the PLK1 inhibitor in the preparation of medicaments for the prevention or treatment of non-small cell lung cancer, liver cancer, ovarian cancer or bladder cancer.

[0027] The present invention also provides a pharmaceutical composition comprising a therapeutically effective dose of the compound or a pharmaceutically acceptable salt thereof.

[0028] In summary, due to the adoption of the above technical solution, the present invention has the following beneficial effects:

[0029] This invention demonstrates effective and specific inhibition of PLK1. Surface plasmon resonance experiments revealed that the PLK1 inhibitor B2 synthesized in this invention exhibits better affinity for PLK1 compared to Volasertib. Further experiments showed that other compounds prepared in this invention also possess good affinity. In vitro thermodynamic migration experiments confirmed that B2 binds to the PLK1 protein, enhancing its thermal stability. Other compounds prepared in this invention also enhanced the thermal stability of the PLK1 protein. Furthermore, in vitro antitumor activity experiments showed that this invention exhibits broad and significant inhibitory effects on the proliferation of tumor cells from various sources, including human lung cancer, liver cancer, bladder cancer, and breast cancer, with IC50 values ​​ranging from 1 to 10 μM. Attached Figure Description

[0030] Figure 1 This is a graph showing the in vitro binding ability of the final product obtained in Example 1 of this invention with PLK1.

[0031] Figure 3 This is a high-resolution mass spectrum of the final product obtained in Example 1 of the present invention.

[0032] Figure 2 This is a high-performance liquid chromatogram of the final product obtained in Example 1 of the present invention. Detailed Implementation

[0033] To more clearly illustrate the present invention, the following specific embodiments will be used to further explain the invention.

[0034] This invention provides a PLK1 inhibitor, wherein the PLK1 inhibitor is a compound having the structure of formula (I) or a pharmaceutically acceptable salt thereof.

[0035]

[0036] Among them, R1 is selected from alkyl, R2 is selected from -H, methyl, methoxy, fluorine, or trifluoromethyl.

[0037] Preferably, the compound is: N-2-butyl-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide, N-2-butyl-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide, N-2-butyl-6-sulfonamide-(trifluoromethyl)benzyl)-1,8-naphthalenediamide, N-2-butyl-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide, N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-benzyl-1,8-naphthalenediamide, N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide, N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide Amine-(4-methoxybenzyl)-1,8-naphthalenediformimide, N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediformimide, N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediformimide, N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(2-fluorobenzyl)-1,8-naphthalenediformimide, N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(3-fluorobenzyl)-1,8-naphthalenediformimide, N-2-(2-(pyrrolidinyl)ethyl)-6-sulfonamide-benzyl ... (alkyl)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide, N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide, N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide, N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide, N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-benzyl-1,8-naphthalenediamide, N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide, N-2- (2-(piperidinyl)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide, N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide, N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide, N-2-(2-(morpholinyl)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide, N-2-(2-(morpholinyl)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide, N-2-(2-(morpholinyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-Naphthalenedicarboximide, N-2-(2-(morpholinyl)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenedicarboximide, N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-benzyl-1,8-naphthalenedicarboximide, N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenedicarboximide, N-2 -(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide, N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide, or N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide.

[0038] The present invention also provides a method for preparing the above-mentioned compound, comprising the following steps:

[0039] (1) Under nitrogen protection, potassium salt of 4-sulfon-1,8-naphthalenedicarboxylic anhydride reacts with the corresponding amine in the first solvent to give intermediate product 1;

[0040] The structural formula of the potassium salt of 4-sulfonyl-1,8-naphthalenedicarboxylic anhydride is:

[0041]

[0042] The structural formula of intermediate product 1 is:

[0043]

[0044] Where R1 represents the base, The corresponding amines are N,N-dimethylethylenediamine, n-butylamine, 1-(2-aminoethyl)pyrrolidine, 1-(2-aminoethyl)piperidine, N-(2-aminoethyl)morpholine, and 3-dimethylamino-1-propylamine;

[0045] (2) Under nitrogen protection, intermediate 1 reacts with thionyl chloride in a second solvent as a catalyst to obtain intermediate 2; the structural formula of intermediate 2 is:

[0046]

[0047] (3) Intermediate product 2 and the corresponding raw material are dissolved in dichloromethane, and triethylamine is added under nitrogen protection to react and obtain the PLK1 inhibitor. The corresponding raw material is selected according to the different substituents and the different substituent positions. The general formula of the corresponding raw material is R2-benzylamine, wherein R2 is -H, methyl, methoxy, fluorine or trifluoromethyl.

[0048] In the above preparation method, in step (1), the molar ratio of the potassium 4-sulfonyl-1,8-naphthalenedicarboxylic anhydride to the corresponding amine is 1:2, and the solid-liquid ratio of the potassium 4-sulfonyl-1,8-naphthalenedicarboxylic anhydride to the first solvent is 1:40.

[0049] In the above preparation method, in step (2), the mass-volume ratio of intermediate product 1 to thionyl chloride is 1:15, and the solid-liquid ratio of intermediate product 1 to the second solvent is 1:30.

[0050] In the above preparation method, in step (3), the molar ratio of the intermediate product 2 to the corresponding raw material is 1:1.5-3, and the solid-liquid ratio of the intermediate product 2 to dichloromethane is 1:20.

[0051] In the above preparation method, the first solvent and the second solvent are selected from one or more combinations of ethanol, methanol, dichloromethane and N,N-dimethylformamide, and the reaction temperature in steps (1) and (2) is from 40°C to the reflux temperature of the first solvent.

[0052] In the above preparation method, the general formula of the corresponding raw material is R2-benzylamine, wherein R2 is -H, methyl, methoxy, fluorine or trifluoromethyl.

[0053] Preferably, the preparation method further includes step (4), which is a method for purifying the product of step (3): the product of step (3) is purified by silica gel column chromatography, eluted with an eluent consisting of dichloromethane and methanol in a volume ratio of 20-50:1, and recrystallized with dichloromethane and methanol in a volume ratio of 5:1 to obtain the purified target compound.

[0054] Example 1

[0055] This embodiment prepares a PLK1 inhibitor: N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (denoted as B2), specifically including the following steps:

[0056] (1) Under nitrogen protection, 1.58 g of 4-sulfono-1,8-naphthalenedicarboxylic anhydride potassium salt (5 mmol, 1.0 eq) and 0.88 g of N,N-dimethylethylenediamine (2.0 eq) were weighed, 40 mL of ethanol was added, and the mixture was refluxed for 5 hours. The reaction was monitored by TLC to determine if it was complete. After the reaction was complete, the mixture was cooled to room temperature, filtered, washed with ethanol, and dried under vacuum to obtain 1.7 g of white solid, which is intermediate product 1, with a yield of 88%.

[0057] (2) Under nitrogen protection, 15 mL of thionyl chloride was added to intermediate product 1 (0.38 g, 1.0 mmol, 1.0 eq) and 30 mL of DMF. The mixture was refluxed for 10 h. The reaction was monitored by TLC to determine if it was complete. After the reaction was complete, the mixture was cooled to room temperature and the solvent was removed by vacuum to obtain a white solid, which is intermediate product 2. No purification was required, and it was used directly for the next step of the reaction.

[0058] (3) The intermediate product 2 (0.67 mmol) obtained in step (2) and p-fluorobenzylamine (0.25 g, 2.0 mmol, 2.0 eq) were dissolved in 20 mL of dry dichloromethane. Under nitrogen protection, 1 mL of triethylamine was added. The reaction mixture was stirred for 5 min, and then a solution of molecule 2 in dichloromethane (20 mL) was added dropwise at 0 °C. The reaction was carried out at room temperature for 5 h. The mixture was filtered, and the filtrate was washed twice with saturated brine and then dried with anhydrous Na2SO4 to obtain the crude target compound. The crude target compound was purified by silica gel column chromatography (V MeOH :V DCM Recrystallization from dichloromethane-methanol (5:1) yielded 0.2 g of yellow needle-like crystals, which is the target product N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide, with a yield of 44%.

