阅读说明：本技术 具有抑制dna拓扑异构酶活性的喹噁啉-n1,n4-二氧化物衍生物、制备方法及应用 (quinoxaline-N 1, N 4 -dioxide derivative with DNA topoisomerase activity inhibition function, preparation method and application ) 是由 袁宗辉 张鹤营 潘源虎 张洁 瞿玮 谢书宇 陶燕飞 陈冬梅 黄玲利 刘振利 谢长 于 2019-10-18 设计创作，主要内容包括：本发明属于生物化学技术领域,尤其涉及具有抑制DNA拓扑异构酶活性的喹噁啉-N~1,N~4-二氧化物衍生物、制备方法及应用。该类化合物的合成是以4,5-二氟-2-硝基苯胺为原料,在碱性催化剂氢氧化钠的催化下,与次氯酸钠发生反应,得到5,6-二氟-N-氧化苯并呋咱；再与不同的底物发生Beirut反应和取代反应,即得到一系列喹噁啉-N~1,N~4-二氧化物衍生物。本发明所述的喹噁啉-N~1,N~4-二氧化物不仅对以前报道的革兰氏阴性菌具有良好的抑菌活性,对胸膜肺炎放线杆菌及革兰氏阳性菌如上述金黄色葡萄球菌和肺炎链球菌也具有良好的抑菌活性。(the invention belongs to the technical field of biochemistry, and particularly relates to quinoxaline-N 1 and N 4 -dioxide derivatives with activity of inhibiting DNA topoisomerase, a preparation method and application thereof.)

1. quinoxaline-N ¹, N ⁴ -dioxide derivatives having an inhibitory activity on DNA topoisomerase, characterized by having the following general structural formula:

In formula I:

R ₁ is alkyl or hydroxy or alkoxy substituted, R ₂ is methyl or trifluoromethyl, R ₆ is nitrogen-containing heterocycle or fluorine atom substituted, the nitrogen-containing heterocycle is connected with quinoxaline mother ring through C-N bond, R ₇ is hydrogen, chlorine or fluorine atom.

2. The quinoxaline-N ¹, N ⁴ -dioxide derivative with the activity of inhibiting DNA topoisomerase according to claim 1, wherein the nitrogen-containing heterocycle is any one of piperazine, pyrrole, triazole, piperidine, imidazole or morpholine.

3. A method for preparing quinoxaline-N ¹, N ⁴ -dioxide derivatives having an inhibitory activity on DNA topoisomerase, comprising the steps of:

step 1: dissolving 4, 5-difluoro-2-nitroaniline in tetrahydrofuran, taking sodium hydroxide as a catalyst, and reacting with sodium hypochlorite in an ice bath to generate 5, 6-difluoro-N-benzofuroxan;

Step 2, carrying out a Beiriut reaction on 5, 6-difluoro-N-benzofuroxan and ethyl acetoacetate at room temperature to obtain 6,7-2F-3-CH ₃ -2-quinoxaline ethyl formate-N ¹, N ⁴ -dioxide;

and step 3: and (3) carrying out substitution reaction on the product obtained in the step (2) and different nitrogenous heterocyclic substrates to obtain a final product, wherein the chemical structural general formula of the final product is shown in claim 1 or 2.

4. the method for preparing quinoxaline-N ¹, N ⁴ -dioxide derivatives having a DNA topoisomerase activity inhibiting activity according to claim 3, wherein 4, 5-dichloro-2-nitroaniline is substituted for 4, 5-difluoro-2-nitroaniline when the C ₇ position of the quinoxaline ring is substituted by a chlorine atom in the step 1.

5. The method for preparing quinoxaline-N ¹, N ⁴ -dioxide derivatives with DNA topoisomerase activity inhibiting activity according to claim 4, wherein in the step 2, when the C ₂ position of the quinoxaline ring is replaced by acetyl, acetylacetone is selected to replace ethyl acetoacetate to obtain 6,7-2F-3-CH ₃ -2-acetylquinoxaline-N ¹, N ⁴ -dioxide.

6. The method for preparing quinoxaline-N ¹, N ⁴ -dioxide derivatives having DNA topoisomerase activity according to claim 5, wherein the C ₃ position of the quinoxaline ring is substituted by trifluoromethyl in the step 2, and the substrate is ethyl trifluoroacetoacetate instead of ethyl acetoacetate or trifluoroacetylacetone instead of acetylacetone.

7. Use of a quinoxaline-N ¹, N ⁴ -dioxide derivative having DNA topoisomerase activity inhibiting according to claim 1 or 2 for inhibiting the activity of any one or more strains of mycobacterium tuberculosis, staphylococcus aureus, streptococcus pneumoniae or actinobacillus pleuropneumoniae.

