阅读说明：本技术 一种黄酮类端锚聚合酶2抑制剂及其制备方法和应用 (Flavonoid tankyrase 2 inhibitor and preparation method and application thereof ) 是由 庞婉 李堂 王杰 赵泽圣 张孝礼 于 2021-07-20 设计创作，主要内容包括：本发明涉及一种黄酮类端锚聚合酶2抑制剂及其制备方法和应用,具有式I所示的结构：该抑制剂采用4-氧代-4H-1-苯并吡喃-2-羧酸和苯胺衍生物或苯甲酰肼衍生物作为起始原料,使用脲鎓盐作为缩合剂,在添加碱的条件下,于有机溶剂中进行缩合反应获得。与现有技术相比,本发明制备方法简单,合成的化合物结构新颖,具有较好的抑制端锚聚合酶2的活性以及更好的选择性。(The invention relates to a flavonoid tankyrase 2 inhibitor, a preparation method and application thereof, and the flavonoid tankyrase 2 inhibitor has a structure shown in a formula I: the inhibitor is obtained by taking 4-oxo-4H-1-benzopyran-2-carboxylic acid and aniline derivatives or benzoyl hydrazine derivatives as starting materials, using uronium salts as condensing agents and carrying out condensation reaction in an organic solvent under the condition of adding alkali. Compared with the prior art, the preparation method is simple, and the synthesized compound has a novel structure, and has better activity for inhibiting tankyrase 2 and better selectivity.)

1. A flavonoid tankyrase 2 inhibitor is characterized by having a structure shown in formula I:

wherein R is selected from:

any one of them.

2. The flavonoid tankyrase 2 inhibitor according to claim 1, wherein R is selected from the group consisting of:

any one of them.

3. The method for preparing the flavonoid tankyrase 2 inhibitor according to claim 1, wherein the flavonoid tankyrase 2 inhibitor is obtained by performing a condensation reaction in an organic solvent using 4-oxo-4H-1-benzopyran-2-carboxylic acid and an aniline derivative or a benzoyl hydrazine derivative as starting materials and a uronium salt as a condensing agent with the addition of a base;

the aniline derivative is

The benzoyl hydrazine derivative is

Wherein:

R₁selected from H, CH₃、OCH₃Cl, Br or 3,4,5-tri-OCH₃；

R₂Selected from H, CH₃、OCH₃Cl or Br.

4. The method of preparing a flavonoid tankyrase 2 inhibitor of claim 3, wherein said uronium salt is selected from HATU, HBTU, HCTU, TBTU, TSTU or TNTU.

5. The method for preparing the flavonoid tankyrase 2 inhibitor according to claim 3 or 4, wherein the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the condensing agent is 1:1 to 1: 5.

6. The preparation method of the flavonoid tankyrase 2 inhibitor according to claim 3, wherein the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the aniline derivative or the benzoyl hydrazine derivative is 1:1 to 1: 3.

7. The method for preparing a flavonoid tankyrase 2 inhibitor according to claim 3, wherein any one or more of the following conditions are included:

(i) the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the alkali is 1: 1-1: 5;

(ii) the base is selected from DIPEA or triethylamine.

8. The method for preparing the flavonoid tankyrase 2 inhibitor according to claim 3, wherein the organic solvent is selected from DMSO, DMF or NMP.

9. The method for preparing the flavonoid tankyrase 2 inhibitor according to claim 3, wherein the reaction temperature of the condensation reaction is 0 ℃ to 70 ℃.

10. Use of the flavonoid tankyrase 2 inhibitor of claim 1 or 2 for the preparation of a medicament for inhibiting tankyrase 2.

Technical Field

The invention belongs to the technical field of medicinal chemistry, and particularly relates to a flavonoid tankyrase 2 inhibitor as well as a preparation method and application thereof.