[0059] The high-resolution mass spectrum of the final product obtained in this embodiment is as follows: Figure 2 As shown, its high-performance liquid chromatogram is as follows: Figure 3 As shown; using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0060] N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (B2): yellow solid, yield 44%, mp 171-173℃; HPLC (MeOH:H2O=45:55, 1% TFA, 0.5mL / min), t R =6.435 min, 99.02%; 1H NMR (400MHz, DMSO-d6) δ9.01(d,J=8.6Hz,1H),8.90(s,1H),8.58(d,J=7.1Hz,1H),8.52(d,J=7.6Hz,1H),8.34(d,J=7.6Hz,1H),8.01(dd,J= 8.8,7.4Hz,1H),7.08(dd,J=8.5,5.6Hz,2H),6.88(t,J=8.8Hz,2H),4.16(t,J=6.9Hz,2H),4.08(s,2H),2.52(d,J=7.0Hz,2H),2.22(s,6H). 13 C NMR (100MHz, DMSO-d6) δ163.5,162.9,161.6(d,J=241.7Hz),141.7,133.7(d,J=3.0Hz),131.6,131.3,130.0,129 .9,129.2(d,J=17.7Hz),128.7,126.6,126.3,123.2,115.0(d,J=21.3Hz),56.8,45.9,45.7,38.3.HRMS(ESI)m / z calcd for C 23 H 22 N3O4FS[MH] - 454.1242, found 454.1238.

[0061] Example 2: Preparation of a PLK1 inhibitor: N-2-butyl-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (denoted as A2)

[0062] The difference between this embodiment and embodiment 1 lies in steps (1) and (3), while the other steps are the same as in embodiment 1;

[0063] The amine used in step (1) of this embodiment is n-butylamine;

[0064] In this embodiment, step (3) is as follows: The intermediate product 2 (1.0 mmol) obtained in step (2) and p-fluorobenzylamine (0.25 g, 2.0 mmol, 2.0 eq) are dissolved in 20 mL of dry dichloromethane. Under nitrogen protection, 1 mL of triethylamine is added. After stirring the reaction mixture for 5 min, a solution of molecule 2 in dichloromethane (20 mL) is added dropwise at 0 °C. The reaction is carried out at room temperature for 5 h. The mixture is filtered, and the filtrate is washed twice with saturated saline solution, then dried with anhydrous Na2SO4 to obtain the crude target compound. The crude target compound is purified by silica gel column chromatography (V... MeOH :V DCM=1:20), dichloromethane-methanol (5:1) recrystallization yields the target product N-2-butyl-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide.

[0065] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0066] N-2-Butyl-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (A2): White powdery solid, yield 54%, mp 159-160℃; 1 H NMR (400MHz, DMSO-d6) δ8.99(d,J=8.6Hz,1H),8.89(s,1H),8.56(d,J=7.2Hz,1H),8.50(d,J=7.7Hz,1H),8.32(d,J=7.7Hz,1H),8.03-7.97 (m,1H),7.05(dd,J=8.4,5.6Hz,2H),6.84(t,J=8.9Hz,2H),4.10-4.01(m,4H),1.68-1.56(m,2H),1.41-1.30(m,2H),0.93(t,J=7.3Hz,3H). 13 C NMR (100MHz, DMSO-d6) δ163.5, 163.0, 161.5 (d, J = 241.7Hz), 141.6, 133.6 (d, J = 3.1Hz), 131.4 (d, J = 20.7Hz), 129.9 (d, J = 8.2H z),129.8,129.2(d,J=21.5Hz),128.7,126.6,126.3,123.2,115.0(d,J=21.3Hz),45.7,40.0,30.0,20.2,14.2.HRMS(ESI)m / z calcd for C 23 H 21 N₂O₄FS[MH] - 439.1133, found 439.1128.

[0067] Example 3: Preparation of a PLK1 inhibitor: N-2-butyl-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (denoted as A3)

[0068] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is n-butylamine, and the corresponding raw material used in step (3) is 4-methoxybenzylamine. The other steps are the same as those in embodiment 1.

[0069] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0070] N-2-Butyl-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (A3): Pale yellow powder solid, yield 40%, mp 141-142℃; 1 H NMR (400MHz, DMSO-d6) δ8.96(d,J=8.4Hz,1H),8.78(s,1H),8.54(d,J=6.9Hz,1H),8.46(d,J=7.7Hz,1H),8.28(d,J=7.7Hz,1H),7.97(dd,J=8.7,7.3 Hz,1H),6.85(d,J=8.6,Hz,2H),6.47(d,J=8.6Hz,2H),4.11-3.95(m,4H), 3.55(s,3H),1.69-1.56(m,2H),1.42-1.31(m,2H),0.93(t,J=7.3Hz,3H). 13 C NMR(100MHz,DMSO-d6)δ163.5,163.0,158.5,141.9,131.4,131.4,129.8,129.3,129.3,129.0,1 28.9,128.6,126.6,126.2,123.2,113.4,55.3,46.1,40.1,30.0,20.2,14.2.HRMS(ESI)m / zcalcd for C 24 H 24 N₂O₅S[M+Na] + 475.1299, found 475.1286.

[0071] Example 4: Preparation of a PLK1 inhibitor: N-2-butyl-6-sulfonamide (trifluoromethyl)benzyl)-1,8-naphthalenediamide (denoted as A4)

[0072] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is n-butylamine, and the corresponding raw material used in step (3) is 4-(trifluoromethyl)benzylamine. The other steps are the same as in embodiment 1.

[0073] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0074] N-2-Butyl-6-sulfonamide (trifluoromethyl)benzyl)-1,8-naphthalenediamide (A4): White powdery solid, yield 51%, mp 174-175℃; 1 H NMR (400MHz, DMSO-d6) δ9.04(s,1H),8.94(d,J=8.5Hz,1H),8.54(d,J=7.1Hz,1H),8.43(d,J=7.6Hz,1H),8.29(d,J=7.6Hz,1H),7.98(dd,J=8.4,7.6H z,1H),7.21(d,J=8.2Hz,2H),7.15(d,J=8.1Hz,2H),4.20(s,2H),4.01(t, J=7.4Hz,2H),1.67-1.54(m,2H),1.43-1.29(m,2H),0.93(t,J=7.3Hz,3H). 13 C NMR (100MHz, DMSO-d6) δ163.4,162.8,141.1(d,J=1.1Hz),141.5,131.4,131.2,129.7,129.5,129.0,128.8,128.6,128.5,127.8(d,J=32.4H z),126.5,126.3,124.7(d,J=3.7Hz),161.5(d,J=241.7Hz),124.7,124.4(d,J=270.1Hz),123.1,45.9,40.1,30.0,20.2,14.2.HRMS(ESI)m / z calcd for C 24 H 21 N₂O₄F₃S[MH] - 489.1101, found 489.1104.

[0075] Example 5: Preparation of a PLK1 inhibitor: Preparation of N-2-butyl-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (denoted as A5)

[0076] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is n-butylamine, and the corresponding raw material used in step (3) is 4-methylbenzylamine. The other steps are the same as those in embodiment 1.