8. Use of a quinoxaline-N ¹, N ⁴ -dioxide derivative having DNA topoisomerase activity inhibiting according to claim 1 or 2 in the manufacture of a medicament for the treatment of an infection caused by any one or more of mycobacterium tuberculosis, staphylococcus aureus, streptococcus pneumoniae or actinobacillus pleuropneumoniae.

Technical Field

The invention belongs to the technical field of biochemistry, and particularly relates to quinoxaline-N ¹, N ⁴ -dioxide derivatives with DNA topoisomerase activity inhibition activity, a preparation method and application thereof.

background

The quinoxaline-N ¹, N ⁴ -dioxide are benzo piperazine heterocyclic compounds, have broad-spectrum antibacterial activity and are generally applied to the fields of medicine and agriculture, the antibacterial activity of the quinoxaline-N ¹, N ⁴ -dioxide is firstly researched by the compound (Landquist et al, 1956; Silk, 1956), and particularly have good inhibitory effect on gram-negative bacteria (Wang Xue et al, 2009), such as remarkable antibacterial effect on avian pasteurella, Escherichia coli, salmonella gallinarum and dysentery bacillus (New Aphrough, 2008), meanwhile, the compound can also improve the feed conversion rate and promote the growth and development of animals (Cihak et al, 1983; Cihak et al, 1985), the quinoxaline-N ¹, N ⁴ -dioxide and anti-tumor (Mielcke et al, 2012; Rajule et al, antiviral, antifungal and the like), wherein the carboxin and ethanol are used as medicaments in 20 th century, have good antiviral and antifungal effects on the livestock, antiviral and anti-toxic and anti-diarrhea effects (European drugs, 1988), and veterinary drug for the pig, pig, pig.

Porcine infectious pleuropneumonia is an acute lethal respiratory infectious disease caused by Actinobacillus pleuropneumoniae (APP), is one of the most important bacterial diseases threatening the swine industry of various countries, and is mainly characterized by hemorrhagic cellulosic pleuropneumonia and cellulosic necrotic pleuropneumonia. Actinobacillus pleuropneumoniae is classified into the genus Actinobacillus, the family Pasteobacteriaceae, gram-negative small bacilli. Although the quinoxaline compound has broad-spectrum antibacterial activity, the activity of actinobacillus pleuropneumoniae is not reported, so that the determination of the antibacterial activity of the quinoxaline compound on APP is of great importance, and a foundation is provided for the development of an application approach of the quinoxaline compound.

Bacterial DNA topoisomerases have been extensively studied and applied as targets for antibacterial drugs (Yorgey et al, 1994). DNA topoisomerases control the topology of the double strand of DNA within bacteria or cells and play a crucial role in protein translation and cell replication. During DNA replication, DNA topoisomerase binds to the DNA double strand, removing the topology of the duplex DNA by cleaving either the DNA single or double strand, and then initiating the DNA replication process (sisi et al, 2010; Bush et al, 2015). Bacterial DNA topoisomerases include DNA topoisomerase I and DNA topoisomerase II (dnatop II), wherein DNA TOP II is extensively studied as a target for fluoroquinolones. The DNA TOP II comprises DNA Gyrase (DNA Gyrase) and DNA topoisomerase IV (DNA TOP IV), the two enzymes have high structural homology, the homology similarity is as high as 99%, but the two enzymes play different roles in the DNA replication process, the DNA Gyrase can promote relaxed circular DNA to be converted into negative supercoiled DNA while hydrolyzing ATP, and the DNA TOP IV can promote the negative supercoiled DNA to be converted into relaxed DNA after being unfolded from the topological state under the premise that the ATP provides energy.

Fluoroquinolones (FQs) are broad-spectrum antibacterial drugs and widely applied to various microbial infections, the action targets of the fluoroquinolones are DNA topoisomerase II (DNA Gyrase and DNA TOP IV) of bacteria, FQs is combined to a DNA-topoisomerase complex in the DNA replication process to form an irreversible drug-enzyme-DNA ternary covalent complex, the complex can cause DNA cracking and is also called a cracking complex, the activity of enzymes in the cracking complex is inhibited by FQs, the processes of DNA replication, transcription, translation and the like are prevented, and finally bacteria die (Mustaev et al, 2014). While FQs was used in large quantities, the bacteria gradually developed resistance to FQs; therefore, in order to solve the increasingly severe problem of drug resistance, the development of novel and highly effective antibacterial drugs is urgently needed.

Disclosure of Invention

Aiming at the problems in the prior art, the invention provides quinoxaline-N ¹, N ⁴ -dioxide derivatives with DNA topoisomerase activity inhibition activity, a preparation method and application thereof.

