ON-DNA 3, 4-substituted 3, 4-dihydroquinoxaline-2 (1H)-ketone compound and preparation method thereof

Abstract

The invention provides an ON-DNA 3, 4-substituted 3, 4-dihydroquinoxaline-2 (1H)-ketone compound and a preparation method thereof, and relates to the technical field of coding compound libraries. According to the invention, by controlling the conditions of solvent, temperature, pH and the like during reaction, the ON-DNA halogenated nitrobenzene compound can be used for preparing the ON-DNA 3, 4-substituted 3, 4-dihydroquinoxaline-2 (1H)-ketone compound through a two-step method. The method can be carried out in a mixed aqueous phase of an organic solvent / aqueous phase, has the advantages of single product, simple operation and environmental friendliness, and is suitable for synthesis of a DNA coding compound library by using a porous plate.

Description

Technical Field

[0001] This invention relates to the field of encoded compound library technology, and more particularly to an ON-DNA 3,4-dihydroquinoxaline-2(1H)-one compound and its preparation method. Background Technology

[0002] DNA-encoded library (DEL) technology is a revolutionary drug discovery technique that uses DNA as a molecular barcode to label and track massive amounts of compounds. As an emerging small-molecule drug screening method, it has become an indispensable technological platform in new drug development. On-DNA (ON-DNA) chemical reactions are of great significance for constructing DELs with rich chemical space and structural diversity. In recent years, the number of ON-DNA chemical reactions has been increasing, greatly expanding the range of chemical reactions that can be used for DEL construction.

[0003] ON-DNA chemical reactions are essential for constructing and modifying DNA-encoded compound libraries. These reactions must be carried out efficiently and specifically under mild, aqueous conditions while maintaining the stability and decodeability of DNA. ON-DNA reactions form the chemical basis for constructing large and diverse DEL libraries. Successful screening using DEL technology relies on high-quality compound libraries successfully constructed using ON-DNA reactions in the past. Without widely applicable, efficient, and reliable ON-DNA chemical reactions, DEL technology cannot realize its enormous potential.

[0004] Quinoxalone compounds, as drug scaffold structures, possess potential and broad application value. Introducing quinoxalone scaffolds into DNA-encoded compound libraries could expand their diversity and increase the probability of screening effective compounds. However, no method has been reported for constructing 3,4-dihydroquinoxalon-2(1H)-one compounds with 3,4-substituted positions on ON-DNA using halogenated nitrobenzene compounds. Therefore, a new method for synthesizing 3,4-dihydroquinoxalon-2(1H)-one compounds with 3,4-substituted positions on ON-DNA is needed for high-volume multi-well plate operations. This would increase the diversity of DNA-encoded compound libraries and further enhance the application value of DNA-encoded compound library technology.

[0005] Therefore, this invention is proposed. Summary of the Invention

[0006] To address the aforementioned technical problems, this invention provides ON-DNA 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-position substitution and their preparation method. The synthetic method used in this invention features mild reaction conditions, stable raw materials, high conversion rate, good substrate versatility, and minimal DNA damage. It is suitable for the batch synthesis of DNA-encoded compound libraries using multi-well plates and can rapidly convert DNA-encoded halonitrobenzene compound libraries into DNA-encoded 3,4-dihydroquinoxaline-2(1H)-one compound libraries through a two-step reaction.

[0007] To achieve the objectives of this invention, a method for synthesizing ON-DNA 3,4-dihydroquinoxaline-2(1H)-ketone compounds with 3,4-position substitution is provided, comprising the following steps: S1. Synthesis of ON-DNA halonitrobenzene compounds; S2. Dissolve the ON-DNA halonitrobenzene compound in borate buffer, then add secondary amine amino acid compound and base in sequence, and react at 55℃-65℃ for 1h-12h. After the reaction is completed, precipitate with ethanol, discard the supernatant, and obtain a solid. S3. Dissolve the solid in deionized water, add sodium dithionite, and react for 1-12 hours. After the reaction is complete, precipitate with ethanol and discard the supernatant to obtain 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA positions.

