Nitro thienyl phenyl oxygen thioether compound and application thereof

Abstract

The invention relates to a nitro thienyl phenyl oxygen thioether compound and application thereof. The invention aims to provide the compound as shown in the formula (I) or pharmaceutically acceptable salts, solvates or prodrugs thereof, so that the selection range of drugs for treating inflammatory bowel diseases is remarkably widened, clear scientific basis and direction are provided for subsequent development of drugs for treating inflammatory bowel diseases, and the compound has important clinical significance.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine and relates to a nitrothiophene phenyl oxysulfide compound and its applications. Background Technology

[0002] When an organism is infected by external pathogens or suffers internal damage, the acute inflammatory response usually activates the innate immune system, producing inflammatory mediators such as various pro-inflammatory factors and chemokines, activating immune cells, and clearing pathogens or damaged cells. A healthy inflammatory response is one of the important ways the body defends itself, helping people prevent damage and restore a healthy state. However, if acute inflammation fails to eliminate the inflammatory triggers, the persistent inflammatory state will develop into chronic inflammation. The negative effects of inflammation are widespread and pose serious threats to people's health. As an important component of the innate immune system, increased secretion of type I interferon under physiological or pathological conditions can activate the immune system to regulate various inflammatory responses. Under normal physiological conditions, it is one of the key links in the organism's fight against the invasion of external pathogens and its own abnormal life activities; while under pathological conditions, its dysfunction or low response can lead to infectious diseases and even malignant tumors, and chronic persistent activation is one of the important triggers for many autoimmune diseases.

[0003] Innate immunity is a crucial innate defense mechanism for organisms to resist external infections and maintain homeostasis. At the heart of this mechanism lies the pattern recognition receptor (PRR), which specifically recognizes pathogen-associated molecular patterns (PAMPs) or damage-associated molecular patterns (DAMPs). PAMPs primarily originate from pathogens (such as lipopolysaccharides, cytoplasmic DNA, and RNA), while DAMPs are mostly released by damaged or dead cells (such as ATP and HMGB1). Upon recognizing PAMPs / DAMPs, the PRR activates downstream signaling pathways, triggering immune cell activation and immune responses. Inducing host cells to produce type I interferons (such as IFN-β) is one of the key mechanisms for activating the innate immune response and mediating host defense.

[0004] Inflammatory bowel disease (IBD) primarily includes Crohn's disease (CD) and ulcerative colitis (UC). The treatment goals are to induce and maintain clinical remission and mucosal healing, thereby improving patients' quality of life. Current treatment options include traditional medications (such as aminosalicylic acid, glucocorticoids, and immunosuppressants), biologics (such as anti-TNF-α, anti-integrin, and anti-IL-12 / 23 drugs), and small molecule drugs (such as JAK inhibitors). In recent years, the use of biologics and small molecule drugs has significantly improved efficacy in moderate to severe cases, and treatment strategies are gradually shifting towards "target-to-target" (T2T) therapy, emphasizing mucosal healing as a key endpoint.

[0005] However, IBD treatment still faces many challenges: First, the disease is highly heterogeneous, with significant individual differences in efficacy; some patients respond poorly or not at all to existing drugs, necessitating the exploration of predictive biomarkers. Second, long-term use of immunosuppressive drugs may lead to safety issues such as infections and malignancies. Third, treatment options are costly, increasing the economic burden on healthcare. Furthermore, patients' lack of understanding of the disease and their mental health issues are often overlooked. Future research should focus on precision medicine, novel drug development (such as stem cell therapy and microbiome intervention), and optimization of comprehensive management to overcome current difficulties.

[0006] P005091 (CAS: 882257-11-6) is a dual-target inhibitor of USP7 / 47, with an IC50 of USP7. 50 The value was 4.2 μM. Studies have reported that the mechanism of action of thiophene sulfide USP7 inhibitors involves covalent modification of the cysteine ​​residue at position 233 of USP7, inducing a conformational change in the enzyme associated with the active site. Because USP7 plays an important role in various carcinogenic pathways, its inhibitor P005091 is often used as a tool in anticancer drug research; however, its therapeutic activity in inflammatory bowel disease is rarely reported. Summary of the Invention

[0007] The purpose of this invention is to provide compounds as shown in formula (I) or their pharmaceutically acceptable salts, solvates or prodrugs, which significantly broaden the range of drugs for inflammatory bowel disease and provide a clear scientific basis and direction for the development of subsequent drugs for inflammatory bowel disease, and have important clinical significance.

[0008] To solve the above-mentioned technical problems, the present invention is achieved through the following technical solution.

[0009] The first aspect of this invention provides a nitrothiophene phenyloxysulfide compound or a pharmaceutically acceptable salt, solvate, or prodrug thereof, comprising tautomers, cis or trans isomers, meso compounds, racemic compounds, enantiomers, diastereomers, or mixtures thereof, wherein the structure of the nitrothiophene phenyloxysulfide compound is shown in formula (I):

[0010] (I);

[0011] Where R 1 It is selected from one of hydrogen, halogen, alkyl, alkoxy, haloalkyl, and amino;

[0012] R 2 It is selected from one of hydrogen, halogen, alkyl, alkoxy, haloalkyl, and amino;

[0013] R 3 Selected from one of hydrogen, alkyl, alkoxy, and alkylamino;

[0014] R 4 Selected from hydrogen and halogen;

[0015] R 5 Selected from , , , , , , , , One of them;

[0016] Among them G 1 Selected from hydrogen, hydroxyl, alkyl, alkenyl, alkynyl, alkoxy, -NH(CH2)4, -NHC2H4NH2, -NHC2H4NHCOOC(CH3)3, 2-acrylamidomethyl (-CH2NHCOCH=CH2), , , One of the following; the alkyl, alkenyl or alkynyl group is either unsubstituted or substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl and heteroaryl;

[0017] J 1 Selected from one of alkyl, phenyl, benzoylmethyl (-CH2COPh), acetylmethyl (-CH2COCH3), carboxymethyl (-CH2COOH), and (N,N-dimethylcarbamoyl)methyl (-CH2CON(CH3)2);

[0018] K 1 Selected from hydrogen, carboxycarboxamide group (-CH2CONH2) One of them, K 2It is selected from alkyl, hydroxy, alkoxy, and amino; the alkyl or amino group is not substituted or is substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl.

[0019] Q 1 Selected from , , , One of them;

[0020] T 1 It is selected from alkyl, amino, N-(2-aminoethyl)amino (-NHCH2CH2NH2), -NH-nBu; the alkyl or amino group is not substituted or is substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl and heteroaryl;

[0021] U 1 It is selected from one of alkyl, halogen, alkoxy, and amino; the alkyl or amino group is not substituted or is substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl.

[0022] W 1 Selected from alkyl, , One of the following; the alkyl and amino groups are either unsubstituted or substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl;

[0023] X 1 Selected from alkyl, alkoxy, amino, One of the following; wherein the alkyl, alkoxy, and amino groups are unsubstituted or substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl;

[0024] Y 1 Selected from , , One of them;

[0025] R 6 Selected from H, alkyl, and alkylene groups;

[0026] R 5 and R 6 These are either independently substituted substituents or connected to form a five-membered ring or a six-membered ring;

[0027] X is selected from O and S;

[0028] The compound of formula (I) is not one of the following compounds:

[0029] .

[0030] Preferably, the R 5 Selected from , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , One of them.

[0031] Preferably, the alkyl group is C10. 1-6 alkyl.

[0032] Preferably, the alkylene group is C64. 1-6 Alkylene.

[0033] Preferably, the sub-alkenyl group is C 2-6 Alkenyl group.

[0034] Preferably, the acetylenoid group is C. 2-6 Alynyl group.

[0035] Preferably, the alkoxy group is C. 1-6 Alkyl group.

[0036] Preferably, the alkyl haloide is C10. 1-6 Halogenated alkyl groups.

[0037] Preferably, the hydroxyalkyl group is C 1-6 Hydroxyalkyl.

[0038] Preferably, the cycloalkyl group is a 3- to 6-membered cycloalkyl group.

[0039] Preferably, the aryl group is a 6- to 10-membered aryl group.

[0040] Preferably, the heteroaryl group is a 5- to 10-membered heteroaryl group.

[0041] Preferably, the nitrothiophene phenyl oxysulfide compound is selected from one or more of the following structural formulas:

[0042] , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , .

[0043] A second aspect of the present invention provides a pharmaceutical composition comprising the above-described nitrothiophene phenyl oxythioether compounds or pharmaceutically acceptable salts, solvates or prodrugs thereof, including tautomers, cis or trans isomers, meso compounds, racemic compounds, enantiomers, diastereomers or mixtures thereof, and one or more active ingredients for treating inflammatory bowel disease.

[0044] Preferably, the active ingredient for treating inflammatory bowel disease is selected from one or more of mesalazine, budesonide, prednisone, azathioprine, methotrexate, infliximab, adalimumab, vedozizumab, ustekinumab, gusejinumab, tofacitinib, and utpatinib.

[0045] Preferably, the inflammatory bowel disease is selected from one or more of Crohn's disease and ulcerative colitis.