[0077] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0078] N-2-Butyl-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (A5): White needle-like crystals, yield 39%, mp 170-171℃; 1 H NMR (400MHz, DMSO-d6) 1 H NMR (400MHz, DMSO-d6) δ8.98(d,J=8.4Hz,1H),8.83(s,1H),8.55(d,J=7.2Hz,1H),8.46(d,J=7.7Hz,1H),8.28(d,J=7.7Hz,1H),7.98(dd,J=8.3,7. 6Hz,1H),6.84(d,J=7.9Hz,2H),6.75(d,J=7.8Hz,2H),4.08-3.99(m,4H), 2.07(s,3H),1.68-1.57(m,2H),1.43-1.30(m,2H),0.93(t,J=7.3Hz,3H). 13 C NMR(100MHz,DMSO-d6)δ163.6,163.0,141.8,136.5,134.1,131.4,131.4,129.8,129.3,12 9.0,128.7,127.9,126.6,126.2,123.2,46.3,40.0,30.1,20.9,20.3,14.2.HRMS(ESI)m / z calcd for C 24 H 24 N₂O₄S[MH] - 435.1384, found 435.1387.

[0079] Example 6: Preparation of a PLK1 inhibitor: N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-benzyl-1,8-naphthalenediamide (denoted as B1)

[0080] The difference between this embodiment and embodiment 1 is that the corresponding raw material used in step (3) of this embodiment is benzylamine, while the other steps are the same as in embodiment 1.

[0081] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0082] N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-benzyl-1,8-naphthalenediamine (B1): white powdery solid, yield 48%, mp 167-168℃; 1 H NMR (400MHz, DMSO-d6) δ9.03(dd,J=8.7,0.9Hz,1H),8.89(s,1H),8.57(dd,J=7.3,1.1Hz,1H),8.51(d,J=7.7Hz,1H),8.34(d,J= 7.7Hz,1H),8.01(dd,J=8.7,7.3Hz,1H),7.15-6.98(m,5H),4.15(t,J=6.8Hz,2H),4.08(s,2H),2.57-2.47(m,4H),2.21(s,6H). 13 C NMR(100MHz,DMSO-d6)δ162.9,162.4,141.1,136.8,130.9,130.8,129.3,128.6,128.5,12 8.1,127.8,127.3,126.8,126.1,125.7,122.6,56.2,45.9,45.2,37.6.HRMS(ESI)m / zcalcd for C 23 H 23 N3O4S[M+H] + 438.1483, found 438.1467.

[0083] Example 7: Preparation of a PLK1 inhibitor: N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (denoted as B3)

[0084] The difference between this embodiment and embodiment 1 is that the corresponding raw material used in step (3) of this embodiment is 4-methoxybenzylamine, while the other steps are the same as in embodiment 1.

[0085] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0086] N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (B3): Yellow powder solid, yield 44%, mp 141-142℃; 1H NMR (400MHz, DMSO-d6) δ8.99(d,J=8.5Hz,1H),8.79(s,1H),8.56(d,J=7.1Hz,1H),8.48(d,J=7.7Hz,1H),8.29(d,J=7.7Hz,1H),7.99(dd,J=8.7 ,7.3Hz,1H),6.88(d,J=8.6Hz,2H),6.50(d,J=8.6Hz,2H),4.15(t,J=6.9Hz,2H),4.01(s,2H),3.57(s,3H),2.52(d,J=7.1Hz,2H),2.22(s,6H). 13 C NMR(100MHz,DMSO-d6)δ163.5,163.0,158.6,141.9,131.5,131.4,129.9,129.3,129.1 ,129.0,128.7,126.6,126.2,123.1,113.5,56.8,55.4,46.1,45.9,38.3.HRMS(ESI)m / z calcd forC 24 H 25 N3O5S[M+H] + 468.1588, found 468.1564.

[0087] Example 8: Preparation of a PLK1 inhibitor: N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (denoted as B4)

[0088] The difference between this embodiment and embodiment 1 is that the corresponding raw material used in step (3) of this embodiment is 4-(trifluoromethyl)benzylamine, while the other steps are the same as in embodiment 1.

[0089] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0090] N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (B4): white powdery solid, yield 51%, mp 208-209℃; 1H NMR (400MHz, DMSO-d6) δ9.04(s,1H),8.96(d,J=8.6Hz,1H),8.55(d,J=7.2Hz,1H),8.46(d,J=7.7Hz,1H),8.30(d,J=7.7Hz,1H),8.0 5-7.95(m,1H),7.27(d,J=8.1Hz,2H),7.19(d,J=8.0Hz,2H),4.20(s,2H),4.14(t,J=7.0Hz,2H),2.56(d,J=1.2Hz,2H),2.22(s,6H). 13 C NMR (100MHz, DMSO-d6) δ163.4, 162.8, 142.0, 141.6, 131.5, 129.9 (d, J = 274.1Hz), 129.7 (d, J = 28.6Hz), 12 9.1,128.6,128.0,127.7,126.4(d,J=29.9Hz),124.9,124.9,123.1,56.8,45.9,45.8,38.2.HRMS(ESI)m / z calcd for C 24 H 22 N3O4F3S[M+H] + 506.1356, found 506.1335.

[0091] Example 9: Preparation of a PLK1 inhibitor: N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (denoted as B5)

[0092] The difference between this embodiment and embodiment 1 is that the corresponding raw material used in step (3) of this embodiment is 4-methylbenzylamine, while the other steps are the same as in embodiment 1.

[0093] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0094] N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (B5): yellow square crystals, yield 35%, mp 151-152℃; 1H NMR (400MHz, CDCl3-d1) δ8.99(d,J=8.3Hz,1H),8.83(s,1H),8.56(d,J=6.9Hz,1H),8.48(d,J=7.7Hz,1H),8.29(d,J=7.7Hz,1H),7.99(dd,J=8. 5,7.5Hz,1H),6.85(d,J=8.0Hz,2H),6.76(d,J=7.9Hz,2H),4.03(s,2H) ,3.17(t,J=5.7Hz,2H),2.52(d,J=6.9Hz,2H),2.21(s,6H),2.08(s,3H). 13 C NMR (100MHz, CDCl3-d1) δ163.59,163.02,140.74,137.97,132.36,131.80,130.72,129.57,129.32,129 .24,129.02,128.79,127.82,126.91,126.64,123.27,56.90,47.30,45.76,38.44,20.99.HRMS(ESI)m / z calcd for C 24 H 25 N3O4S[M+H] + 452.1639, found 452.1618.

[0095] Example 10: Preparation of a PLK1 inhibitor: N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(2-fluorobenzyl)-1,8-naphthalenediamide (denoted as B6)

[0096] The difference between this embodiment and embodiment 1 is that the corresponding raw material used in step (3) of this embodiment is 2-fluorobenzylamine, while the other steps are the same as those in embodiment 1.

[0097] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0098] N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(2-fluorobenzyl)-1,8-naphthalenediamide (B6): white powdery solid, yield 58%, mp 172-173℃; 1H NMR (400MHz, DMSO-d6) δ9.01(d,J=8.6Hz,1H),8.57(d,J=6.8Hz,1H),8.50(d,J=7.6Hz,1H),8.33(d,J=7.6Hz,1H),8.00(dd,J=8.5,7.5Hz ,1H),7.17-7.13(m,1H),7.10-7.06(m,1H),6.86(t,J=8.2Hz,2H),4.17(d,J=6.8Hz,2H),4.14(s,2H),2.55(t,J=6.8Hz,2H),2.23(s,6H). 13 C NMR (100MHz, DMSO-d6) δ172.5,163.5,163.0,160.0(d,J=244.2Hz),141.4,131.5,131.3,130.6(d,J=3.9Hz),129.8,129.8,129.7,129.1(d,J= 19.8Hz),128.7,126.5(d,J=24.9Hz),124.4(d,J=3.4Hz),124.2,124.1,123.2,115.2(d,J=21.1Hz),56.78,45.82,38.20,21.56.HRMS(ESI)m / z calcd for C 23 H 22 N3O4FS[M+H] + 456.1388, found 456.1385.