[0008] Furthermore, the structural formula of the ON-DNA halonitrobenzene compound is as follows: ; The structural formula of the secondary amine amino acid compound is as follows: ; The structural formula of the 3,4-dihydroquinoxaline-2(1H)-one compound with 3,4-substituted ON-DNA is as follows: ; Wherein, X is any one of F, Cl, Br or I; R1 and R2 are groups with a molecular weight of less than 1000.

[0009] Furthermore, R1 and R2 are selected from any one of alkyl, substituted alkyl, aromatic or substituted aromatic groups; The alkyl group is C1-C20 or C3-C8 cycloalkyl.

[0010] Furthermore, the alkyl group is a straight-chain or branched C1-C12 alkyl group or a C3-C6 saturated cycloalkyl group; Furthermore, the alkyl group is selected from any one of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, or hexyl. Furthermore, the saturated cycloalkyl group is selected from any one of cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl.

[0011] The number of substituents in the substituted alkyl group is one or more; The substituents of the substituted alkyl group are independently selected from any one or more of alkyl, halogen, nitro, alkoxy, halophenyl, phenyl, and alkylphenyl groups; Furthermore, the alkyl group is selected from any one of cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl; Furthermore, the alkylphenyl group is selected from any one of methylphenyl, ethylphenyl, n-propylphenyl, isopropylphenyl, n-butylphenyl, isobutylphenyl, tert-butylphenyl, pentylphenyl, or hexylphenyl; Furthermore, the halophenyl group is selected from any one of fluorophenyl, chlorophenyl, bromophenyl, or iodophenyl.

[0012] The aromatic group is selected from any one of pyridyl, quinolinyl, thiazolyl, thiophenyl or phenyl; The number of substituents of the substituted aromatic group is one or more; Furthermore, the aromatic group is phenyl.

[0013] The substituents of the substituted aromatic group are independently selected from one or more of halogen, cyano, nitro, alkoxy, aminoacyl, alkyl, or C1-C20 alkyl groups.

[0014] Furthermore, the substituent of the aromatic group is any one of C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkanoyl or alkylaminoyl; Furthermore, the C1-C6 alkyl group is selected from any one of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, or hexyl. Furthermore, the C1-C6 alkoxy groups are selected from any one of methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, pentoxy, or hexoxy. Furthermore, the alkylaminoyl group is selected from any one of methylaminoyl, ethaminoyl, n-alanaminoyl, isoalanaminoyl, n-butaminoyl, isobutaminoyl, tert-butaminoyl, pentaminoyl, or hexaminoyl; Furthermore, the C1-C6 alkyl acyl group is selected from any one of methane acyl, ethane acyl, n-propane acyl, isopropane acyl, n-butane acyl, isobutane acyl, tert-butane acyl, pentane acyl, or hexane acyl.

[0015] Furthermore, the DNA molecule in the structural formula comprises a double-stranded nucleotide chain obtained by polymerizing artificially modified and / or unmodified nucleotide monomers, the nucleotide chain being linked to a halonitrobenzene via an amide bond.

[0016] Furthermore, the base is any one or more of sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, N,N-diisopropylethylamine, or triethylamine.

[0017] Furthermore, the reaction time in step S2 is 2 hours; The reaction time in step S3 is 2 hours.

[0018] Furthermore, the ON-DNA halonitrobenzene compound has a molar equivalent of 1 and a molar concentration of 0.5 mM-2 mM; The secondary amine amino acid compound has a molar equivalent of 200-400 times; The base has a molar equivalent of 200-400 times; The sodium dithionite has a molar equivalent of 200-400 times.

[0019] Furthermore, the secondary amine amino acid compound has a molar equivalent of 200, 300, or 400 times; Furthermore, the base has a molar equivalent of 200, 300, or 400 times; Furthermore, the sodium dithionite has a molar equivalent of 200, 300, or 400 times.