[0046] A third aspect of the present invention provides a pharmaceutical formulation comprising the above-described nitrothiophene phenyl oxysulfide compounds or their pharmaceutically acceptable salts, solvates or prodrugs, including tautomers, cis or trans isomers, meso compounds, racemates, enantiomers, diastereomers or mixtures thereof, and one or more pharmaceutically acceptable carriers.

[0047] Preferably, the pharmaceutically acceptable carrier includes one or more of the following: solvent, filler, binder, lubricant, disintegrant, preservative, colorant, chelating agent, antioxidant, flavoring agent, and surfactant.

[0048] Preferably, the dosage form of the pharmaceutical preparation is selected from one or more of tablets, lozenges, capsules, granules, powders, solutions, suspensions, emulsions, gels, aerosols, and sprays.

[0049] The fourth aspect of the present invention provides the use of the above-mentioned nitrothiophene phenyl oxythioether compounds or pharmaceutically acceptable salts, solvates or prodrugs thereof, including their tautomers, cis or trans isomers, meso compounds, racemic compounds, enantiomers, diastereomers or mixtures thereof, in the preparation of products for inhibiting inflammatory bowel disease.

[0050] Preferably, the inflammatory bowel disease is selected from one or more of Crohn's disease and ulcerative colitis.

[0051] The compositions of the present invention can be formulated using one or more pharmaceutically acceptable carriers according to conventional methods. Therefore, the active compounds of the present invention can be formulated into various dosage forms for oral, buccal, nasal, parenteral (e.g., intravenous, intramuscular, or subcutaneous), rectal, inhalation, or blow-through administration. As is well known to those skilled in the art, the dosage of a drug depends on a variety of factors, including but not limited to: the activity of the specific compound used, the patient's age, the patient's weight, the patient's health condition, the patient's behavior, the patient's diet, the time of administration, the method of administration, the rate of excretion, and the combination of drugs. Furthermore, optimal treatment methods, such as the mode of treatment, the daily dosage of compound (I), or the type of pharmaceutically acceptable salt thereof, can be adjusted and validated according to conventional treatment regimens. Unless specifically defined, any terminology in this invention will be used in its general sense as understood by those skilled in the art. Unless otherwise stated, the terms used in the specification and claims have the meanings described below.

[0052] Wherein “alkyl” refers to C1-C 20Saturated aliphatic hydrocarbon groups with straight and branched chains. Preferably, the alkyl group is an alkyl group having 1-12, sometimes preferably 1-6, and sometimes more preferably 1-4 carbon atoms. Representative examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl 2,3-Dimethylpentyl, 2,4-Dimethylpentyl, 2,2-Dimethylpentyl, 3,3-Dimethylpentyl, 2-Ethylpentyl, 3-Ethylpentyl, n-Octyl, 2,3-Dimethylhexyl, 2,4-Dimethylhexyl, 2,5-Dimethylhexyl, 2,2-Dimethylhexyl, 3,3-Dimethylhexyl, 4,4-Dimethylhexyl, 2-Ethylhexyl, 3-Ethylhexyl, 4-Ethylhexyl, 2-Methyl-2-Ethylpentyl, 2-Methyl-3-Ethylpentyl, n-Nonyl, 2-Methyl-2-Ethylhexyl, 2-Methyl-3-Ethylhexyl, 2,2-Diethylpentyl, n-Decyl, 3,3-Diethylhexyl, 2,2-Diethylhexyl, and their branched isomers. More preferably, the alkyl group is a lower alkyl group having 1 to 6 carbon atoms. Representative examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, etc. The alkyl group may be substituted or unsubstituted. When substituted, the substituent can be substituted at any available connection point. Preferably, the substituent is one or more substituents independently selected from alkyl, halogen, alkoxy, alkenyl, alkynyl, alkylsulfonyl, alkylamino, mercapto, hydroxyl, nitro, cyano, amino, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocyclol, cycloalkylthio, heterocycloalkylthio, and oxo.

[0053] "Cycloalkyl" refers to a saturated and / or partially unsaturated monocyclic or polycyclic hydrocarbon group having 3 to 20 carbon atoms, preferably 3 to 12 carbon atoms, more preferably 3 to 10 carbon atoms, and most preferably 3 to 8 carbon atoms or 3 to 6 carbon atoms. Representative examples of monocyclic cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cyclohepttrienyl, and cyclooctyl. Polycyclic cycloalkyl groups include cycloalkyl groups having spirocyclic, fused, or bridged rings.

[0054] "Alkoxy" refers to -O- (alkyl) and -O- (unsubstituted cycloalkyl), wherein the alkyl group is as defined above. Representative examples include, but are not limited to, methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy, cyclopentoxy, cyclohexoxy, etc. Alkoxy groups may be substituted or unsubstituted. When substituted, the substituent is preferably selected independently from one or more of alkyl, alkenyl, alkynyl, alkoxy, alkylsulfonyl, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, and heterocycloalkylthio, sometimes preferably one to five, and sometimes more preferably one to three substituents. "Hydroxyalkyl" refers to an alkyl group substituted with a hydroxyl group, wherein the alkyl group is as defined above.

[0055] "Alkenyl" refers to an alkyl compound containing a carbon-carbon double bond in its molecule, where alkyl is defined as described above. Alkenyl groups can be substituted or unsubstituted. When substituted, the substituent is preferably one or more of the following groups, independently selected from hydrogen, alkyl, alkoxy, halogen, haloalkyl, haloalkoxy, cycloalkyloxy, heterocyclic oxy, hydroxy, hydroxyalkyl, cyano, amino, nitro, cycloalkyl, heterocyclic, aryl, and heteroaryl substituents.

[0056] "Alkyne" refers to an alkyl compound containing a carbon-carbon triple bond in its molecule, where alkyl is defined as described above. Alkyne can be substituted or unsubstituted. When substituted, the substituent is preferably one or more of the following groups, independently selected from hydrogen, alkyl, alkoxy, halogen, haloalkyl, haloalkoxy, cycloalkyloxy, heterocyclic oxy, hydroxy, hydroxyalkyl, cyano, amino, nitro, cycloalkyl, heterocyclic, aryl, and heteroaryl substituents.

[0057] "Heterocyclic group" refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic substituent containing 3 to 20 ring atoms, wherein one or more ring atoms are heteroatoms selected from nitrogen, oxygen, and sulfur, wherein the sulfur may optionally be oxidized (i.e., forming sulfoxide or sulfone), but does not include the -OO-, -OS-, or -SS- ring moiety, and the remaining ring atoms are carbon. Preferably, it contains 3 to 12 (e.g., 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12) ring atoms, wherein 1 to 4 (e.g., 1, 2, 3, and 4) are heteroatoms; more preferably, it contains 3 to 8 ring atoms (e.g., 3, 4, 5, 6, 7, and 8), wherein 1 to 3 are heteroatoms (e.g., 1, 2, and 3); even more preferably, it contains 3 to 6 ring atoms, wherein 1 to 3 are heteroatoms; most preferably, it contains 5 or 6 ring atoms, wherein 1 to 3 are heteroatoms. Non-limiting examples of monocyclic heterocyclic groups include pyrrolidinyl, tetrahydropyranyl, 1,2,3,6-tetrahydropyridyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, and homopiperazinyl. Polycyclic heterocyclic groups include spirocyclic, fused-ring, and bridged-ring heterocyclic groups.

[0058] "Aryl" refers to a 6- to 14-membered all-carbon monocyclic or fused polycyclic (fused polycyclic) group having a conjugated π-electron system, preferably 6- to 10-membered, such as phenyl and naphthyl. The aryl ring comprises an aryl ring fused to a heteroaryl, heterocyclic, or cycloalkyl ring as described above, wherein the ring connected to the parent structure is an aryl ring. The aryl group can be substituted or unsubstituted; when substituted, it can be substituted at any usable connection point. The substituent is preferably selected from one or more of hydrogen, halogen, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyloxy, heterocyclicoxy, hydroxyl, hydroxyalkyl, cyano, amino, nitro, cycloalkyl, heterocyclic, aryl, and heteroaryl groups.

[0059] "Heteroaryl" refers to a heteroaryl system comprising 1 to 4 (e.g., 1, 2, 3, and 4) heteroatoms and 5 to 14 ring atoms, wherein the heteroatoms are selected from oxygen, sulfur, and nitrogen. The heteroaryl group is preferably 5 to 10-membered (e.g., 5, 6, 7, 8, 9, or 10-membered), more preferably 5- or 6-membered, such as furanyl, thiophene, pyridinyl, pyrroleyl, N-alkylpyrroleyl, pyrimidinyl, pyrazinyl, pyridazinyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, etc. The heteroaryl ring comprises a heteroaryl group fused to an aryl, heterocyclic, or cycloalkyl ring as described above, wherein the ring connected to the parent structure is a heteroaryl ring. The heteroaryl group can be substituted or unsubstituted. When substituted, it can be substituted at any usable connection point. The substituent is preferably selected from one or more of hydrogen, halogen, alkyl, alkoxy, haloalkyl, haloalkoxy, cycloalkyloxy, heterocyclic oxy, hydroxy, hydroxyalkyl, cyano, amino, nitro, cycloalkyl, heterocyclic, aryl, and heteroaryl.