Background

Tankyrase is a multifunctional protein translation modifying enzyme, a member of the polyadenylic diphosphoribose polymerase family, and its main domain is ankyrin repeat. The enzyme can be combined with a telomere repetitive sequence factor, and can also be combined with a highly conserved structural domain of a carboxyl terminal of an axial inhibitory protein, so that the inhibition effect on the telomere enzyme is realized, and an extracellular factor (Wnt)/beta-catenin (beta-catenin) signal path is adjusted, so that the proliferation and the growth of tumor cells are inhibited. A tankyrase inhibitor XAV939 reported in Nature in 2009 is directly synthesized based on the crystal structure of tankyrase 1/2, and can selectively block the activation of a Wnt/beta-catenin signaling pathway through the expression of a stable axis inhibiting protein, so as to limit the proliferation and growth of tumor cells. In recent years, tankyrase 2(TNKS2) has attracted much attention as a new target for antitumor drugs. More and more researches on the medicines developed by TNKS show the huge development prospect of the antitumor medicines. Flavonoids are a large group of compounds that occur widely in nature and have antioxidant properties. In addition, flavonoid derivatives have been shown to inhibit tankyrase 2 and have antiproliferative properties in lung, prostate, colorectal, pancreatic and ovarian cancer cells. Most of the reported flavonoid tankyrase 2 inhibitors act only on the nicotinamide binding site of tankyrase 2.

Disclosure of Invention

In order to find a novel tankyrase 2 inhibitor, the invention provides a novel flavonoid tankyrase 2 inhibitor, a preparation method thereof and application thereof in preparing a medicament for inhibiting tankyrase 2.

The inhibitor which acts on the binding site of nicotinamide and adenosine has better inhibitory activity and selectivity. The newly designed and synthesized flavonoid compound has longer branched chains and can simultaneously occupy the nicotinamide and adenosine binding sites of tankyrase 2, thereby improving the inhibitory activity and selectivity of the compound.

In the invention, a computer-aided drug design method is utilized to design the structure of the tankyrase 2 and synthesize a plurality of tankyrase 2 inhibitors with novel structures, and finally the tankyrase 2 inhibitors are subjected to tankyrase design2 inhibitory Activity and the reported inhibitory Activity of tankyrase 2 inhibitor GK-007 (IC of GK-007)₅₀0.035uM), the synthesized multiple tankyrase 2 inhibitors were found to have better tankyrase 2 inhibitory activity, especially IC of 5 compounds preferred among them₅₀8.74uM, 3.54uM, 1.25uM, 0.89uM, 0.082uM, respectively.

The purpose of the invention can be realized by the following technical scheme:

the invention provides a flavonoid tankyrase 2 inhibitor, which has a structure shown in a formula I:

wherein R is selected from:

any one of them.

Preferably, said R is selected from:

any one of them.

The second aspect of the present invention provides a method for preparing the flavonoid tankyrase 2 inhibitor, wherein the method comprises the steps of using 4-oxo-4H-1-benzopyran-2-carboxylic acid and aniline derivatives or benzoyl hydrazine derivatives as starting materials, using uronium salts as a condensing agent, and performing a condensation reaction in an organic solvent under the condition of adding alkali to obtain the flavonoid tankyrase 2 inhibitor;

the aniline derivative is

The benzoyl hydrazine derivative is

Wherein:

R₁selected from H, CH₃、OCH₃Cl, Br or 3,4,5-tri-OCH₃；

R₂Selected from H, CH₃、OCH₃Cl or Br.

The reaction equations are shown in the following three processes a, b and c respectively:

preferably, the uronium salt is selected from HATU, HBTU, HCTU, TBTU, TSTU or TNTU. Preferably, the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the condensing agent is 1: 1-1: 5

Preferably, the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the aniline derivative or the benzoyl hydrazine derivative is 1: 1-1: 3.

Preferably, the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the alkali is 1: 1-1: 5.

Preferably, the base is selected from DIPEA or triethylamine.

Preferably, the organic solvent is selected from DMSO, DMF or NMP.

Preferably, the reaction temperature of the condensation reaction is from 0 ℃ to 70 ℃.

The third aspect of the invention provides an application of the flavonoid tankyrase 2 inhibitor in preparation of a drug for inhibiting tankyrase 2.

Compared with the prior art, the invention has the beneficial effects that:

(1) the preparation method is simple, low in cost and mild in reaction condition.

(2) The compound synthesized by the invention has a novel structure, and is reported for the first time.

(3) The flavonoid tankyrase 2 inhibitor synthesized by the invention has good effect, wherein the compound 5 (see example 5) has similar inhibitory activity to GK-007 and better selectivity.