[0020] Furthermore, the reaction is carried out in solution, which is one or more aqueous mixed solvents selected from acetonitrile, methanol, ethanol, DMF, DMA, DMSO, THF, water, inorganic salt buffer, organic acid buffer, and organic base buffer.

[0021] The present invention also provides a 3,4-dihydroquinoxaline-2(1H)-one compound with 3,4-substituted ON-DNA obtained by the above method.

[0022] The present invention also provides an application of the above method in batch operations of perforated plates.

[0023] The present invention also provides an application of the above method in the synthesis of DNA-encoded compound libraries in multi-well plates.

[0024] The embodiments of the present invention have the following technical effects: This invention provides a method for synthesizing 3,4-dihydroquinoxaline-2(1H)-one compounds substituted at the 3,4 positions of ON-DNA. The method uses ON-DNA halonitrobenzene compounds and secondary amine amino acid compounds as raw materials, and obtains the ON-DNA substituted 3,4-dihydroquinoxaline-2(1H)-one compounds through a two-step reaction of SNAr and intramolecular reductive amination under alkaline and sodium dithionite conditions.

[0025] This reaction is characterized by mild reaction conditions, stable raw materials, high conversion rate, good substrate versatility, and minimal DNA damage. It is suitable for the synthesis of DNA-encoded compound libraries in batches using multi-well plates. It can rapidly convert DNA-encoded halonitrobenzene compound libraries into DNA-encoded 3,4-dihydroquinoxaline-2(1H)-one compound libraries through a two-step reaction, further increasing the diversity of DNA-encoded compound libraries and enhancing the application value of DNA-encoded compound library technology. Attached Figure Description

[0026] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0027] Figure 1 The conversion rate distribution of 11 ON-DNA 3,4-dihydroquinoxaline-2(1H)-one compounds obtained in this invention; Figure 2 Example 2: Liquid chromatogram of the reaction product; Figure 3 Mass spectrum of the reaction product in Example 2; Figure 4 Example 3: Liquid chromatogram of the reaction product; Figure 5 Mass spectrum of the reaction product in Example 3; Figure 6 Example 4: Liquid chromatogram of the reaction product; Figure 7 Mass spectrum of the reaction product in Example 4; Figure 8 : Liquid chromatogram of the reaction product in Example 5; Figure 9 Mass spectrum of the reaction product in Example 5; Figure 10 Example 6: Liquid chromatogram of the reaction product; Figure 11 Mass spectrum of the reaction product in Example 6; Figure 12 Example 7: Liquid chromatogram of the reaction product; Figure 13 Mass spectrum of the reaction product in Example 7; Figure 14 Example 8: Liquid chromatogram of the reaction product; Figure 15 Mass spectrum of the reaction product in Example 8; Figure 16 Example 9: Liquid chromatogram of the reaction product; Figure 17 Mass spectrum of the reaction product in Example 9; Figure 18 Example 10: Liquid chromatogram of the reaction product; Figure 19 Mass spectrum of the reaction product in Example 10; Figure 20 Example 11: Liquid chromatogram of the reaction product; Figure 21 Mass spectrum of the reaction product in Example 11; Figure 22 Example 12: Liquid chromatogram of the reaction product; Figure 23 Mass spectrum of the reaction product in Example 12. Detailed Implementation

[0028] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of this invention, and not all of them. Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this invention.

[0029] In a first aspect, the present invention provides a method for synthesizing 3,4-dihydroquinoxaline-2(1H)-ketone compounds substituted at the 3,4 positions of ON-DNA, comprising the following steps: S1. Synthesis of ON-DNA halonitrobenzene compounds; S2. Dissolve the ON-DNA halonitrobenzene compound in borate buffer, then add secondary amine amino acid compound and base in sequence, and react at 55℃-65℃ for 1h-12h. After the reaction is completed, precipitate with ethanol, discard the supernatant, and obtain a solid. S3. Dissolve the solid in deionized water, add sodium dithionite, and react for 1-12 hours. After the reaction is complete, precipitate with ethanol and discard the supernatant to obtain 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA positions.