[0060] “Bond” refers to a covalent bond marked with “—”. “Hydroxyalkyl” refers to an alkyl group substituted with a hydroxyl group, wherein the alkyl group is as defined above. “Hydroxy” refers to a -OH group. “Halogen” refers to a fluorine, chlorine, bromine, or iodine atom. “Amino” refers to a -NH2 group. “Cyano” refers to a -CN group. “Nitro” refers to a -NO2 group. “Oxo-group” refers to a =O group. “Carboxyl” refers to a -C(O)OH group. “Mercapto” refers to -SH. “Carbonyl” refers to C=O. “Carboxyl ester” refers to -C(O)O (alkyl) or -C(O)O (cycloalkyl), wherein the alkyl and cycloalkyl groups are as defined above. “Substituted” means that one or more hydrogen atoms in the group, preferably up to five, more preferably one to three hydrogen atoms, are independently substituted by the corresponding number of substituents. It goes without saying that substituents are only in their possible chemical positions. Those skilled in the art can determine (by experiment or theory) possible or impossible substitutions without much effort. For example, amino or hydroxyl groups having free hydrogen may be unstable when combined with carbon atoms having unsaturated bonds (such as alkenes). As used in the context of this invention, the term "solvent" refers to the physical bond of the compound of this invention with one or more, preferably one to three solvent molecules (whether organic or inorganic). This physical bond includes hydrogen bonds. In some cases, such as when one or more, preferably one to three solvent molecules are incorporated into the lattice of a crystalline solid, the solvate can be separated. Exemplary solvates include, but are not limited to, hydrates, ethanolates, methanolates, and isopropanolates. Methods of solvation are generally known in the art. "Prodrug" refers to a compound that can be converted in vivo under physiological conditions, such as by hydrolysis in the blood, to produce the active parent compound. Common examples include, but are not limited to, ester and amide forms of compounds with a carboxylic acid moiety as the active form.

[0061] P005091 is a dual-target inhibitor of USP7 / 47 containing a thiophene sulfide structure. Studies have reported that the mechanism of action of thiophene sulfide-based USP7 inhibitors involves covalent modification of the cysteine ​​residue at position 233 of USP7, inducing a conformational change in the enzyme associated with the active site. Because USP7 plays a crucial role in various oncogenic pathways, its inhibitor P005091 is frequently used as a tool in anticancer drug research. This invention, through extensive research, has found that the USP7 / 47 inhibitor P005091 can efficiently inhibit the expression of inflammatory factors such as IFN-β1. Based on this research, this invention further investigated the function and structure-activity relationship of P005091, designing and screening a series of molecular compounds with significant therapeutic activity against inflammatory bowel disease (IBD). This provides a clear scientific basis and direction for the subsequent development of drugs for IBD and related diseases, and has significant clinical implications. Attached Figure Description

[0062] Figure 1 This is a schematic diagram showing the effects of different concentrations of P005091 on diABZI-mediated IFN-β1 in BJ cells.

[0063] Figure 2 This is a schematic diagram showing the effects of different concentrations of P005091 on S-cddA-mediated IFN-β1 in THP-1 cells.

[0064] Figure 3 This is a schematic diagram showing the effect of different concentrations of P005091 on diABZI-mediated IFN-β1 in RAW264.7 cells.

[0065] Figure 4 This is a schematic diagram showing the effect of different concentrations of P005091 on S-cddA-mediated IFN-β1 in L929 cells.

[0066] Figure 5 This is a schematic diagram showing the effect of different concentrations of P005091 on S-cddA-mediated IFN-β1 in DC2.4 cells.

[0067] Figure 6 This is a schematic diagram of the cytotoxicity test results for P005091.

[0068] Figure 7 This is a schematic diagram of an animal experimental protocol for P005091 to inhibit DSS-induced acute colitis.

[0069] Figure 8 Photograph of the colon of mice with DSS-induced acute colitis induced by P005091 inhibition.

[0070] Figure 9 This is a schematic diagram showing the changes in colon length in mice with DSS-induced acute colitis induced by P005091 inhibition.

[0071] Figure 10 This is a schematic diagram showing the results of body weight changes in mice with DSS-induced acute colitis induced by P005091 inhibition.

[0072] Figure 11 This is a schematic diagram showing the serum levels of IFN-β1, CXCL10, and IL-6 in mice with DSS-induced acute colitis after P005091 treatment.

[0073] Figure 12 This is a schematic diagram of the H&E staining, p-IRF3, and p-STAT3 immunohistochemical staining results of colon tissue from mice with DSS-induced acute colitis after P005091 treatment.

[0074] Figure 13This is a schematic diagram showing the H&E staining, p-IRF3, and p-STAT3 immunohistochemical scoring results of colon tissue from mice with DSS-induced acute colitis after P005091 treatment.

[0075] Figure 14 This diagram illustrates the results of different concentrations of compounds (H-151, NTP1, NTP14, NTP16, NTP19) inhibiting relative luciferase activity in THP1-Luc cells.

[0076] Figure 15 IC50 values ​​for H-151, NTP1, NTP14, NTP16, and NTP19 in THP1-Luc cells 50 A diagram illustrating the test results.

[0077] Figure 16 This diagram illustrates the results of different concentrations of NTP14 and NTP16 inhibiting the IFN-β1 response mediated by diABZI or S-cddA in BJ cells.

[0078] Figure 17 This is a schematic diagram showing the results of different concentrations of NTP14 and NTP16 inhibiting the IFN-β1 response mediated by diABZI or S-cddA in DC2.4 cells.

[0079] Figure 18 A schematic diagram of the colon of mice with DSS-induced acute colitis, showing NTP14 and NTP16 inhibition.

[0080] Figure 19 A schematic diagram showing the changes in colon length in mice with DSS-induced acute colitis induced by NTP14 and NTP16 inhibition.

[0081] Figure 20 This is a schematic diagram showing the results of NTP14 and NTP16 inhibiting DSS-induced acute colitis in mice.

[0082] Figure 21 This is a schematic diagram of the H&E staining results of colon tissue in mice with DSS-induced acute colitis after P005091 treatment.

[0083] Figure 22 This is a schematic diagram of the H&E score results of colon tissue in mice with DSS-induced acute colitis after P005091 treatment.

[0084] Figure 23 This is a schematic diagram showing the serum levels of IFN-β1, CXCL10, and IL-6 in mice with DSS-induced acute colitis after NTP14 and NTP16 treatment. Detailed Implementation

[0085] To make the objectives, technical solutions, and effects of this invention clearer and more explicit, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

[0086] Unless otherwise specified, the cell lines listed in this invention, including BJ, RAW264.7, THP1-Luc, L929, and DC2.4, were cultured according to existing techniques. All cell lines were identified by short tandem repeat analysis at the China Center for Type Culture Collection (Wuhan), and the presence of mycoplasma contamination was verified using a PCR detection kit (Shanghai Biothrive Sci). All cell lines were cryopreserved in liquid nitrogen for subsequent experiments. All reagents used in this invention were commercially available. The experimental methods used in this invention, such as molecular biology experiments and cell experiments, are conventional methods and techniques in the art. Representative results from biological experiment replicates are presented in the accompanying figures, and data are displayed as mean ± SD and mean ± SEM as specified in the figures. All in vitro experiments were repeated at least three times. Data were analyzed using GraphPadPrism 8.0 or SPSS 22.0 software. Conventional medical statistical methods such as t-tests, chi-square tests, and analysis of variance were used to compare the differences between the means of two or more groups. p < 0.05 was considered a significant difference.

[0087] Example 1

[0088] BJ, RAW264.7, THP1-Luc, L929, and DC2.4 cell models were used. Different concentrations of P005091 were added for treatment (SATE-cddA (0.4 μM in THP-1 cells; 0.5 μM in RA2264.7, L929, and DC2.4 cells), S-cddA, CAS: 2447159-29-5, or diABZI (1 μM in BJ cells), CAS: 2138299-33-7 served as positive controls). Changes in the expression activities of inflammatory factors such as IFN-β1 and CXCL10 were detected using qPCR or Renal luciferase reporter gene assay. Specifically, the qPCR procedure was as follows: 800,000 cells / well were seeded into six-well plates before drug administration. After culture for at least 12 hours, cell adhesion was observed; cells in good condition were then drug-treated. THP1-Luc cells were seeded at 1,200,000 cells / well in six-well plates after counting, followed by direct drug administration. In subsequent experiments, all pipette tips were sterilized at high temperatures, and all centrifuge tubes, eight-tube sets, and other disposable sterile, RNase-free products were used.