[0099] Example 11: Preparation of a PLK1 inhibitor: N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(3-fluorobenzyl)-1,8-naphthalenediamide (denoted as B7)

[0100] The difference between this embodiment and embodiment 1 is that the corresponding raw material used in step (3) of this embodiment is 3-fluorobenzylamine, while the other steps are the same as in embodiment 1.

[0101] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0102] N-(2-N,N-dimethylamino)ethylamino-6-sulfonamide-(3-fluorobenzyl)-1,8-naphthalenediamide (B7): white powdery solid, yield 50%, mp 144-145℃; 1H NMR (400MHz, DMSO-d6) δ9.01(dd,J=8.8,1.1Hz,2H),8.57(dd,J=7.4,1.0Hz,1H),8.51(d,J=7.6Hz,1H),8.34(d,J=7.6Hz,1H),8.02( dd,J=8.8,7.4Hz,1H),7.12-7.06(m,1H),6.91-6.80(m,3H),4.17(t,J=6.8Hz,2H),4.12(s,2H),2.56(t,J=6.8Hz,2H),2.24(s,6H). 13 C NMR (100MHz, DMSO-d6) δ172.5,163.5,162.92,162.2(d,J=242.1Hz),141.6,140.4,131.6,131.3,130.3,130.2,129.9,129.2(d,J=24.2Hz ),128.7,126.5(d,J=24.0Hz),123.9(d,J=2.62Hz),123.2,114.6,114.4,114.1(d,J=20.9Hz),56.81,45.86,38.25,21.59.HRMS(ESI)m / z calcd for C 23 H 22 N3O4FS[MH] - 454.1242, found 454.1240.

[0103] Example 12: Preparation of a PLK1 inhibitor: N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-benzyl-1,8-naphthalenediamide (denoted as C1)

[0104] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)pyrrolidine, and the corresponding raw material used in step (3) is benzylamine. The other steps are the same as those in embodiment 1.

[0105] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0106] N-2-(2-(pyrrolidinyl)ethyl)-6-sulfonamide-benzyl-1,8-naphthalenediamide (C1): white powdery solid, yield 49%, mp 150-151℃; 1H NMR (400MHz, DMSO-d6) δ9.03(dd,J=8.8,1.1Hz,1H),8.90(s,1H),8.57(dd,J=7.4,1.1Hz,1H),8.51(d,J=7.6Hz,1H),8.34(d,J=7.6Hz,1H),8.01( dd,J=8.6,7.4Hz,1H),7.07(d,J=3.2Hz,5H),4.17(t,J=7.0Hz,2H),4.09 (s,2H),2.68(t,J=7.0Hz,2H),2.61-2.45(m,4H),1.67(t,J=3.3Hz,4H). 13 C NMR(100MHz,DMSO-d6)δ163.5,163.0,141.8,137.5,131.5,131.4,129.9,129.2,129.1,12 8.7,128.4,127.9,127.4,126.7,126.3,123.2,54.2,53.4,49.1,46.5,23.7.HRMS(ESI)m / z calcd forC 25 H 25 N3O4S[M+H] + 464.1639, found 464.1628.

[0107] Example 13: Preparation of a PLK1 inhibitor: N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (denoted as C2)

[0108] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)pyrrolidine, and the corresponding raw material used in step (3) is 4-fluorobenzylamine. The other steps are the same as in Example 1.

[0109] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0110] N-2-(2-(pyrrolidinyl)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (C2): White powdery solid, yield 55%, mp 108-109℃; 1H NMR(400MHz, DMSO-d6)δ9.00(dd,J=8.8,1.1Hz,1H),8.91(s,1H),8.58(dd,J=7 .4,1.1Hz,1H),8.52(d,J=7.6Hz,1H),8.33(d,J=7.6Hz,1H),8.01(dd,J=8.8,7 .4Hz,1H),7.08(dd,J=8.6,5.6Hz,2H),6.91-6.83(m,2H),4.18(t,J=7.1Hz,2H ),4.08(s,2H),2.68(t,J=7.1Hz,2H),2.59-2.51(m,4H),1.67(t,J=3.1Hz,4H). 13 C NMR (100MHz, DMSO-d6) δ163.4,162.9,161.6(d,J=241.8Hz),141.7,133.7(d,J=2.9Hz),131.6,131.3,130.0(d,J=8.2Hz),12 9.9,129.2(d,J=17.1Hz),128.7,126.5(d,J=3.0Hz),123.2,115.0(d,J=21.2Hz),54.2,53.3,49.1,45.7,23.7.HRMS(ESI)m / z calcd for C 25 H 24 N3O4FS[M+H] + 482.1545, found 482.1541.

[0111] Example 14: Preparation of a PLK1 inhibitor: N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (denoted as C3)

[0112] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)pyrrolidine, and the corresponding raw material used in step (3) is 4-methoxybenzylamine. The other steps are the same as in Example 1.

[0113] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0114] N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (C3): yellow flaky crystals, yield 38%, mp 197-198℃;1 H NMR(400MHz,DMSO-d6)δ8.98(dd,J=8.8,1.1Hz,1H),8.79(s,1H),8.56(dd,J =7.4,1.1Hz,1H),8.48(d,J=7.6Hz,1H),8.30(d,J=7.6Hz,1H),7.99(dd,J=8. 7,7.3Hz,1H),6.91-6.85(m,2H),6.54-6.47(m,2H),4.22-4.14(m,2H),4.01( s,2H),3.57(s,3H),2.72-2.64(m,2H),2.58-2.51(m,4H),1.69-1.67(m,4H). 13 C NMR(100MHz,DMSO-d6)δ163.5,162.9,158.6,141.9,131.5,131.4,129.9,129.3,129.1,12 9.0,128.7,126.7,126.2,123.1,113.5,55.4,54.2,53.4,49.1,46.1,23.7.HRMS(ESI)m / z calcd for C 26 H 27 N3O5S[M+H] + 494.1745, found 494.1739.

[0115] Example 15: Preparation of a PLK1 inhibitor: N-2-(2-(pyrrolidinyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (denoted as C4)

[0116] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)pyrrolidine, and the corresponding raw material used in step (3) is 4-(trifluoromethyl)benzylamine. The other steps are the same as in embodiment 1.

[0117] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0118] N-2-(2-(pyrrolidinyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (C4): white velvety crystals, yield 42%, mp 220-221℃; 1H NMR (400MHz, DMSO-d6) δ9.06 (s, 1H), 8.96 (dd, J=8.8, 1.1Hz, 1H), 8.56 (dd, J=7 .4,1.1Hz,1H),8.46(d,J=7.6Hz,1H),8.31(d,J=7.6Hz,1H),8.00(dd,J=8.8,7. 4Hz,1H),7.27(d,J=8.1Hz,2H),7.19(d,J=8.1Hz,2H),4.20(s,2H),4.15(t,J= 7.2Hz, 2H), 2.66 (t, J = 7.2Hz, 2H), 2.53 (d, J = 5.5Hz, 4H), 1.68 (t, J = 3.2Hz, 4H). 13 C NMR(100MHz,DMSO-d6)δ163.33,162.8,142.0,141.5,131.5,131.2,129.6(d,J=274.0Hz),129.1,128.6,1 28.5,128.0,127.7,126.5,126.2,124.8(d,J=3.8Hz),123.1,54.2,53.3,49.1,45.9,23.7.HRMS(ESI)m / z calcd for C 26 H 24 N3O4F3S[M+H] + 532.1473, found 532.1513.

[0119] Example 16: Preparation of a PLK1 inhibitor: N-2-(2-(pyrrolyl)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (denoted as C5)

[0120] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)pyrrolidine, and the corresponding raw material used in step (3) is 4-methylbenzylamine. The other steps are the same as in Example 1.