Drawings

FIG. 1 shows a hydrogen spectrum of the product obtained in example 1: (¹H NMR(DMSO))。

FIG. 2 is a carbon spectrum of the product obtained in example 1: (¹³C NMR(DMSO))。

FIG. 3 is a hydrogen spectrum of the product obtained in example 2: (¹H NMR(DMSO))。

FIG. 4 is a carbon spectrum of the product obtained in example 2 (C:)¹³C NMR(DMSO))。

FIG. 5 is a hydrogen spectrum of the product obtained in example 3 (¹H NMR(DMSO))。

FIG. 6 is a carbon spectrum of the product obtained in example 3 (C:)¹³C NMR(DMSO))。

FIG. 7 is a hydrogen spectrum (C) of the product obtained in example 4¹H NMR(DMSO))。

FIG. 8 is a carbon spectrum of the product obtained in example 4 (C:)¹³C NMR(DMSO))。

FIG. 9 is a hydrogen spectrum (C) of the product obtained in example 5¹H NMR(DMSO))。

FIG. 10 is a carbon spectrum of the product obtained in example 5 (C:)¹³C NMR(DMSO))。

Detailed Description

A flavonoid tankyrase 2 inhibitor has a structure shown in formula I:

wherein R is selected from:

any one of them.

Preferably R is selected from:

any one of them.

The preparation method of the flavonoid tankyrase 2 inhibitor adopts 4-oxo-4H-1-benzopyran-2-carboxylic acid and aniline derivatives or benzoyl hydrazine derivatives as starting materials, uses uronium salts as a condensing agent, and carries out condensation reaction in an organic solvent under the condition of adding alkali to obtain the flavonoid tankyrase 2 inhibitor;

the aniline derivative is

The benzoyl hydrazine derivative is

Wherein:

R₁selected from H, CH₃、OCH₃Cl, Br or 3,4,5-tri-OCH₃；

R₂Selected from H, CH₃、OCH₃Cl or Br.

Preferably the uronium salt is selected from HATU, HBTU, HCTU, TBTU, TSTU or TNTU. The molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the condensing agent is preferably 1:1 to 1: 5. Preferably, the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the aniline derivative or the benzoyl hydrazine derivative is 1: 1-1: 3. Preferably, the molar ratio of the 4-oxo-4H-1-benzopyran-2-carboxylic acid to the base is 1:1 to 1: 5. Preferably the base is selected from DIPEA or triethylamine. Preferably the organic solvent is selected from DMSO, DMF or NMP. The reaction temperature of the condensation reaction is preferably 0 ℃ to 70 ℃.

The third aspect of the invention provides an application of the flavonoid tankyrase 2 inhibitor in preparation of a drug for inhibiting tankyrase 2.

The invention is described in detail below with reference to the figures and specific embodiments.

In the present invention, IC₅₀Test method of value: the inhibitory activity of the compounds on TNKS2 was determined by an enzymatic reaction. Dissolving a compound to be detected in a DMSO solution to prepare a solution with an initial concentration of 200 mu M/L, diluting the solution 5 times each time according to a proportion, and configuring 7 concentration gradients to obtain a solution concentration range of each compound to be detected of 0.0128 mu M/L to 200 mu M/L. The enzymatic reaction was carried out in 96-well plates (Greiner bio-one U-shaped blackboard) at room temperature. TNKS2 at 5nM was incubated with compound and 500nM NAD' in assay buffer (50mM HEPES pH 7.0, 1mM CHAPS) for 1 hour, with four parallel experimental groups being set up simultaneously per concentration gradient. Then, assay buffer (50mM HEPES,0.8M KF and 20mM EDTA in 0.1% BSA) was added to react at room temperature for 1 hour. And finally, adding 20mL of 20% acetophenone ethanol solution and 20mL of 2MKOH to terminate the reaction, chemically converting the unreacted NAD' into a fluorescent substance, and reading the fluorescence intensity under the conditions of an excitation wavelength of 355nm and an absorption wavelength of 450 nm. According to the formula: inhibition rate (sample fluorescence intensity-blank)/(enzyme value fluorescence intensity-blank) the corresponding inhibition rate for each inhibitor was calculated and then half the effective inhibitory concentration was fitted using Graph Pad Prism software.