[0089] (1) Cell RNA extraction: For RAW264.7 and L929 cells, at the end of incubation, the drug-containing culture medium was aspirated, the cells were washed once with PBS buffer, 800 μL of Trizol was added to each well, and the cells were incubated at 4°C for 30 min. The lysis buffer was then transferred to a 1.5 mL centrifuge tube. 160 μL of chloroform was added to each sample, and the mixture was thoroughly mixed and incubated at room temperature for 10 min. Then, the tube was centrifuged at 4°C and 13,000 rpm for 20 min. 300 μL of the supernatant was transferred to a new 1.5 mL centrifuge tube, an equal volume of isopropanol was added, and the mixture was thoroughly mixed and incubated at room temperature for 10 min. Then, the tube was centrifuged at 4°C and 13,000 rpm for 10 min. The supernatant was discarded, and 500 μL of 75% ethanol (ethanol:DEPC water = 3:1) was added to the white precipitate at the bottom of the tube. After mixing, the tube was directly centrifuged at 4°C and 13,000 rpm for 5 min. Discard the supernatant and allow it to air dry at room temperature for 1 hour until the white precipitate at the bottom of the tube is no longer visible to the naked eye. Then add 20 μL of DEPC water and test the concentration using a micro-quantitative analyzer.

[0090] (2) DNA removal and reverse transcription: Take 2000 ng of the RNA obtained in the previous step and adjust the volume to 12 μL with DEPC water. Add 4×g DNA wiper to the RNA and heat in a metal bath at 42℃ for 3 min. Then add reverse transcriptase to the system and heat in a metal bath at 50℃ for 15 min. Subsequently, raise the temperature of the metal bath to 85℃ and heat for 3 min to obtain cDNA. Add 80 μL of DEPC water to dilute the cDNA, and adjust the total volume of each sample to 100 μL.

[0091] (3) Real-time quantitative PCR system preparation and procedure: Prepare the system in eight-tube strips as shown in the table below. qPCR system: 4.6 μL cDNA sample, 5 μL qPCR SYBR Green Master Mix, 0.2 μL Forward Primer (10 ng / μL), 0.2 μL Reverse Primer (10 ng / μL). After preparation, centrifuge briefly to mix. Perform qPCR according to the procedure shown in the table below: pre-denaturation 95℃ for 3 min, denaturation 95℃ for 15 s, annealing 55℃ for 30 s, extension 72℃ for 20 s, and melting curve determination. The denaturation-annealing-extension cycle is performed 44 times.

[0092] (4) Data processing: The δcq value is the difference between the target gene and the internal reference gene; the δcq value is the difference between the δcq value of the control group and the δcq value of the experimental group. 2 -δδcq The calculated value represents the relative expression level of the target gene. Independent experiments should be repeated at least three times, and the results are statistically expressed as mean ± standard deviation.

[0093] The results showed that in the above cell model, P005091 could increase its inhibition rate on S-cddA-mediated IFN-β1 and CXCL10 expression in a concentration-dependent manner; the IC50 of P005091 in different cell types was obtained through calculation and fitting. 50 Value (see) Figure 1-5 These results indicate that P005091 has a strong inhibitory effect on S-cddA-mediated IFN-β expression in RAW264.7 cells, and that the duration of interaction between P005091 and cells can affect its activity. Repeating the above experiments in mouse cells (L929) and human cells (THP1-Luc) revealed that P005091 also exhibited similarly significant inhibitory activity against S-cddA-mediated IFN-β1 expression.

[0094] To rule out whether the inhibitory effect of P005091 was due to its toxicity, different concentrations of P005091 were incubated in BJ, RAW264.7, L929, THP-1, and HeLa cells, and cell viability was measured after 24 hours. The results showed that cell viability decreased in a concentration-dependent manner, and different cell lines exhibited varying sensitivities; the half-maximal inhibitory concentration (IC50) was above 20 μM for all cell lines (see [link to relevant documentation]). Figure 6 Therefore, it can be concluded that P005091 does not inhibit the IFN-β1 response through cytotoxicity. In summary, P005091 can effectively inhibit IFN-β1 transcriptional activation and has a regulatory effect on the viability of different cell lines, providing a basis for further analysis of this pathway and the development of related disease intervention strategies.

[0095] Example 2

[0096] The foregoing embodiments clearly demonstrate that P005091 is a broad-spectrum and highly effective type I interferon inhibitor. To obtain more active molecules with excellent inhibitory effects, the molecule of P005091 was modified, and its structure-activity relationship was explored, as detailed below:

[0097] (1) Synthesis of intermediate 1-(5-chloro-4-nitrothiophen-2-yl)ethan-1-one: 500 mg of 2-acetyl-5-chlorothiophene was dissolved in 3 mL of concentrated sulfuric acid and stirred in an ice bath until the solid was completely dissolved. 1 mL of pre-cooled nitric acid was added dropwise to 1 mL of pre-cooled concentrated sulfuric acid, and then the mixed acid was slowly added dropwise to the reaction system. The reaction was monitored by TLC. After the starting material was completely converted, the reaction solution was slowly added dropwise to ice water, and a pale yellow solid precipitated. The solid was extracted with an ethyl acetate / water system, the organic layer was concentrated, and the crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase. The NMR identification data are as follows: 1-(5-chloro-4-nitrothiophen-2-yl)ethan-1-one pale yellow solid, yield 50%; 1HNMR (400MHz, CDCl3) δ8.07 (s, 1H), 2.58 (s, 3H).

[0098] (2) Synthesis of NTP1 (P005091), NTP2, NTP3, NTP4, NTP5, NTP6, NTP7, NTP8, NTP9, NTP10, NTP11, NTP12, and NTP13: The synthesis proceeds according to the following reaction formula:

[0099]

[0100] Specifically, 50 mg (0.24 mmol) of 1-(5-chloro-4-nitrothiophene-2-yl) acetone was dissolved in 4 mL of methanol and stirred at room temperature. Then, 2,3-dichlorothiophenol (NTP1), 4-methoxythiophenol (NTP2), 2-methoxythiophenol (NTP3), 3-aminothiophenol (NTP4), 4-methylthiophenol (NTP5), 3-methoxythiophenol (NTP6), 4-aminothiophenol (NTP7), 4-methylaminothiophenol (NTP8), 2-aminothiophenol (NTP9), 2,6-dichlorothiophenol (NTP10), 2-fluorothiophenol (NTP11), 2-chlorothiophenol (NTP12), and thiophenol (NTP13) were added, with 0.24 mmol of the corresponding thiophenol added. After dissolution, 13 mg (0.3 mmol) of sodium methoxide was added, and the reaction was carried out at room temperature for 4-8 h. The reaction was monitored by TLC until the thiophene feedstock was essentially converted. The pH was then adjusted to neutral with 1N hydrochloric acid. Extraction was performed using an ethyl acetate / water system, and the organic layer was concentrated. The crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase. The NMR identification data are as follows:

[0101] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP1) pale yellow solid, yield 46%; 1 HNMR (400MHz, CDCl3) δ8.08 (s, 1H), 7.74-7.68 (m, 2H), 7.37 (t, J=7.9Hz, 1H), 2.50 (s, 3H); HRMScalcd for [M+Na] + :369.9137, found 369.9160.

[0102] 1-(5-((4-methoxyphenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((4-methoxyphenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP2) pale yellow solid, yield 45%; 1 HNMR (400MHz, CDCl3) δ8.06 (s, 1H), 7.56 (d, J=8.6Hz, 2H), 7.03 (d, J=8.7Hz, 2H), 3.89 (s, 3H), 2.47 (s, 3H); HRMS calcd for [M+Na] + 332.0022, found 332.0025. The structure is as follows:

[0103] (NTP2).

[0104] 1-(5-((2-methoxyphenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((2-methoxyphenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP3) pale yellow solid, yield 47%; 1 HNMR (400MHz, CDCl3) δ8.07 (s, 1H), 7.63–7.54 (m, 2H), 7.10–7.04 (m, 2H), 3.87 (s, 3H), 2.47 (s, 3H); HRMS calcd for [M+Na] + 332.0022, found 332.0028. The structure is as follows:

[0105] (NTP3).

[0106] 1-(5-((3-aminophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((3-aminophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP4) yellow solid, yield 37%; 1 HNMR (400MHz, CDCl3) δ8.06 (s, 1H), 7.29 (t, J=7.9Hz, 1H), 7.02 (d, J=7.3Hz, 1H), 6.94 (s, 1H), 6.84 (d, J=8.2Hz, 1H), 2.48 (s, 3H); HRMScalcdfor[M+H] + : 295.0206, found 295.0238. The structure is as follows:

[0107] (NTP4).

[0108] 1-(4-nitro-5-(p-tolylthio)thiophen-2-yl)ethan-1-one (1-(4-nitro-5-(p-tolylthio)thiophen-2-yl) ethyl ketone, NTP5) white solid, yield 44%; 1 H NMR (400 MHz, CDCl3) δ 8.07 (s, 1H), 7.53 (d, J = 8.1 Hz, 2H), 7.33 (d, J = 7.9 Hz, 2H), 2.47 (s, 3H), 2.45 (s, 3H). The structural formula is shown below:

[0109] (NTP5).

[0110] 1-(5-((3-methoxyphenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((3-methoxyphenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP6) pale yellow solid, yield 45%; 1 HNMR (400MHz, CDCl3) δ8.07 (s, 1H), 7.48–7.41 (m, 1H), 7.26–7.23 (m, 1H), 7. 20–7.17 (m, 1H), 7.11 (dd, J=8.4, 2.6Hz, 1H), 3.85 (s, 3H), 2.48 (s, 3H); HRMS calcd for [M+Na] + 332.0022, found 332.0039. The structure is as follows:

[0111] (NTP6).