[0030] In some embodiments, the structural formula of the ON-DNA halonitrobenzene compound is as follows: ; The structural formula of the secondary amine amino acid compound is as follows: ; The structural formula of the 3,4-dihydroquinoxaline-2(1H)-one compound with 3,4-substituted ON-DNA is as follows: ; Wherein, X is any one of F, Cl, Br or I; R1 and R2 are groups with a molecular weight of less than 1000.

[0031] In some embodiments, R1 and R2 are selected from any one of alkyl, substituted alkyl, aromatic or substituted aromatic groups; The alkyl group is C1-C20 or C3-C8 cycloalkyl.

[0032] In some embodiments, the alkyl group is a straight-chain or branched C1-C12 alkyl group or a C3-C6 saturated cycloalkyl group; In some embodiments, the alkyl group is selected from any one of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, or hexyl. In some embodiments, the saturated cycloalkyl group is selected from any one of cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl.

[0033] The number of substituents in the substituted alkyl group is one or more; The substituents of the substituted alkyl group are independently selected from any one or more of alkyl, halogen, nitro, alkoxy, halophenyl, phenyl, and alkylphenyl groups; In some embodiments, the alkyl group is selected from any one of cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl; In some embodiments, the alkylphenyl is selected from any one of methylphenyl, ethylphenyl, n-propylphenyl, isopropylphenyl, n-butylphenyl, isobutylphenyl, tert-butylphenyl, pentylphenyl or hexylphenyl; In some embodiments, the halophenyl group is selected from any one of fluorophenyl, chlorophenyl, bromophenyl, or iodophenyl.

[0034] The aromatic group is selected from any one of pyridyl, quinolinyl, thiazolyl, thiophenyl or phenyl; The number of substituents of the substituted aromatic group is one or more; In some embodiments, the aromatic group is phenyl.

[0035] The substituents of the substituted aromatic group are independently selected from one or more of halogen, cyano, nitro, alkoxy, aminoacyl, alkyl, or C1-C20 alkyl groups.

[0036] In some embodiments, the substituent of the aromatic group is any one of C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkanoyl or alkylaminoyl; In some embodiments, the C1-C6 alkyl group is selected from any one of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, or hexyl. In some embodiments, the C1-C6 alkoxy groups are selected from any one of methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, pentoxy, or hexoxy. In some embodiments, the alkylaminoyl group is selected from any one of methylaminoyl, ethaminoyl, n-alanaminoyl, isoalanaminoyl, n-butaminoyl, isobutaminoyl, tert-butaminoyl, pentaminoyl, or hexaminoyl; In some embodiments, the C1-C6 alkyl group is selected from any one of methaneyl, ethaneyl, n-propaneyl, isopropaneyl, n-butaneyl, isobutaneyl, tert-butaneyl, pentaneyl, or hexaneyl.

[0037] In some embodiments, the DNA molecule in the structure comprises a double-stranded nucleotide chain obtained by polymerizing artificially modified and / or unmodified nucleotide monomers, the nucleotide chain being linked to a halonitrobenzene via an amide bond.

[0038] In some embodiments, the base is any one or more of sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, N,N-diisopropylethylamine, or triethylamine.

[0039] In some embodiments, the reaction time in step S2 is 2 hours; The reaction time in step S3 is 2 hours.

[0040] In some embodiments, the ON-DNA halonitrobenzene compound has a molar equivalent of 1 and a molar concentration of 0.5 mM to 2 mM; The secondary amine amino acid compound has a molar equivalent of 200-400 times; The base has a molar equivalent of 200-400 times; The sodium dithionite has a molar equivalent of 200-400 times.

[0041] In some embodiments, the secondary amine amino acid compound is present in a molar equivalent of 200, 300, or 400 times; In some embodiments, the base is in a molar equivalent of 200, 300, or 400 times; In some embodiments, the sodium dithionite is in a molar equivalent of 200, 300, or 400 times.