[0121] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0122] N-2-(2-(pyrrolidinyl)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (C5): white flaky crystals, yield 40%, mp 155-156℃; 1H NMR(400MHz, DMSO-d6)δ8.99(dd,J=8.8,1.1Hz,1H),8.84(s,1H),8.56(dd,J=7.4 ,1.1Hz,1H),8.48(d,J=7.6Hz,1H),8.30(d,J=7.6Hz,1H),7.99(dd,J=8.8,7.4Hz, 1H),6.86(d,J=8.0Hz,2H),6.76(d,J=7.9Hz,2H),4.17(t,J=7.0Hz,2H),4.03(s, 2H),2.68(t,J=7.0Hz,2H),2.52(d,J=6.4Hz,4H),2.08(s,3H),1.72-1.61(m,4H). 13 C NMR(100MHz,DMSO-d6)δ163.5,163.0,141.8,136.5,134.6,131.5,131.4,129.9,129.3,12 9.0,128.7,127.9,126.6,126.2,123.1,54.2,53.4,49.1,46.3,23.7,20.7.HRMS(ESI)m / z calcd for C 26 H 27 N3O4S[M+H] + 478.1756, found 478.1796.

[0123] Example 17: Preparation of a PLK1 inhibitor: N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-benzyl-1,8-naphthalenediamide (denoted as D1)

[0124] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)piperidine, and the corresponding raw material used in step (3) is benzylamine. The other steps are the same as in Example 1.

[0125] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0126] N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-benzyl-1,8-naphthalenediamide (D1): white powdery solid, yield 62%, mp 102-103℃; 1H NMR (400MHz, DMSO-d6) δ9.02(dd,J=8.8,1.1Hz,1H),8.90(s,1H),8.56(dd,J=7.4,1.1Hz,1H),8.50(d,J=7.6Hz,1H),8.33(d,J=7.6Hz,1H), 8.01(dd,J=8.8,7.4Hz,1H),7.06(s,5H),4.16(t,J=7.0Hz,2H),4.10(s,2H),2.64-2.52(m,2H),2.45(s,4H),1.42(dd,J=35.9,4.7Hz,6H). 13 C NMR(100MHz,DMSO-d6)δ163.4,162.9,141.8,137.4,131.5,131.4,129.8,129.2,129.0,128.7,1 28.3,127.9,127.4,126.7,126.3,123.2,56.1,54.7,49.1,46.5,37.9,26.1,24.4.HRMS(ESI)m / z calcd forC 26 H 27 N3O4S[M+H] + 478.1796, found 478.1787.

[0127] Example 18: Preparation of a PLK1 inhibitor: N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (denoted as D2)

[0128] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)piperidine, and the corresponding raw material used in step (3) is 4-fluorobenzylamine. The other steps are the same as in embodiment 1.

[0129] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0130] N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (D2): white needle-like crystals, yield 45%, mp 105-106℃; 1H NMR (400MHz, DMSO-d6) δ9.00(dd,J=8.8,1.1Hz,1H),8.91(s,1H),8.57(dd,J=7.4,1.1Hz,1H),8.51(d,J=7.8Hz,1H),8.33(d,J=7.8Hz,1H),8.0 1(dd,J=8.8,7.4Hz,1H),7.13-7.01(m,2H),6.92-6.81(m,2H),4.17(t, J=7.1Hz,2H),4.08(s,2H),2.54(s,2H),2.44(s,4H),1.55-1.30(m,6H). 13 C NMR(100MHz,DMSO-d6)δ13C NMR (101MHz, DMSO) δ163.4,162.9,161.6(d,J=241.7Hz),141.7,133.7(d,J=3.0Hz),131.5,131.3,129.98,129.9(d,J=8.2H z),129.2,129.1,128.7,126.6,126.3,123.2,115.0(d,J=21.3Hz),56.1,54.6,49.1,45.7,37.8,26.1,24.4.HRMS(ESI)m / z calcd forC 26 H 26 N3O4FS[M+H] + 496.1662, found 496.1691.

[0131] Example 19: Preparation of a PLK1 inhibitor: N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (denoted as D3)

[0132] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)piperidine, and the corresponding raw material used in step (3) is 4-methoxybenzylamine. The other steps are the same as in Example 1.

[0133] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0134] N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (D3): white powdery solid, yield 60%, mp 152-153℃; 1H NMR (400MHz, DMSO-d6) δ8.98 (dd, J=8.8, 1.1Hz, 1H), 8.79 (s, 1H), 8.55 (d, J=7. 2Hz,1H),8.48(d,J=7.6Hz,1H),8.29(d,J=7.6Hz,1H),7.98(t,J=8.0Hz,1H),6 .88(d,J=8.4Hz,2H),6.50(d,J=8.5Hz,2H),4.16(t,J=7.2Hz,2H),4.02(s,2H) ,3.57(s,3H),2.53(d,J=6.0Hz,2H),2.43(s,4H),1.42(dd,J=37.2,4.6Hz,6H). 13 C NMR (100MHz, DMSO-d6) δ163.4,162.9,158.6,141.9,131.4,131.4,129.8,129.3,129.1,129. 0,128.6,126.6,126.2,123.1,113.5,56.1,55.4,54.7,46.1,37.9,26.1,24.5.HRMS(ESI)m / z calcd for C 27 H 29 N3O5S[M+H] + 508.1861, found 508.1893.

[0135] Example 20: Preparation of a PLK1 inhibitor: N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (denoted as D4)

[0136] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)piperidine, and the corresponding raw material used in step (3) is 4-(trifluoromethyl)benzylamine. The other steps are the same as in embodiment 1.

[0137] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0138] N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (D4): white needle-like crystals, yield 46%, mp 177-178℃; 1H NMR (400MHz, DMSO-d6) δ9.06(s,1H),8.96(d,J=8.6Hz,1H),8.56(d,J=7.2Hz,1H),8.46(d,J=7.6Hz,1H),8.31(d,J=7.6Hz,1H),8.00(dd,J=8 .8,7.4Hz,1H),7.27(d,J=8.1Hz,2H),7.19(d,J=8.1Hz,2H),4.21(s,2H),4.14(t,J=7.2Hz,2H),2.51(s,2H),2.45(s,4H),1.55-1.31(m,6H). 13 C NMR(100MHz,DMSO-d6)δ13C NMR (101MHz, DMSO) δ163.3,162.8,142.0,141.5,131.5,131.2,129.7,129.5,129.1,128.6,128.5,128.0,12 7.8(d,J=31.4Hz),126.2,124.8(d,J=3.7Hz),123.1,56.1,54.6,49.1,45.9,37.9,26.1,24.4.HRMS(ESI)m / z calcd for C 27 H 26 N3O4F3S[M+H] + 546.1630, found 546.1658.

[0139] Example 21: Preparation of a PLK1 inhibitor: N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (denoted as D5)

[0140] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 1-(2-aminoethyl)piperidine, and the corresponding raw material used in step (3) is 4-methylbenzylamine. The other steps are the same as in Example 1.

[0141] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0142] N-2-(2-(piperidinyl)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (D5): pale yellow needle-like crystals, yield 30%, mp 162-163℃; 1H NMR (400MHz, DMSO-d6) δ8.99 (dd, J=8.8, 1.0Hz, 1H), 8.82 (s, 1H), 8.56 (dd, J= 7.4,1.0Hz,1H),8.48(d,J=7.6Hz,1H),8.29(d,J=7.6Hz,1H),7.99(dd,J=8.8, 7.4Hz,1H),6.86(d,J=8.0Hz,2H),6.77(d,J=7.8Hz,2H),4.16(t,J=7.2Hz,2H) ,4.04(s,2H),2.57-2.52(m,2H),2.43(s,4H),2.08(s,3H),1.53-1.30(m,6H). 13 C NMR (100MHz, DMSO-d6) δ163.5,163.0,141.9,136.5,134.2,131.4,129.8,129.3,129.0,128. 7,128.7,127.9,126.6,126.2,123.1,56.2,54.7,46.3,37.9,26.1,24.4,20.9.HRMS(ESI)m / z calcd for C 27 H 29 N3O4S[M+H] + 492.1912, found 492.1950.