[0112] 1-(5-((4-aminophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((4-aminophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP7) yellow solid, yield 34; 1 HNMR (400MHz, CDCl3) δ8.05 (s, 1H), 7.41–7.35 (m, 2H), 6.78–6.72 (m, 2H), 2.47 (s, 3H); HRMS calcd for [M+Na] + 317.0025, found 317.0036. The structure is as follows:

[0113] (NTP7).

[0114] 1-(5-((4-(methylamino)phenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one(1-(5-((4-(methylamino)phenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one, NTP8) orange solid, yield 42; 1 HNMR (400MHz, CDCl3) δ8.05 (s, 1H), 7.39 (d, J=8.6Hz, 2H), 6.65 (d, J=8.6Hz, 2H), 2.90 (s, 3H), 2.46 (s, 3H); HRMS calcd for [M+Na] + : 331.0182, found 331.0188. The structure is as follows:

[0115] (NTP8).

[0116] 1-(5-((2-aminophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((2-aminophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP9) yellow solid, yield 35%; 1 HNMR (400MHz, DMSO) δ8.44 (s, 1H), 7.38 (d, J=7.5Hz, 1H), 7.31 (t, J=7.5Hz, 1H), 6.87 ( d, J=8.0Hz, 1H), 6.66 (t, J=7.4Hz, 1H), 5.82 (s, 2H), 2.50 (s, 3H); HRMScalcdfor[M+H] + : 295.0206, found 3295.0238. The structure is as follows:

[0117] (NTP9).

[0118] 1-(5-((2,6-dichlorophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((2,6-dichlorophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP10) pale yellow solid, yield 43%; 1HNMR (400MHz, CDCl3) δ8.09 (s, 1H), 7.57 (d, J=1.1Hz, 1H), 7.55 (s, 1H), 7.47 (dd, J=8.9, 7.1Hz, 1H), 2.50 (s, 3H); HRMS calcd for [M+Na] + 349.9137, found 369.9140. The structure is as follows:

[0119] (NTP10).

[0120] 1-(5-((2-fluorophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((2-fluorophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP11) white solid, yield 36%; 1 HNMR (400MHz, Acetone) δ8.32 (s, 1H), 7.88–7.77 (m, 2H), 7.49 (t, J=8.2Hz, 2H), 2.55 (s, 3H); HRMS calcdfor [M+Na] + 319.9822, found 319.9845. The structure is as follows:

[0121] (NTP11).

[0122] 1-(5-((2-chlorophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-((2-chlorophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one, NTP12) pale yellow solid, yield 45%; 1 HNMR (400MHz, CDCl3) δ8.08 (s, 1H), 7.76 (d, J=7.7Hz, 1H), 7.64 (d, J=8.0Hz, 1H), 7.55 (t, J=7.7Hz, 1H), 7.43 (t, J=7.5Hz, 1H), 2.49 (s, 3H); HRMS calcd for [M+Na] + 335.9526, found 335.9543. The structure is as follows:

[0123] (NTP12).

[0124] 1-(4-nitro-5-(phenylthio)thiophen-2-yl)ethan-1-one (1-(4-nitro-5-(phenylthio)thiophen-2-yl)ethan-1-one, NTP13) white solid, yield 43%; 1 HNMR (400MHz, CDCl3) δ8.07 (s, 1H), 7.67 (dd, J=5.2, 3.1Hz, 2H), 7.62–7.51 (m, 3H), 2.48 (d, J=5.3Hz, 3H); HRMScalcdfor[M+H] + : 280.0097, found 280.0084. The structure is as follows:

[0125] (NTP13).

[0126] (3) Synthesis of NTP14, NTP15, and NTP16: The synthesis is carried out according to the following reaction formula.

[0127]

[0128] Specifically, 1-(5-chloro-4-nitrothiophene-2-yl) was dissolved in 4 mL of toluene and stirred at room temperature. 0.24 mmol of 2,3-dichlorophenol (NTP14) was added and dissolved, followed by the addition of 41.4 mg (0.30 mmol) of potassium carbonate. The reaction was carried out at room temperature for 12 h. The reaction was monitored by TLC until the thiophene precursor was essentially converted. The mixture was then extracted with an ethyl acetate / water system, and the organic layer was concentrated. The crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase.

[0129] Ethyl ketone (0.24 mmol) or methyl 5-chloro-4-nitrothiophene-2-carboxylic acid (0.24 mmol) was dissolved in 4 mL of toluene and stirred at room temperature. 0.24 mmol of 2,3-dichlorophenol (NTP15) or 2,3-dichlorothiophenol (NTP16) was added, and the reaction was carried out at room temperature for 12 h. The reaction was monitored by TLC. After the thiophene starting material was substantially converted, the mixture was extracted with an ethyl acetate / water system, the organic layer was concentrated, and the crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase.

[0130] Its NMR identification data are as follows:

[0131] 1-(5-(2,3-dichlorophenoxy)-4-nitrothiophen-2-yl)ethan-1-one (1-(5-(2,3-dichlorophenoxy)-4-nitrothiophen-2-yl) ethyl ketone, NTP14) white solid, yield 44%; 1HNMR (400MHz, CDCl3) δ8.02 (s, 1H), 7.52 (d, J=6.9Hz, 1H), 7.34 (t, J=8.2Hz, 1H), 7.27 (d, J=5.4Hz, 1H), 2.52 (s, 3H); HRMS calcd for [M+Na] + 353.9365, found 353.9340. The structure is as follows:

[0132] (NTP14).

[0133] methyl5-((2,3-dichlorophenyl)thio)-4-nitrothiophene-2-carboxylate (NTP15) pale yellow solid, 58% yield; 1 HNMR (400MHz, CDCl3) δ8.18 (s, 1H), 7.71 (d, J=8.3Hz, 2H), 7.38 (t, J=7.9Hz, 1H), 3.85 (s, 3H); HRMS calcd for [M+Na] + : 385.9086, found 385.9086. The structure is as follows:

[0134] (NTP15).

[0135] methyl 5-(2,3-dichlorophenoxy)-4-nitrothiophene-2-carboxylate (NTP16) was a pale yellow solid in 62% yield. 1 HNMR (400MHz, CDCl3) δ8.12 (s, 1H), 7.51 (dd, J=8.1, 1.4Hz, 1H), 7.33 (t, J=8.2Hz, 1H), 7.24 (d, J=1.4Hz, 1H), 3.87 (s, 3H); HRMS calcd for [M+Na] + 369.9314, found 369.9316. The structure is as follows:

[0136] (NTP16).

[0137] (4) Synthesis of NTP17 and NTP18: The synthesis proceeds according to the following reaction:

[0138]

[0139] Specifically, 150 mg (0.41 mmol) of NTP15 was dissolved in 8 mL of a methanol-tetrahydrofuran-water mixture (volume ratio 2:4:2), and 66 mg (1.65 mmol) of lithium hydroxide was added. The mixture was stirred at room temperature for 6 h. The reaction was monitored by TLC. After the thiophene feedstock was basically converted, 1 N hydrochloric acid was added to adjust the pH to neutral. The mixture was extracted with an ethyl acetate / water system, and the organic layer was then extracted with sodium chloride solution. The organic layer was concentrated and saturated to obtain the product NTP17. Its NMR identification data are as follows:

[0140] 5-((2,3-dichloro-5-phenyl)thio)-4-nitrothiophene-2-carboxylic acid (NTP17) pale brown solid, 92% yield; ¹H NMR (400 MHz, DMSO) δ 8.06 (s, 1H), 7.98 (m, 2H), 7.62 (t, J = 8.0 Hz, 1H); HRMS calcd for [M+Na] + : 371.8929, found 371.8916. 100mg (0.29mmol). The structural formula is shown below:

[0141] (NTP17).

[0142] NTP17 (200 mg, 0.58 mmol) was dissolved in 6 mL of dichloromethane, and 150 mg (0.72 mmol) of HATU, 117 mg (0.90 mmol) of N,N-diisopropylethylamine, and 42.4 mg (0.58 mmol) of n-butylamine were added sequentially. The reaction was carried out at room temperature for 4 h. The reaction was monitored by TLC until the product spot no longer increased. The reaction solution was concentrated, extracted with an ethyl acetate / water system, and the organic layer was concentrated. The crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase. The NMR identification data are as follows:

[0143] N-butyl-5-((2,3-dichlorophenyl)thio)-4-nitrothiophene-2-carboxamide (NTP18) is a light brown solid, 12% yield; 1HNMR (400MHz, CDCl3) δ7.83 (s, 1H), 7.69 (dd, J=7.9, 2.7Hz, 2H), 7.36 (t, J=7.9Hz, 1H), 5.97 (s, 1H), 3.70 (t, J=6.3Hz, 1H ), 3.59 (t, J=6.5Hz, 1H), 3.38 (dd, J=13.2, 6.9Hz, 2H), 1.93–1.85 (m, 1H), 1.76-1.69 (m, 1H), 0.94 (t, J=7.3Hz, 3H); HRMS calcd for [M+Na] + : 426.9715, found 426.9710. The structure is as follows:

[0144] (NTP18).