Example 1

N- (4-acetylphenyl) -4-oxo-4H-benzopyran-2-carboxamide

4-oxo-4H-1-benzopyran-2-carboxylic acid (95mg,0.5mmol) was dissolved in 25ml DMF, HATU (228mg, 0.6mmol) and DIPEA (129mg, 1mmol) were added to a DMF solution containing 4-oxo-4H-1-benzopyran-2-carboxylic acid under ice-bath conditions with stirring, the temperature was slowly raised to room temperature, and 4-aminoacetophenone (81mg, 0.6mmol) was added after 15 minutes. Stirring was continued at room temperature for 6H until the 4-oxo-4H-1-benzopyran-2-carboxylic acid reaction was complete. After completion of the reaction, the reaction solution was poured into ice water. Separating out solid, collecting solid, adding water and 10% Na₂CO₃Saline and ethanol ultrasonic washing. Purification of the solid by flash silica gel chromatography (petroleum ether/ethyl acetate 6:1) afforded the product.

IC₅₀The value was 8.74 uM.

¹H NMR(400MHz,DMSO)δ＝10.94(s,1H),8.06(d,J＝8.0Hz,1H),7.98(q,J＝8.6Hz,4H),7.90(d,J＝7.6Hz,1H),7.83(d,J＝8.5Hz,1H),7.55(t,J＝7.6Hz,1H),6.98(s,1H),2.55(s,3H).¹³C NMR(101MHz,DMSO)δ＝196.73,177.32,158.20,155.34,155.20,141.93,135.15,133.12,129.38,126.23,125.02,123.78,120.36,119.09,111.43,26.59.

The hydrogen and carbon spectra are shown in FIGS. 1 and 2.

Example 2

N' -benzoyl-4-oxo-4H-benzopyran-2-carbohydrazide

4-oxo-4H-1-benzopyran-2-carboxylic acid (95mg,0.5mmol) was dissolved in 25ml DMF and HATU (228mg, 0.6mmol) and DIPEA (129mg, 1mmol) were added to a DMF solution containing 4-oxo-4H-1-benzopyran-2-carboxylic acid under ice bath with stirring, slowly warmed to room temperature and benzoyl hydrazine (82mg, 0.6mmol) was added after 15 min. Stirring was continued at room temperature for 6H until the 4-oxo-4H-1-benzopyran-2-carboxylic acid reaction was complete. After completion of the reaction, the reaction solution was poured into ice water. Separating out solid, collecting solid, adding water and 10% Na₂CO₃Saline and ethanol ultrasonic washing. Purification of the solid by flash silica gel chromatography (petroleum ether/ethyl acetate 4:1) afforded the product.

IC₅₀The value was 3.54 uM.

¹H NMR(400MHz,DMSO)δ11.17(s,1H),10.75(s,1H),8.07(t,J＝7.5Hz,1H),7.92(t,J＝7.6Hz,3H),7.76(d,J＝7.9Hz,1H),7.57(dt,J＝24.0,7.6Hz,4H),6.90(d,J＝6.7Hz,1H).¹³C NMR(101MHz,DMSO)δ＝177.13,165.76,158.73,155.18,154.73,135.28,132.27,132.18,128.68,127.57,126.27,125.07,123.83,118.90,111.32.

The hydrogen and carbon spectra are shown in fig. 3 and 4.

Example 3

N' - (4-methylbenzoyl) -4-oxo-4H-benzopyran-2-carbohydrazide

4-oxo-4H-1-benzopyran-2-carboxylic acid (95mg,0.5mmol) was dissolved in 25ml DMF and HATU (228mg, 0.6mmol) and DIPEA (129mg, 1mmol) were added to a DMF solution containing 4-oxo-4H-1-benzopyran-2-carboxylic acid under ice bath with stirring, slowly warmed to room temperature and 4-methylbenzoyl hydrazine (90mg,0.6mmol) was added after 15 minutes. Stirring was continued at room temperature for 6H until the 4-oxo-4H-1-benzopyran-2-carboxylic acid reaction was complete. After completion of the reaction, the reaction solution was poured into ice water. Separating out solid, collecting solid, adding water and 10% Na₂CO₃Saline and ethanol ultrasonic washing. Purification of the solid by flash silica gel chromatography (petroleum ether/ethyl acetate 4:1) afforded the product.

IC₅₀The value was 1.25 uM.