[0143] Example 22: Preparation of a PLK1 inhibitor: N-2-(2-(morpholino)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (denoted as E2)

[0144] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is N-(2-aminoethyl)morpholine, and the corresponding raw material used in step (3) is 4-fluorobenzylamine. The other steps are the same as those in embodiment 1.

[0145] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0146] N-2-(2-(morpholino)ethyl)-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (E2): Brown powdery solid, yield 41%, mp 143-144℃; 1H NMR (400MHz, DMSO-d6) δ8.99(d,J=8.6Hz,1H),8.90(s,1H),8.57(d,J=7.2Hz,1H),8.51(d,J=7.6Hz,1H),8.32(d,J=7.8Hz,1H),8.00(dd,J=8.8,7.4H z,1H),7.07(dd,J=8.5,5.6Hz,2H),6.86(t,J=8.9Hz,2H),4.18(t,J=6.9H z,2H),4.08(s,2H),3.57-3.49(m,4H),2.57(t,J=6.9Hz,2H),2.46(s,4H). 13 C NMR (100MHz, DMSO-d6) δ163.5,162.9,161.6(d,J=241.7Hz), 141.7,133.7(d,J=3.0Hz), 131.5,131.3,129.9(d,J=8.2Hz), 129.9,129.2(d,J=17.7Hz),128.7,126.6,126.3,123.2,115.0(d,J=21.2Hz),66.7,55.9,53.9,45.7,37.5.HRMS(ESI)m / z calcd forC 25 H 24 N3O5FS[MH] - 496.1347, found 496.1338.

[0147] Example 23: Preparation of a PLK1 inhibitor: N-2-(2-(morpholino)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (denoted as E3)

[0148] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is N-(2-aminoethyl)morpholine, and the corresponding raw material used in step (3) is 4-methoxybenzylamine. The other steps are the same as in Example 1.

[0149] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0150] N-2-(2-(morpholino)ethyl)-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (E3): Brown powdery solid, yield 48%, mp 182-183℃; 1H NMR (400MHz, DMSO-d6) δ8.98(d,J=8.6Hz,1H),8.79(s,1H),8.56(d,J=7.2Hz,1H),8.48(d,J=7.6Hz,1H),8.30(d,J=7.6Hz,1H),8.04-7.95(m,1H) ,6.88(d,J=8.6Hz,2H),6.51(d,J=8.6Hz,2H),4.18(t,J=7.0Hz,2H),4.0 2(d,J=3.2Hz,2H),3.62-3.49(m,7H),2.58(t,J=7.0Hz,2H),2.48(s,4H). 13 C NMR (100MHz, DMSO-d6) δ163.5,163.0,158.6,141.9,131.5,131.4,129.9,129.3,129.1,129. 0,128.7,126.7,126.2,123.1,113.5,66.7,55.9,55.4,53.9,46.1,37.5.HRMS(ESI)m / zcalcd for C 26 H 27 N3O6S[M+H] + 510.1654, found 510.1688.

[0151] Example 24: Preparation of a PLK1 inhibitor: N-2-(2-(morpholino)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (denoted as E4)

[0152] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is N-(2-aminoethyl)morpholine, and the corresponding raw material used in step (3) is 4-(trifluoromethyl)benzylamine. The other steps are the same as in Example 1.

[0153] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0154] N-2-(2-(morpholinyl)ethyl)-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (E4): white needle-like crystals, yield 43%, mp 175-176℃; 1H NMR (400MHz, DMSO-d6) δ9.05(s,1H),8.97(d,J=8.5Hz,1H),8.56(d,J=7.0Hz,1H),8.46(d,J=7.6Hz,1H),8.31(d,J=7.6Hz,1H),8.00(dd,J=8.6 ,7.4Hz,1H),7.27(d,J=8.1Hz,2H),7.19(d,J=8.1Hz,2H),4.18(dd,J=15.9,8.9Hz,4H),3.59-3.51(m,4H),2.56(t,J=7.0Hz,2H),2.47(s,4H). 13 C NMR (100MHz, DMSO-d6) δ163.4, 162.8, 142.0, 142.0, 141.6, 131.5, 129.9 (d, J = 272.1Hz), 129.6 (d, J = 29.0Hz), 129. 1,127.8(d,J=31.6Hz),126.5,126.2,125.8,124.9,124.8,123.1,66.7,55.9,53.9,45.9,37.5.HRMS(ESI)m / zcalcd for C 26 H 24 N3O5F3S[M+H] + 548.1422, found 548.1453.

[0155] Example 25: Preparation of a PLK1 inhibitor: N-2-(2-(morpholino)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (denoted as E5)

[0156] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is N-(2-aminoethyl)morpholine, and the corresponding raw material used in step (3) is 4-methylbenzylamine. The other steps are the same as in Example 1.

[0157] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0158] N-2-(2-(morpholino)ethyl)-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (E5): pale yellow needle-like crystals, yield 34%, mp 167-168℃; 1H NMR (400MHz, DMSO-d6) δ8.99(d,J=8.6Hz,1H),8.83(t,J=6.0Hz,1H),8.56(d,J =7.4Hz,1H),8.49(d,J=7.6Hz,1H),8.30(d,J=7.6Hz,1H),8.04-7.95(m,1H),6. 87(d,J=7.8Hz,2H),6.78(d,J=7.8Hz,2H),4.18(t,J=6.8Hz,2H),4.04(d,J=5.8 Hz,2H),3.54(t,J=4.8Hz,4H),2.59(t,J=6.8Hz,2H),2.48(s,4H),2.08(s,3H). 13 C NMR(100MHz,DMSO-d6)δ163.5,163.0,141.9,136.5,134.2,131.5,129.9,129.3,129.0,12 8.7,128.7,127.9,126.7,126.1,123.1,66.7,56.0,53.9,46.3,37.5,20.9.HRMS(ESI)m / z calcd for C 26 H 27 N3O5S[M+H] + 494.1705, found 494.1738.

[0159] Example 26: Preparation of a PLK1 inhibitor: N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-benzyl-1,8-naphthalenediamide (denoted as F1)

[0160] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is 3-dimethylamino-1-propylamine, and the corresponding raw material used in step (3) is benzylamine. The other steps are the same as those in embodiment 1.

[0161] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0162] N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-benzyl-1,8-naphthalenediamide (F1): white needle-like crystals, yield 44%, mp 113-114℃; 1H NMR(400MHz, DMSO-d6)δδ9.04-9.00(m,1H),8.91(s,1H),8.57(d,J=6.7Hz,1H),8.51(d,J=7.6Hz,1H),8.34(d,J=7.6Hz,1H ),8.01(dd,J=8.8,7.4Hz,1H),7.11-7.04(m,5H),4.10-4.04(m,4H),2.32(t,J=7.0Hz,2H),2.14(s,6H),1.86-1.69(m,2H). 13 CNMR(100MHz,DMSO-d6)δ163.6,163.0,141.7,137.5,131.4,131.3,129.8,129.1,129.0,12 8.8,128.4,127.9,127.4,126.7,126.4,123.3,57.2,46.5,45.5,38.9,25.9.HRMS(ESI)m / z calcd for C 24 H 25 N3O4S[M+H] + 452.1639, found 452.1617.

[0163] Example 27: Preparation of a PLK1 inhibitor: N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (denoted as F2)

[0164] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is 3-dimethylamino-1-propylamine, and the corresponding raw material used in step (3) is 4-fluorobenzylamine. The other steps are the same as in embodiment 1.