[0145] (5) Synthesis of NTP20 and NTP21: The synthesis is carried out according to the following reaction formula:

[0146] 500 mg (1.43 mmol) of NTP17 was dissolved in 8 mL of DMF. Then, 412 mg (1.72 mmol) of HATU, 180.6 mg (2.15 mmol) of triethylamine, and 229 mg (1.43 mmol) of mono-Boc ethylenediamine were added sequentially, and the mixture was stirred at room temperature for 12 h. The reaction was monitored by TLC until the product spot no longer increased. Extraction was performed with an ethyl acetate / water system, the organic layer was concentrated, and the crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase. The NMR identification data are as follows:

[0147] tert-butyl(2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophene-2-carboxamido)ethyl)carbamate (NTP21) yellow solid, yield 27%; 1 HNMR (400MHz, DMSO): δ8.84–8.77 (m, 1H), 8.35 (s, 1H), 7.97 (t, J=8.3Hz, 2H), 7.61 (t, J=8 .0Hz, 1H), 6.84 (t, J=5.4Hz, 1H), 3.23–3.16 (m, 2H), 3.08–3.01 (m, 2H), 1.35 (s, 9H); HRMS calcd for [M+Na] + 514.0035, found 514.0087. The structure is as follows:

[0148] (NTP21).

[0149] 90 mg (0.18 mmol) of NTP21 was dissolved in 3 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was stirred at room temperature for 12 h. The reaction was monitored by TLC until the starting material was completely converted. The reaction solution was concentrated, extracted with a dichloromethane / saturated sodium carbonate solution, and the organic layer was concentrated to obtain the product. Its NMR identification data are as follows:

[0150] N-(2-aminoethyl)-5-((2,3-dichlorophenyl)thio)-4-nitrothiophene-2-carboxamide (N-(2-aminoethyl)-5-((2,3-dichlorophenyl)thio)-4-nitrothiophene-2-carboxamide, NTP20) is a yellow solid, 65% yield; 1 HNMR (400MHz, DMSO): δ8.90 (s, 1H), 8.36 (s, 1H), 7.97 (t, J = 8.3Hz, 2H), 7.62 (t, J = 8.0Hz, 1H), 2.84 (t, J = 6.2Hz, 2H); HRMS calcd for [M+H] + 391.9692, found 3919649. The structure is as follows:

[0151] (NTP20).

[0152] (6) Synthesis of NTP19, NTP22, NTP23, NTP24, NTP35, NTP37, NTP42, NTP43, NTP44, NTP54, NTP55, NTP56, NTP57, NTP60, NTP62, NTP63, and NTP67: The synthesis proceeds according to the following reaction formula:

[0153]

[0154] Specifically, 1 g (8.43 mmol) of 2-chlorothiophene was dissolved in 8 mL of chloroform, and 1.5 equivalents (12.65 mmol) of substituted acyl chlorides were added: propionyl chloride (NTP19), acryloyl chloride (NTP22), trifluoroacetyl chloride (NTP23), (E)-butyronyl chloride (NTP24), propynoyl chloride (NTP35), methyl-3-chloro-3-oxopropionate (NTP37), 3-oxobutyryl chloride (NTP42), and chloroacetyl chloride (NTP... 43) Hydroxyacetyl chloride (NTP44), butyryl chloride (NTP54), isobutyryl chloride (NTP55), 2-oxopropionyl chloride (NTP56), 3-chloro-2-oxopropionyl chloride (NTP57), methyl 2-chloro-2-oxoacetate (NTP60), methyl 4-chloro-4-oxobutyrate (NTP62), and 4-oxopentanoyl chloride (NTP63) were added slowly, followed by the addition of 2.25 g (16.86 mmol) of aluminum trichloride. The mixture was stirred at room temperature for 8 h. The reaction was monitored by TLC until the product spot no longer increased. The reaction solution was concentrated, extracted with an ethyl acetate / water system, and the organic layer was concentrated. The crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase to obtain product P1. 0.40 mmol of P1 was dissolved in 3 mL of concentrated sulfuric acid. The nitrating reagent, equivalent, and reaction conditions were the same as those used in (1) to prepare the nitration intermediate 1-(5-chloro-4-nitrothiophen-2-yl)ethan-1-one, yielding intermediate P2. 0.27 mmol of P2 was dissolved in 4 mL of methanol. The thiophenolizing reagent, equivalent, and reaction conditions were the same as those used in (2) to prepare NTP1, yielding the final product. Its NMR identification data are as follows:

[0155] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)propan-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)propan-1-one, NTP19) pale yellow solid, yield 52%; 1 HNMR (400MHz, CDCl3) δ8.09 (s, 1H), 7.74–7.66 (m, 2H), 7.37 (t, J=7.9Hz, 1H), 2.85 (q, J=7.3Hz, 2H), 1.19 (t, J=7.3Hz, 3H); HRMS calcd for [M+Na] + 383.9323, found 383.9324. The structure is as follows:

[0156] (NTP19).

[0157] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)prop-2-en-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)prop-2-en-1-one, NTP22), pale yellow solid, 21% yield; 1 HNMR (400MHz, DMSO-d6) 8.07 (s, 1H), 7.78 (dd, J=7.9Hz, 2H), 6.30–6.05 (m, 3H); HRMScalcd for [M+Na] + 381.9144, found 381.9142. The structure is as follows:

[0158] (NTP22).

[0159] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2,2,2-trifluoroethan-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2,2,2-trifluoroethan-1-one, NTP23), pale yellow solid, 43% yield; 1 HNMR (400MHz, DMSO-d6) δ8.09 (s, 1H), 7.92 (dd, J=7.9Hz, 1H), 7.32 (dd, J=7.9Hz, 1H) 7.17 (t, J=7.9Hz, 1H); HRMS calcd for [M+Na] + : 423.8862, found 423.8876. The structure is as follows:

[0160] (NTP23).

[0161] (E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)but-2-en-1-one ((E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)but-2-en-1-one, NTP24), pale yellow solid, 37% yield; 1HNMR (400MHz, DMSO-d6) δ8.08 (s, 1H), 7.53 (dd, J=7.9Hz, 1H), 7.33 (dd, J=7.9Hz, 1H) 7.23 (t , J=7.9Hz, 1H), 6.26 (dd, J=7.3Hz, 1H), 6.02 (dd, J=7.3Hz, 1H), 2.45 (dd, J=7.3Hz, 3H); HRMS calcd for [M+Na] + 395.9301, found 395.9289. The structure is as follows:

[0162] (NTP24).

[0163] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)but-3-yn-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)but-3-yn-1-one, NTP35), pale yellow solid, yield 41%; 1 HNMR (400MHz, CDCl3) δ8.09 (s, 1H), 7.72 (dd, J=7.9Hz, 1H), 7.52 (dd, J=7.9Hz, 1H) 7.12 (t , J=7.9Hz, 1H), 3.73 (dt, J=7.2Hz, 2H), 6.02 (t, J=7.2Hz, 1H), 2.45 (dd, J=7.3Hz, 3H); HRMS calcd for [M+Na] + 393.9144, found 393.9134. The structure is as follows:

[0164] (NTP35).

[0165] methyl 3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopropanoate (NTP37), pale yellow solid, 42% yield; 1 HNMR (400MHz, DMSO-d6) 8.09 (s, 1H), 7.57 (dd, J=7.9Hz, 1H), 7.37 (dd, J=7.9Hz, 1H) 7.13 (t, J=7.9Hz, 1H), 3.93 (s, 2H), 3.84 (s, 2H); HRMS calcd for [M+Na]+ : 427.9199, found 427.9205. The structure is as follows:

[0166] (NTP37).

[0167] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)butane-1,3-dione (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)butane-1,3-dione, NTP42), pale yellow solid, 51% yield; 1 HNMR (400MHz, DMSO-d6) 8.09 (s, 1H), 7.56 (dd, J=7.9Hz, 1H), 7.36 (dd, J=7.9Hz, 1H) 7.13 (t, J=7.9Hz, 1H), 3.73 (s, 2H), 2.98 (s, 3H), 3.84 (s, 2H); HRMS calcd for [M+Na] + : 411.9250, found 411.9275. The structure is as follows:

[0168] (NTP42).

[0169] 2-chloro-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (2-chloro-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP43), pale yellow solid, 16% yield; 1 HNMR (400MHz, DMSO-d6) 8.09 (s, 1H), 7.58 (dd, J=7.9Hz, 1H), 7.38 (dd, J=7.9Hz, 1H) 7.09 (t, J=7.9Hz, 1H), 4.24 (s, 2H); HRMS calcd for [M+Na] + : 403.8755, found403.8749. The structure is as follows:

[0170] (NTP43).