[0042] In some embodiments, the reaction is carried out in solution, which is one or more aqueous mixed solvents selected from acetonitrile, methanol, ethanol, DMF, DMA, DMSO, THF, water, inorganic salt buffer, organic acid buffer, and organic base buffer.

[0043] Secondly, the present invention also provides a 3,4-dihydroquinoxaline-2(1H)-one compound with 3,4-substituted ON-DNA obtained by the above method.

[0044] Thirdly, the present invention also provides an application of the above method in batch operations of perforated plates.

[0045] Fourthly, the present invention also provides an application of the above method in the synthesis of DNA-encoded compound libraries in multi-well plates.

[0046] The following is a detailed explanation using specific embodiments: Example 1: Synthesis of ON-DNA Halonitrobenzene Compounds All raw materials and equipment used in this invention are known products, obtained by purchasing commercially available products; in this invention, "room temperature" refers to 20-25°C; HATU: 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate; DIPEA: N,N-diisopropylethylamine; DMSO: dimethyl sulfoxide; in this invention, DNA-NH2 is a DNA structure with a -NH2 linker formed by double-stranded DNA and a linker group, such as the DNA-NH2 structure of "compound 1" in WO2005058479.

[0047] 400 molar equivalents of o-fluoronitrobenzic acid (400 mM, dissolved in DMSO) were mixed with 400 molar equivalents of HATU (400 mM, dissolved in DMSO) and 400 molar equivalents of DIPEA (400 mM, dissolved in DMSO), respectively. The mixture was thoroughly mixed by vortexing. After 5 min, the mixture was added to DNA-NH2 solution (1 mM concentration; borate buffer, pH 9.4, 250 mM solution). The reaction solution was then thoroughly mixed by vortexing and reacted at room temperature for 2 h.

[0048] After the reaction is complete, ethanol precipitation is performed: 10% of the total volume of 5M sodium chloride solution is added to the solution, followed by 3 times the total volume of anhydrous ethanol. After shaking and mixing, the reaction is placed in a -80℃ freezer for 2 hours, then centrifuged at 13000 ×g for half an hour. The supernatant is discarded and used directly for the next step of the reaction.

[0049] Example 2: Synthesis of 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA positions Step 1: Put Dissolve in borate buffer (0.5M, pH=9.4) to prepare a 1mM concentration solution (10μL, 10nmol). Add N-methylglycine (400 molar equivalent, 400mM aqueous solution) and DIPEA (400 molar equivalent, 400mM DMSO solution) to the solution in sequence and react at 60℃ for 2h. After the reaction is complete, ethanol precipitation is performed: 10% of the total volume of 5M sodium chloride solution is added to the solution, followed by 3 times the total volume of anhydrous ethanol. After shaking and mixing, the reaction is placed in a -80℃ freezer for 2 hours, then centrifuged at 13000 ×g for half an hour. The supernatant is discarded and used directly for the next step of the reaction.

[0050] Step 2: Dissolve in water to prepare a 1 mM concentration solution (10 μL, 10 nmol), add Na2S2O4 (400 molar equivalent, 400 mM H2O solution) to the solution, and react at room temperature for 4 h; After the reaction was complete, ethanol precipitation was performed: 10% (total volume) of 5M sodium chloride solution was added to the solution, followed by 3 times the total volume of anhydrous ethanol. After shaking and mixing, the reaction mixture was placed in a -80°C freezer for 0.5 hours. Then, it was centrifuged at 13000 × g for 15 minutes. The supernatant was discarded, and the remaining precipitate was dissolved in deionized water to obtain... The solution was sent to LCMS to confirm that the reaction conversion rate was 86%.

[0051] In summary, this invention, by controlling the solvent, temperature, pH, and other conditions during the reaction, enables a two-step method to obtain 3,4-dihydroquinoxalin-2(1H)-one compounds substituted at the 3,4 positions of ON-DNA from halogenated nitrobenzene compounds. This method can be carried out in a mixed aqueous phase of organic solvent / aqueous phase, yields a single product, is simple to operate, environmentally friendly, and suitable for the synthesis of DNA-encoded compound libraries using multi-well plates.