[0165] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0166] N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-fluorobenzyl)-1,8-naphthalenediamide (F2): White blocky crystals, yield 56%, mp 198-199℃; 1H NMR (400MHz, DMSO-d6) δ8.99(d,J=8.1Hz,1H),8.91(s,1H),8.57(d,J=6.7Hz,1H),8.51(d,J=7.6Hz,1H),8.33(d,J=7.6Hz,1H),8.01( dd,J=8.8,7.4Hz,1H),7.13-7.00(m,2H),6.92-6.79(m,2H),4.21-3.92(m,4H),2.32(t,J=7.2Hz,2H),2.13(s,6H),1.86-1.69(m,2H). 13 C NMR (100MHz, DMSO-d6) δ163.4,162.8,142.0,142.0,141.6,131.5,129.9(d,J=272.1Hz),129.6(d,J=29.0Hz),12 9.1,127.8(d,J=31.6Hz),126.5,126.2,125.8,124.9,124.8,123.1,66.7,55.9,53.9,45.9,37.5.HRMS(ESI)m / z calcd for C 24 H 24 N3O4FS[M+H] + 470.1545, found 470.1533.

[0167] Example 28: Preparation of a PLK1 inhibitor: N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (denoted as F3)

[0168] The difference between this embodiment and embodiment 1 is that the corresponding amine used in step (1) of this embodiment is 3-dimethylamino-1-propylamine, and the corresponding raw material used in step (3) is 4-methoxybenzylamine. The other steps are the same as in embodiment 1.

[0169] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0170] N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-methoxybenzyl)-1,8-naphthalenediamide (F3): yellow flaky crystals, yield 50%, mp 209-210℃; 1H NMR (400MHz, DMSO-d6) δ8.97(dd,J=8.8,1.1Hz,1H),8.79(s,1H),8.54(dd,J=7.4,1.0Hz,1H),8.46(d,J=7.7Hz,1H),8.28(d,J=7.6Hz,1H),7.98(dd,J=8 .8,7.6Hz,1H),6.98-6.79(m,2H),6.60-6.38(m,2H),4.09-4.02(m,2H),4.0 1(s,2H),3.55(s,3H),2.32(t,J=6.8Hz,2H),2.14(s,6H),1.81-1.74(m,2H). 13 C NMR(100MHz,DMSO-d6)δ163.6,163.0,158.5,141.8,131.4,131.3,129.8,129.3,129.0,12 8.9,128.7,126.6,126.3,123.2,113.4,57.2,55.3,46.1,45.5,38.9,25.9.HRMS(ESI)m / z calcd for C 25 H 27 N3O5S[M+H] + 482.1745, found 482.1735.

[0171] Example 29: Preparation of a PLK1 inhibitor: N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (denoted as F4)

[0172] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 3-dimethylamino-1-propylamine, and the corresponding raw material used in step (3) is 4-(trifluoromethyl)benzylamine. The other steps are the same as in Example 1.

[0173] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0174] N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-(trifluoromethyl)benzyl)-1,8-naphthalenediamide (F4): white needle-like crystals, yield 40%, mp 171-172℃; 1H NMR (400MHz, DMSO-d6) δ9.05(s,1H),8.95(dd,J=8.6,1.1Hz,1H),8.56(dd,J=7.4,1.1Hz,1H),8.45(d,J=7.6Hz,1H),8.30(d,J=7.6Hz,1H),8.00(dd,J= 8.6,7.4Hz,1H),7.23(d,J=8.0Hz,2H),7.16(d,J=8.0Hz,2H),4.20(s,2H),4 .10(t,J=7.2Hz,2H),2.35(t,J=7.2Hz,2H),2.17(s,6H),1.80-1.73(m,2H). 13 C NMR(100MHz,DMSO-d6)δ163.5,162.9,141.8,141.5,131.4,131.2,129.7,129.6,129.1,128.6 ,128.5,126.49,126.3,124.7(d,J=2.4Hz),123.2,57.0,45.9,45.3,38.8,25.7.HRMS(ESI)m / z calcd for C 25 H 24 N3O4F3S[M+H] + 520.1473, found 520.1495.

[0175] Example 30: Preparation of a PLK1 inhibitor: N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (denoted as F5)

[0176] The difference between this embodiment and Example 1 is that the corresponding amine used in step (1) of this embodiment is 3-dimethylamino-1-propylamine, and the corresponding raw material used in step (3) is 4-methylbenzylamine. The other steps are the same as in Example 1.

[0177] Using nuclear magnetic resonance (NMR) 1 H NMR and 13 The target product obtained in this embodiment was characterized by C10 NMR and ESI-MS, and the results are as follows:

[0178] N-2-(2-N,N-dimethylamino)propyl-6-sulfonamide-(4-methylbenzyl)-1,8-naphthalenediamide (F5): white square crystals, yield 44%, mp 212-213℃; 1H NMR(400MHz, DMSO-d6)δ8.99(dd,J=8.8,1.1Hz,1H),8.83(s,1H),8.56(dd,J=7 .4,1.1Hz,1H),8.48(d,J=7.6Hz,1H),8.29(d,J=7.6Hz,1H),7.99(dd,J=8.8,7. 4Hz,1H),6.85(d,J=8.0Hz,2H),6.77(d,J=8.0Hz,2H),4.07(t,J=6.8Hz,2H),4 .03(s,2H),2.32(t,J=6.8Hz,2H),2.14(s,6H),2.08(s,3H),1.81-1.74(m,2H). 13 C NMR(101MHz,DMSO-d6)δ163.57,163.04,141.78,136.47,134.16,131.37,129.76,129.27,128.95, 128.68,127.91,126.63,126.26,123.21,57.16,46.31,45.51,38.92,25.90,20.89.HRMS(ESI)m / z calcd for C 25 H 28 N3O4S[M+H] + 466.1796, found 466.1783.

[0179] In addition, in the above preparation method, the molar ratio of the raw materials used in each reaction is a stoichiometric ratio; the organic solvent used can be one or a combination of two or more selected from ethanol, methanol, dichloromethane (DCM) and N,N-dimethylformamide (DMF). When the organic solvent is a combination of two or more of the above substances, their ratio can be arbitrary; the amount of organic solvent can be determined as needed. Generally, based on 1 mmol of the compound shown in Example 1, all the raw materials participating in the reaction are dissolved in 5-10 mL of organic solvent.

[0180] In actual operation, the reaction can be carried out under no heating or heating conditions, preferably under no heating conditions, and the reaction is monitored by TLC to detect whether the reaction is complete. In step (1), the first solvent is preferably ethanol; the reaction is usually carried out at 40°C to the reflux temperature of the first solvent, preferably under stirring conditions; the product obtained in this step can be further purified (such as recrystallization, the recrystallization solvent can be a common solvent such as ethanol) before being used in subsequent operations; in step (2), the second solvent is preferably DMF; the reaction is usually carried out at 40°C to the reflux temperature of the first solvent, and the product of this step does not need to be purified and can be directly used in the next step of the reaction.

[0181] All compounds obtained in this invention can be separated by high performance liquid chromatography, silica gel column chromatography, or thin-layer chromatography.

[0182] To fully illustrate the use of the PLK1 inhibitor described in this invention in pharmaceutical manufacturing, the applicant conducted PLK1 binding capacity determination and in vitro antitumor activity experiments.

[0183] 1. Surface Plasmon Resonance Experiment (SPR)

[0184] The experiment was conducted on a Biacore T200 surface plasmon resonance spectrometer.

[0185] A. Pre-enrichment: Take 3 μL of pre-prepared PLK1 protein solution with a mass concentration of 200 μg·mL. -1 Take 97 μL of sodium acetate solution with pH values ​​of 4.5, 5.0, and 5.5 respectively, dissolve it thoroughly, and place it in a labeled EP tube with the cap removed. Then, measure 500 μL of NaOH solution and place it in another uncapped EP tube for CM5 chip regeneration. During the pre-enrichment test, first select chip channel 2 as the protein coupling channel and channel 1 as the reference channel. Set the sample injection program as follows: sample flow cell temperature to 16℃, detection temperature to 25℃, PLK1 protein solution injection time to 30 s, waiting time to 10 s, NaOH solution regeneration time to 30 s, and flow rate to 10 μL / min. -1 The RU value is obtained and used to determine the optimal coupling condition.