[0171] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-hydroxyethan-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-hydroxyethanone, NTP44), pale yellow solid, 17% yield; 1 HNMR (400MHz, DMSO-d6) 8.09 (s, 1H), 7.67 (dd, J=7.9Hz, 1H), 7.47 (dd, J=7.9Hz, 1H), 5.87 (s, 1H), 5.27 (s, 2H); HRMS calcd for [M+Na] + 385.9094, found385.9104. The structure is as follows:

[0172] (NTP44).

[0173] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)butan-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)butanone, NTP54), pale yellow solid, 44% yield; 1 HNMR (400MHz, CDCl3) 8.06 (s, 1H), 7.72 (dd, J=7.9Hz, 1H), 7.52 (dd, J=7.9Hz, 1H), 3.41 (t, J=5.9Hz, 2H), 1.41-0.98 (m, 5H); HRMS calcd for [M+Na] + 397.9457, found 397.9470. The structure is as follows:

[0174] (NTP54).

[0175] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-methylpropan-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-methylpropanone, NTP55), pale yellow solid, yield 47%; 1HNMR (400MHz, CDCl3) 8.08 (s, 1H), 7.68 (dd, J=7.9Hz, 1H), 7.48 (dd, J=7.9Hz, 1H), 2.58 (m, 1H), 1.61 (t, J=7.5Hz, 6H); HRMS calcd for [M+Na] + 397.9457, found397.9472. The structure is as follows:

[0176] (NTP55).

[0177] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)propane-1,2-dione (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)propane-1,2-dione, NTP56), pale yellow solid, 56% yield; 1 HNMR (400MHz, CDCl3) 8.09 (s, 1H), 7.69 (dd, J=7.9Hz, 1H), 7.49 (dd, J=7.9Hz, 1H), 2.93 (s, 3H); HRMS calcd for [M+Na] + 397.9094, found 397.9106. The structure is as follows:

[0178] (NTP56).

[0179] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoacetylchloride (NTP57), pale yellow solid, 34% yield; 1 HNMR (400MHz, CDCl3) 8.09 (s, 1H), 7.65 (dd, J=7.9Hz, 1H), 7.45 (dd, J=7.9Hz, 1H); HRMS calcd for [M+Na] + : 417.8547, found 417.8532. The structure is as follows:

[0180] (NTP57).

[0181] methyl 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoacetate (NTP60), pale yellow solid, 34% yield; 1 HNMR (400MHz, CDCl3) 8.09 (s, 1H), 7.68 (dd, J=7.9Hz, 1H), 7.48 (dd, J=7.9Hz, 1H), 3.93 (s, 3H); HRMS calcd for [M+Na] + : 413.9043, found413.9050. The structure is as follows:

[0182] (NTP60).

[0183] methyl 4-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-4-oxobutanoate (NTP62), pale yellow solid, 31% yield; 1 HNMR (400MHz, CDCl3) 8.09 (s, 1H), 7.56 (dd, J=7.9Hz, 1H), 7.36 (dd, J=7.9Hz, 1H), 3.91 (s, 3H), 3.43-3.03 (m, 4H); HRMS calcd for [M+Na] + : 441.9356, found 441.9363. The structure is as follows:

[0184] (NTP62).

[0185] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)pentane-1,4-dione (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)pentane-1,4-dione, NTP63), pale yellow solid, 27% yield; 1HNMR (400MHz, CDCl3) 8.09 (s, 1H), 7.56 (dd, J=7.9Hz, 1H), 7.36 (dd, J=7.9Hz, 1H), 3.53-3.13 (m, 4H), 2.91 (s, 3H); HRMS calcd for [M+Na] + : 425.9407, found 425.9412. The structure is as follows:

[0186] (NTP63).

[0187] 2-bromo-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)ethan-1-one (2-bromo-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl) ethyl ketone, NTP67), pale yellow solid, 17% yield; 1 HNMR (400MHz, CDCl3) 8.09 (s, 1H), 7.54 (dd, J=7.9Hz, 1H), 7.34 (dd, J=7.9Hz, 1H), 5.16 (s, 2H); HRMS calcd for [M+Na] + : 447.8250, found 447.8239. The structure is as follows:

[0188] (NTP67).

[0189] (7) Synthesis of NTP25: The process is carried out according to the following reaction formula:

[0190]

[0191] Specifically, 200 mg (0.57 mmol) of NTP1 was dissolved in 10 mL of ethanol and stirred at room temperature until completely dissolved. 46 mg (1.14 mol) of sodium hydroxide was added, followed by the slow addition of 138 mg (0.86 mmol) of 2,4,6-trifluorobenzaldehyde. The mixture was refluxed at 80 °C for 12 h, and then the reaction solution was concentrated. Extraction was performed with ethyl acetate / water. The ethyl acetate layer was collected, and the crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0192] (E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-(2,4,6-trifluorophenyl)prop-2-en-1-one ((E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-(2,4,6-trifluorophenyl)prop-2-en-1-one, NTP25), pale yellow solid, 17% yield; 1 HNMR (400MHz, DMSO-d6) 8.09 (s, 1H), 7.87 (d, J=8.3Hz, 1H), 7.53 (dd, J=7.9Hz, 1H), 7.33 ( dd, J=7.9Hz, 1H), 7.17 (t, J=7.9Hz, 1H), 7.01 (d, J=8.3Hz, 1H); 6.70 (t, J=7.9Hz, 2H); HRMS calcd for[M+Na] + : 513.9175, found 513.9188. The structure is as follows:

[0193] (NTP25).

[0194] (8) Synthesis of NTP26: The process proceeds according to the following reaction:

[0195]

[0196] Specifically, 200 mg (0.57 mmol) of NTP1 was dissolved in 10 mL of dichloromethane and stirred at room temperature until completely dissolved. Then, 288 mg (1.15 mmol) of Fluolead and 114 mg (1.15 mmol) of pyridine hydrogen fluoride were added sequentially. The mixture was stirred in an ice bath for 12 h, then at room temperature for 48 h. The mixture was extracted with dichloromethane / water, and the dichloromethane layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0197] 2-((2,3-dichlorophenyl)thio)-5-(1,1-difluoroethyl)-3-nitrothiophene (2-((2,3-dichlorophenyl)thio)-5-(1,1-difluoroethyl)-3-nitrothiophene, NTP26), yellow solid, 12% yield; 1HNMR (400MHz, DMSO-d6) 7.57 (dd, J=7.9Hz, 1H), 7.30 (s, 1H), 7.33 (dd, J=7.9Hz, 1H), 7.17 (t, J=7.9Hz, 1H), 2.44 (t, J=7.8Hz, 3H), HRMS calcd for [M+Na] + 391.9163, found391.9171. The structure is as follows:

[0198] (NTP26).

[0199] (9) Synthesis of NTP27: The process proceeds according to the following reaction:

[0200]

[0201] Specifically, 5 g (42.2 mmol) of 2-chlorothiophene was dissolved in 20 mL of acetonitrile. After complete dissolution, 8.79 g (63.3 mmol) of (E)-4-chlorobutadiene chloride was added, followed by the slow addition of 8.43 g (63.3 mmol) of aluminum trichloride. The mixture was stirred at room temperature for 8 h to obtain intermediate P1. 4 g (18.1 mmol) of P1 was slowly added to 15 mg of concentrated sulfuric acid, and the mixture was heated to 80 °C and refluxed for 12 h. The reaction solution was slowly added dropwise to ice water, and the mixture was extracted with ethyl acetate / water. The ethyl acetate layer was concentrated, and the crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase to obtain intermediate P2. P3 was synthesized using the same method as intermediate NTP0 (1-(5-chloro-4-nitrothiophen-2-yl)ethan-1-one), and NTP27 was synthesized using the same method as NTP1 to obtain the compound. Its NMR identification data are as follows:

[0202] 2-((2,3-dichlorophenyl)thio)-3-nitrobenzo[b]thiophen-7(4H)-one (2-((2,3-dichlorophenyl)thio)-3-nitrobenzo[b]thiophen-7(4H)-one, NTP27) pale yellow solid, 15% yield; ¹H NMR (400 MHz, DMSO-d6) 7.47 (dd, J = 7.9 Hz, 1H), 7.18 (dd, J = 7.9 Hz, 1H), 7.03 (s, 1H), 6.44 (m, 1H), 6.03 (dt, J = 7.3 Hz, 1H), 3.60 (dd, J = 7.3 Hz, 2H), HRMS calcd for [M+Na] + 393.9144, found 393.9142. The structure is as follows:

[0203] (NTP27).

[0204] (10) Synthesis of NTP28: The process is carried out according to the following reaction formula:

[0205]

[0206] Specifically, 100 mg (0.26 mmol) of NTP-43 was dissolved in 10 mL of dichloromethane. Then, 22.2 mg (0.26 mmol) and 39.7 g (0.39 mmol) of triethylamine were added, and the mixture was stirred at room temperature for 3 hours. The mixture was then extracted with dichloromethane / water, and the dichloromethane layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0207] N-(2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl)acrylamide(N-(2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl)acrylamide, NTP28), white solid, 38% yield; ¹H NMR (400 MHz, DMSO-d6) 8.41 (t, J = 7.3 Hz, 3H), 8.09 (s, 1H), 7.43 (dd, J = 7.9 Hz, 1H), 7.13 (dd, J = 7.9 Hz, 1H), 7.13 (t, J = 7.9 Hz, 1H), 6.48–6.08 (m, 3H), 5.09 (t, J = 7.3 Hz, 2H), HRMS calcd for [M+Na] + : 438.9359, found 438.9354. The structure is as follows:

[0208] (NTP28).