The quinoxaline-N ¹, N ⁴ -dioxide derivative which can inhibit the activity of DNA topoisomerase is characterized by having the following structural general formula:

In formula I:

r ₁ is alkyl or hydroxy or alkoxy substituted, R ₂ is methyl or trifluoromethyl, R ₆ is nitrogen-containing heterocycle or fluorine atom substituted, the nitrogen-containing heterocycle is connected with quinoxaline mother ring through C-N bond, R ₇ is hydrogen, chlorine or fluorine atom.

further, the nitrogen-containing heterocycle is any one of piperazine, pyrrole, triazole, piperidine, imidazole or morpholine.

A method for preparing quinoxaline-N ¹, N ⁴ -dioxide derivatives having an inhibitory activity on DNA topoisomerase, comprising the steps of:

step 1: dissolving 4, 5-difluoro-2-nitroaniline in tetrahydrofuran, taking sodium hydroxide as a catalyst, and reacting with sodium hypochlorite in an ice bath to generate 5, 6-difluoro-N-benzofuroxan;

Step 2, carrying out a Beiriut reaction on 5, 6-difluoro-N-benzofuroxan and ethyl acetoacetate at room temperature to obtain 6,7-2F-3-CH ₃ -2-quinoxaline ethyl formate-N ¹, N ⁴ -dioxide;

And step 3: and (3) carrying out substitution reaction on the product obtained in the step (2) and different nitrogenous heterocyclic substrates to obtain a final product, wherein the chemical structural general formula of the final product is shown as a formula I.

Further, in the step 1, 4, 5-dichloro-2-nitroaniline is used for replacing 4, 5-difluoro-2-nitroaniline when the C ₇ position of the quinoxaline ring is replaced by a chlorine atom.

further, in the step 2, when the position C ₂ of the quinoxaline ring is replaced by acetyl, acetylacetone is selected to replace ethyl acetoacetate to obtain 6,7-2F-3-CH ₃ -2-acetylquinoxaline-N ¹ and N ⁴ -dioxide.

Further, in the step 2, the C ₃ position of the quinoxaline ring is replaced by trifluoromethyl, and the substrate is ethyl trifluoroacetoacetate instead of ethyl acetoacetate or trifluoroacetylacetone instead of acetylacetone.

The quinoxaline-N ¹, N ⁴ -dioxide derivative with the activity of inhibiting DNA topoisomerase is applied to inhibiting the activity of any one or more strains of mycobacterium tuberculosis, staphylococcus aureus, streptococcus pneumoniae or actinobacillus pleuropneumoniae.

use of a quinoxaline-N ¹, N ⁴ -dioxide derivative having DNA topoisomerase inhibitory activity as described above in the manufacture of a medicament for the treatment of an infection caused by any one or more of mycobacterium tuberculosis, staphylococcus aureus, streptococcus pneumoniae or a. pleuropneumoniae.

In summary, the advantages and positive effects of the invention are:

^{1 4}The research of the applicant of the application finds that quinoxaline compounds have DNA TOP IV inhibiting activity, a series of quinoxaline-N1, N4-dioxide derivatives shown in the application are synthesized by carrying out side chain modification on positions C2, C3, C6 and C7 of a parent ring of the quinoxaline compounds based on the FQs structure, and the result of the research on the DNA TOP IV inhibiting activity of the newly synthesized compounds shows that the compounds have obvious inhibiting activity on Escherichia coli DNA TOP IV.

Drawings

FIG. 1 shows the results of inhibition of the enzyme activity of DNA topoisomerase IV by Compound 8;

FIG. 2 shows the results of the inhibition of the enzyme activity of DNA topoisomerase IV by Compound 9;

FIG. 3 shows the results of inhibition of the enzyme activity of DNA topoisomerase IV by Compound 2;

FIG. 4 shows the results of inhibition of the enzyme activity of DNA topoisomerase IV by Compound 6;

fig. 5 is a general chemical molecular structural formula of quinoxaline-N ¹, N ⁴ -dioxide derivatives in the present application.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail below with reference to examples, and the equipment and reagents used in the examples and test examples are commercially available without specific reference. The specific embodiments described herein are merely illustrative of the invention and are not intended to be limiting.

The invention discloses quinoxaline-N ¹, N ⁴ -dioxide derivatives with DNA topoisomerase activity inhibition, a preparation method and application thereof, wherein the quinoxaline-N ¹, N ⁴ -dioxide derivatives with antibacterial activity and DNA TOP IV inhibition activity, which are prepared by the invention, have the following chemical molecular structure general formula and are shown in figure 5.

In formula I:

R ₁ is alkyl, hydroxyl or alkoxy substituted, R ₂ is methyl or trifluoromethyl, R ₆ is nitrogen-containing heterocycle or fluorine atom substituted, the nitrogen-containing heterocycle is preferably selected from piperazine, pyrrole, triazole, piperidine, imidazole, morpholine and the like, the nitrogen-containing heterocycle is connected with the quinoxaline ring through a C-N bond, and R ₇ is hydrogen, chlorine or fluorine atom.