[0052] In Examples 2-12, reaction steps 1 and 2 are the same, and the substrates used in the reactions are all... Only the secondary amine, the substrate participating in the first step of the reaction, is changed, specifically: The reaction equations for Examples 3-12 are shown below.

[0053] Example 3 Conversion rate: 72% Example 4 Conversion rate: 73% Example 5 Conversion rate: 73% Example 6 Conversion rate: 76% Example 7: Conversion rate: 90% Example 8: Conversion rate: 91% Example 9: Conversion rate: 75% Example 10: Conversion rate: 71% Example 11: Conversion rate: 89% Example 12: Conversion rate: 79% Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the technical solutions of the embodiments of the present invention.

Claims

1. A method for synthesizing 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA, characterized in that, Includes the following steps: S1. Synthesis of ON-DNA halonitrobenzene compounds; S2. Dissolve the ON-DNA halonitrobenzene compound in borate buffer, then add secondary amine amino acid compound and base in sequence, and react at 55℃-65℃ for 1h-12h. After the reaction is completed, precipitate with ethanol, discard the supernatant, and obtain a solid. S3. Dissolve the solid in deionized water, add sodium dithionite, and react for 1-12 hours. After the reaction is complete, precipitate with ethanol and discard the supernatant to obtain 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA positions.

2. The method for producing 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA according to claim 1, characterized in that, The structural formula of the ON-DNA halonitrobenzene compound is as follows: ; The structural formula of the secondary amine amino acid compound is as follows: ; The structural formula of the 3,4-dihydroquinoxaline-2(1H)-one compound with 3,4-substituted ON-DNA is as follows: ; Wherein, X is any one of F, Cl, Br or I; R1 and R2 are groups with a molecular weight of less than 1000.

3. The method for producing 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA according to claim 2, characterized in that, R1 and R2 are selected from any one of alkyl, substituted alkyl, aromatic or substituted aromatic groups; The alkyl group is C1-C20 or C3-C8 cycloalkyl; The number of substituents in the substituted alkyl group is one or more; The substituents of the substituted alkyl group are independently selected from any one or more of alkyl, halogen, nitro, alkoxy, halophenyl, phenyl, and alkylphenyl groups; The aromatic group is selected from any one of pyridyl, quinolinyl, thiazolyl, thiophenyl or phenyl; The number of substituents of the substituted aromatic group is one or more; The substituents of the substituted aromatic group are independently selected from one or more of halogen, cyano, nitro, alkoxy, aminoacyl, alkyl, or C1-C20 alkyl groups.

4. The method for producing 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA according to claim 2, characterized in that, The DNA molecule in the structural formula comprises a double-stranded nucleotide chain obtained by polymerizing artificially modified and / or unmodified nucleotide monomers, the nucleotide chain being linked to a halonitrobenzene via an amide bond.

5. The method for producing 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA according to claim 1, characterized in that, The base is any one or more of sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, N,N-diisopropylethylamine, or triethylamine.

6. The method for producing 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA according to claim 1, characterized in that, The reaction time in step S2 is 2 hours. The reaction time in step S3 is 2 hours.

7. The method for producing 3,4-dihydroquinoxaline-2(1H)-one compounds with 3,4-substituted ON-DNA according to claim 1, characterized in that, The ON-DNA halonitrobenzene compound has a molar equivalent of 1 and a molar concentration of 0.5 mM-2 mM; The secondary amine amino acid compound has a molar equivalent of 200-400 times; The base has a molar equivalent of 200-400 times; The sodium dithionite has a molar equivalent of 200-400 times.

8. The ON-DNA 3,4-dihydroquinoxaline-2(1H)-one compound prepared by any one of the methods for synthesizing ON-DNA 3,4-substituted 3,4-dihydroquinoxaline-2(1H)-one compounds as described in any one of claims 1-7.

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