¹H NMR(400MHz,DMSO)δ＝8.07(d,J＝8.0Hz,1H),7.91(t,J＝7.8Hz,1H),7.83(d,J＝7.8Hz,2H),7.75(d,J＝8.5Hz,1H),7.56(t,J＝7.5Hz,1H),7.33(d,J＝7.8Hz,2H),6.90(s,1H),2.38(s,3H).¹³C NMR(101MHz,DMSO)δ177.18,165.60,158.72,155.22,154.88,142.24,135.30,129.49,129.20,127.63,126.27,125.07,123.85,118.93,111.29,21.12.

The hydrogen and carbon spectra are shown in fig. 5 and 6.

Example 4

N' - (4-chlorobenzoyl) -4-oxo-4H-benzopyran-2-carbohydrazide

4-oxo-4H-1-benzopyran-2-carboxylic acid (95mg,0.5mmol) was dissolved in 25ml DMF and HATU (228mg, 0.6mmol) and DIPEA (129mg, 1mmol) were added to a DMF solution containing 4-oxo-4H-1-benzopyran-2-carboxylic acid under ice bath with stirring, slowly warmed to room temperature and 4-chlorobenzoyl hydrazine (102mg, 0.6mmol) was added after 15 min. Stirring is continued at room temperature for 6h until 4-oxo-4 is obtainedThe H-1-benzopyran-2-carboxylic acid is completely reacted. After completion of the reaction, the reaction solution was poured into ice water. Separating out solid, collecting solid, adding water and 10% Na₂CO₃Saline and ethanol ultrasonic washing. Purification of the solid by flash silica gel chromatography (petroleum ether/ethyl acetate 4:1) afforded the product.

IC₅₀The value was 0.89 uM.

¹H NMR(400MHz,DMSO)δ11.05(s,3H),8.10(d,J＝7.9Hz,1H),7.95(dd,J＝15.1,7.9Hz,3H),7.78(d,J＝8.5Hz,1H),7.65(d,J＝8.1Hz,2H),7.59(t,J＝7.6Hz,1H),6.94(s,1H).¹³C NMR(101MHz,DMSO)δ＝177.14,164.79,158.73,155.19,154.67,137.09,135.30,130.97,129.52,128.86,126.29,125.09,123.85,118.90,111.37.

The hydrogen and carbon spectra are shown in fig. 7 and 8.

Example 5

4-oxo-N- (4- (3,4, 5-trimethoxybenzamido) phenyl) -4H-benzopyran-2-carboxamide

4-oxo-4H-1-benzopyran-2-carboxylic acid (95mg,0.5mmol) was dissolved in 25ml of DMF, HATU (228mg, 0.6mmol) and DIPEA (129mg, 1mmol) were added to a DMF solution containing 4-oxo-4H-1-benzopyran-2-carboxylic acid under ice-bath conditions with stirring, the temperature was slowly raised to room temperature, and N- (4-aminophenyl) -3,4, 5-trimethoxybenzamide (181mg, 0.6mmol) was added after 15 minutes. Stirring was continued at room temperature for 6H until the 4-oxo-4H-1-benzopyran-2-carboxylic acid reaction was complete. After completion of the reaction, the reaction solution was poured into ice water. Separating out solid, collecting solid, adding water and 10% Na₂CO₃Saline and ethanol ultrasonic washing. Purification of the solid by flash silica gel chromatography (petroleum ether/ethyl acetate 4:1) afforded the product.

IC₅₀The value was 0.082 uM.

¹H NMR(400MHz,DMSO)δ10.73(s,1H),10.17(s,1H),8.07(d,J＝7.7Hz,1H),7.93–7.87(m,1H),7.83(d,J＝8.6Hz,1H),7.79(s,4H),7.54(t,J＝7.5Hz,1H),7.28(s,2H),6.97(s,1H),3.87(s,8H),3.73(s,3H).¹³C NMR(101MHz,DMSO)δ＝177.39,164.87,157.55,155.83,155.24,152.71,140.41,136.04,135.09,133.33,130.04,126.18,125.02,123.79,121.50,121.04,119.09,111.08,105.38,60.21,56.20.

The hydrogen and carbon spectra are shown in fig. 9 and 10.

The embodiments described above are intended to facilitate the understanding and use of the invention by those skilled in the art. It will be readily apparent to those skilled in the art that various modifications to these embodiments may be made, and the generic principles described herein may be applied to other embodiments without the use of the inventive faculty. Therefore, the present invention is not limited to the above embodiments, and those skilled in the art should make improvements and modifications within the scope of the present invention based on the disclosure of the present invention.