[0186] B. Coupling: According to the conditions of the optimal ligand enrichment experiment, take 9 μL of the PLK1 protein solution to be used and place it at the bottom of an open EP tube. Then add 141 μL of sodium acetate solution at pH 5.0 and mix thoroughly. Take three more open EP tubes and add 100 μL of EDC solution, 100 μL of NHS solution, and 150 μL of ethanolamine solution to each tube. The coupling value is approximately 16000 RU.

[0187] C. Determination of affinity: Prepare analytical samples with gradient concentrations of the analyte, namely 0 μM, 0.16 μM, 0.31 μM, 0.62 μM, 1.25 μM, 2.5 μM, 5 μM, and 10 μM. Inject the analytical samples into the chip at a flow rate of 30 μL / min for 120 s, followed by dissociation for 300 s.

[0188] D. Analysis: Use the 1:1 combined model in the Biaevaluation program to process the dynamic data.

[0189] In this invention, the Polo-like kinase 1 inhibitor Volasetib was selected as a control, and the results are shown in the appendix. Figure 1 As shown, compared to Volasertib, the PLK1 inhibitor B2 synthesized in this invention has a better affinity for PLK1; in addition, other compounds also showed good affinity through experiments.

[0190] 2. In vitro thermodynamic migration experiment

[0191] A. For complete cell assays, A549 cells were collected after treatment with 5 μM PLK1 inhibitor or DMSO for 2 hours and washed several times with PBS buffer to avoid excess compound residue. Each cell suspension was aliquoted into separate 0.2 mL PCR tubes. The cells were then denatured at different temperatures for 3 minutes on an Eppendorf PCR instrument, followed by two freeze-thaw cycles with liquid nitrogen. The samples were centrifuged, and the supernatant was analyzed by Western blotting.

[0192] B. For cell lysate analysis, A549 cells were collected and harvested using PBS buffer. Then, 20 μM PLK1 inhibitor or DMSO was added to the supernatant, and the cells were incubated at 25°C for 2 hours. After denaturation at different temperatures for 3 minutes, the samples were centrifuged, and the supernatant was analyzed by Western blotting.

[0193] The results are as follows Figure 1 As shown, in vitro thermodynamic migration experiments detected that B2 can bind to the PLK1 protein and enhance its thermal stability; in addition, experiments showed that other compounds can also enhance the thermal stability of the PLK1 protein.

[0194] 3. In vitro antitumor activity experiment

[0195] A. Cell lines and cell culture

[0196] This experiment used seven tumor cell lines: human lung cancer cells A549, NCI-H460, human gastric cancer cells MGC80-3, human bladder cancer cells T24, human ovarian cancer cells SKOV-3, human liver cancer cells BEL-7402, and human breast cancer cells MCF-7.

[0197] Human lung cancer cells NCI-H460 and A549, human liver cancer cells BEL-7402, and other tumor cell lines were cultured in RPMI-1640 medium containing 10 wt% fetal bovine serum, 100 U / mL penicillin, and 100 U / mL streptomycin, and incubated at 37°C in an incubator containing 5% CO2. Human gastric cancer cells MGC80-3, human bladder cancer cells T24, human ovarian cancer cells SKOV-3, and human breast cancer cells MCF-7 were cultured in DMEM medium containing 10 wt% fetal bovine serum, 100 U / mL penicillin, and 100 U / mL streptomycin.

[0198] B. Preparation of the test compound

[0199] The compounds used were prepared in Examples 1-30, with a purity ≥95%. Their DMSO stock solution (0.002 mol / L) was sequentially diluted with culture medium to five concentration gradients: 20, 10, 5, 2.5, and 1.25 μmol / L, with the final concentration of the co-solvent DMSO ≤1%. First, the inhibitory rate of the target product at 20 μmol / L on tumor cell proliferation was tested, considered as the initial screening result. Then, the inhibitory degree of the target product on the proliferation of various tumor cells at different concentration gradients was tested to calculate the half-maximal inhibitory concentration (IC50).

[0200] C. Cell growth inhibition assay (crystal violet staining method)

[0201] (1) Take cells in the logarithmic growth phase, digest them with trypsin, and prepare a cell suspension with a concentration of 5000 cells / mL using culture medium containing 10% fetal bovine serum. Seed 180 μL per well in a 96-well culture plate to make the cell density to be tested 1000-10000 cells / well (fill the edge wells with sterile PBS).

[0202] (2) 5% CO2, incubate at 37°C for 24 h until the cell monolayer covers the bottom of the well, add 20 μL of drug at a certain concentration gradient to each well, and set 5 replicates for each concentration gradient;

[0203] (3) Incubate with 5% CO2 at 37°C for 48 hours and observe under an inverted microscope;

[0204] (4) Fixation: Wash twice with PBS, and fix cells with 100 μL of 10% methanol for 15 min;

[0205] (5) Crystal violet staining: Remove 10% methanol fixative, add 100μL 0.1% crystal violet staining solution and stain for 15min, then wash three times with deionized water, slowly wash away the staining solution, and air dry.

[0206] (6) Decolorization: Decolorize with 100 μL of 33% acetic acid solution per well, shake thoroughly, and measure the absorbance at a wavelength of 570 nm using an enzyme-linked immunosorbent assay (ELISA) reader.

[0207] (7) Calculate the inhibition rate: according to the formula:

[0208] (8) Find IC 50 Value: Based on the calculated inhibition rate, the IC is then calculated using SPSS20 software. 50 The values ​​were repeated three times, and the average value was taken. The standard deviation was then calculated.

[0209] The inhibition rate of the compound on cell growth was calculated, and the inhibition rate data of five concentration gradients were further fitted using SPSS software to determine the half-maximal inhibitory concentration (IC50) of the compound for different cell lines. 50 Value (in μmol / L), IC50 of the compound for different cell lines 50 The values ​​are shown in Table 1.

[0210] Table 1: IC50 of the compounds of the present invention against cancer cell lines and normal hepatocyte lines 50 Value (μM)

[0211]

[0212]

[0213] As shown in Table 1, the Polo-like kinase inhibitors described in this invention exhibit significant inhibitory effects on the proliferation of various tumor cell lines. Among these, the R1 substituent in the synthesized compounds has a significant impact on antitumor activity; when the R1 group is... At that time, its IC50 values ​​against multiple tumor cell lines ranged from 1 to 10 μM, while when the R1 group was hexyl, At that time, its IC 50 The concentration is greater than 20 μM. In this application, when the concentrations of A2, A3, A4, A5, E2, E3, E4, E5, F1, F2, F3, F4, and F5 are 20 μmol / L, the inhibition rate can reach 20%-50%, and all of them can be used as effective antitumor drugs.

[0214] The above description is a detailed description of the preferred embodiments of the present invention. However, the embodiments are not intended to limit the scope of the patent application of the present invention. All equivalent changes or modifications made under the technical spirit of the present invention should fall within the patent scope covered by the present invention.

Claims

1. The application of PLK1 inhibitors in the preparation of antitumor drugs, wherein the PLK1 inhibitor is a compound having the structure of formula (I) or a pharmaceutically acceptable salt thereof. ； in, R1 is selected from ethyl, , , , or R2 is selected from -H, methyl, methoxy, fluorine or trifluoromethyl.

2. The application according to claim 1, characterized in that, The application refers to the use of PLK1 inhibitors in the preparation of drugs for the prevention or treatment of non-small cell lung cancer, liver cancer, ovarian cancer, or bladder cancer.

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