[0209] (11) Synthesis of NTP29, NTP30, NTP31, NTP32, and NTP33: The synthesis proceeds according to the following reaction formula:

[0210]

[0211] Specifically, 2 g (13.6 mmol) of 5-chlorothiophene-2-carboxaldehyde was dissolved in 20 mL of ethanol and stirred until completely dissolved. Then, 13.6 mmol of the corresponding substituted ketones were added: acetone (NTP29), acetylbenzene (NTP30), 2,4-pentanedione (NTP31), 1-phenylbutane-1,3-dione (NTP32), and 3-oxobutyric acid (NTP33), respectively. 27.2 mmol of sodium hydroxide was then added, and the mixture was stirred at room temperature for 24 h. The mixture was then extracted with dichloromethane / water, and the dichloromethane layer was concentrated. The crude product was purified by column chromatography using a petroleum ether / ethyl acetate system as the mobile phase to obtain product P1. P2 and the final product R were synthesized using the same method as for NTP0 and NTP1. 1 Substituted nitrated thiophene sulfides. Their NMR identification data are as follows:

[0212] (E)-4-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)but-3-en-2-one ((E)-4-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)but-3-en-2-one, NTP29), yellow solid, 23% yield; ¹H NMR (400 MHz, DMSO-d6) 8.06 (s, 1H), 7.66 (dd, J = 7.9 Hz, 1H), 7.52 (dd, J = 7.3 Hz, 1H), 7.36 (dd, J = 7.9 Hz, 1H), 7.13 (t, J = 7.9 Hz, 1H), 6.91 (dd, J = 7.3 Hz, 1H), 2.34 (s, 3H), HRMS calcd for [M+Na] + 395.9301, found 395.9299. The structure is as follows:

[0213] (NTP29).

[0214] (E)-3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-1-phenylprop-2-en-1-one ((E)-3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-1-phenylprop-2-en-1-one, NTP 30), yellow solid, 25% yield; ¹H NMR (400 MHz, DMSO-d6) 8.08 (s, 1H), 7.92 (dd, J = 7.3 Hz, 1H), 7.81–7.30 (m, 6H), 7.17 (dd, J = 7.9 Hz, 1H), 7.03 (t, J = 7.9 Hz, 1H), HRMS calcd for [M+Na] +: 457.9457, found 457.9453. The structure is as follows:

[0215] (NTP30).

[0216] (E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-1-phenylpent-4-ene-1,3-dione ((E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-1-phenylpent-4-ene-1,3-dione, NTP31), pale yellow solid, 25% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 8.04 (t, J = 7.9 Hz, 1H), 8.00 (t, J = 7.9 Hz, 1H), 7.68–7.50 (m, 5H), 7.10–6.98 (m, 3H), 4.43 (s, 2H), HRMS calcd for [M+Na] + 499.9563, found 499.9571. The structure is as follows:

[0217] (NTP31).

[0218] (E)-6-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)hex-5-ene-2,4-dione ((E)-6-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)hex-5-ene-2,4-dione, NTP32), pale yellow solid, 35% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.56 (dd, J = 7.3 Hz, 1H), 7.47 (dd, J = 7.9 Hz, 1H), 7.16 (dd, J = 7.9 Hz, 1H), 7.07 (t, J = 7.9 Hz, 1H), 6.92 (dd, J = 7.3 Hz, 1H), 4.45 (s, 2H), 2.41 (s, 3H), HRMS calcd for [M+Na] + : 437.9407, found 437.9410. The structure is as follows:

[0219] (NTP32).

[0220] (E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopent-4-enoic acid ((E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopent-4-enoic acid, NTP 33), pale yellow solid, 26% yield; ¹H NMR (400 MHz, DMSO-d6) 13.98 (s, 1H), 8.09 (s, 1H), 7.71 (dd, J=7.3 Hz, 1H), 7.41 (dd, J=7.9 Hz, 1H), 7.16 (dd, J=7.9 Hz, 1H), 7.11 (t, J=7.9 Hz, 1H), 6.87 (dd, J=7.3 Hz, 1H), 4.23 (s, 2H), HRMS calcd for [M+Na] + : 439.9199, found 439.9210. The structure is as follows:

[0221] (NTP33).

[0222] (12) Synthesis of NTP34: The process is carried out according to the following reaction formula:

[0223]

[0224] Specifically, 100 mg (0.24 mmol) was dissolved in 10 mL of dichloromethane, then 21 mg (0.29 mmol) of diethylamine and 29 mg (0.29 mmol) of triethylamine were added. After stirring at room temperature for 3 hours, the mixture was extracted with dichloromethane / water, and the dichloromethane layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0225] (E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N,N-diethyl-3-oxopent-4-enamide ((E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N,N-diethyl-3-oxopent-4-enamide, NTP34), pale yellow solid, yield 56%; HNMR (400MHz, DMSO-d6) 8.09 (s, 1H), 7.65 (dd, J=7.3Hz, 1H), 7.53 (dd, J=7.9Hz, 1H), 7.23 ( dd, J=7.9Hz, 1H), 7.12 (t, J=7.9Hz, 1H), 6.98 (dd, J=7.3Hz, 1H), 3.49 (m, 4H), 1.26 (m, 6H), HRMS calcd for [M+Na] + : 494.9985, found 494.9991. The structure is as follows:

[0226] (NTP34).

[0227] (13) Synthesis of NTP38 and NTP40: The synthesis is carried out according to the following reaction:

[0228]

[0229] 200 mg (0.49 mmol) of NTP37 was dissolved in 10 mL of toluene. 0.6 mmol of the corresponding diamines, diethylamine (NTP38) and di-n-propylamine (NTP40), were added, followed by 0.75 mmol of DMAP. The mixture was refluxed and heated to 110 °C, stirred for 12 h, and then extracted with dichloromethane / water. The dichloromethane layer was concentrated, and the crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0230] 3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N,N-diethyl-3-oxopropanamide (3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N,N-diethyl-3-oxopropanamide, NTP38), pale yellow solid, 39% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.47 (dd, J = 7.9 Hz, 1H), 7.17 (dd, J = 7.9 Hz, 1H), 7.09 (t, J = 7.9 Hz, 1H), 4.09 (s, 2H), 3.49 (m, 4H), 1.27 (m, 6H), HRMS calcd for [M+Na] + : 468.9829, found 468.9838. The structure is as follows:

[0231] (NTP38).

[0232] 3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxo-N,N-dipropylpropanamide (3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxo-N,N-dipropylpropanamide, NTP40), pale yellow solid, 39% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.47 (dd, J = 7.9 Hz, 1H), 7.17 (dd, J = 7.9 Hz, 1H), 7.09 (t, J = 7.9 Hz, 1H), 4.08 (s, 2H), 3.54 (t, 4H), 1.79–1.67 (m, 4H), 1.20–1.04 (m, 6H), HRMS calcd for [M+Na] + : 497.0142, found 497.0145. The structure is as follows:

[0233] (NTP40).

[0234] (14) Synthesis of NTP39: The reaction proceeds as follows:

[0235]

[0236] 200 mg (0.49 mmol) of NTP37 was dissolved in 10 mL of toluene, and 46 mg (0.49 mmol) of chloroacetamide and 52 mg (0.98 mmol) of sodium methoxide were added. The mixture was stirred at room temperature for 8 h, and the reaction solution was concentrated. The solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0237] methyl 4-amino-2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophene-2-carbonyl)-4-oxobutanoate (NTP 39), pale yellow solid, 65% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.45 (dd, J = 7.9 Hz, 1H), 7.15 (dd, J = 7.9 Hz, 1H), 7.09–7.00 (m, 3H), 3.83 (s, 3H), 2.85 (m, 1H), 2.60 (m, 1H); HRMS calcd for [M+Na] + : 484.9414, found 484.9408. The structure is as follows:

[0238] (NTP39).

[0239] (15) Synthesis of NTP41: The process proceeds according to the following reaction:

[0240]

[0241] 200 mg (0.49 mmol) of NTP37 was dissolved in 10 mL of toluene, and 79 mg (0.49 mmol) of 2,4,6-trifluorobenzaldehyde, 50 mg (0.59 mmol) of glacial acetic acid, and 35.5 mg (0.59 mmol) of piperidine were added. The mixture was refluxed at 120 °C for 6 h. The reaction solution was concentrated, and the mixture was extracted with ethyl acetate / water. The ethyl acetate layer was concentrated, and the crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0242] methyl(E)-2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophene-2-carbonyl)-4-(2,4,6-trifluorophenyl)but-3-enoate NTP41), orange-yellow solid, yield 47%; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.45 (dd, J=7.9 Hz, 1H), 7.15 (dd, J=7.9 Hz, 1H), 7.05 (t, J=7.9 Hz, 1H), 6.85 (m, 1H), 6.33 (m, 1H), 3.99 (dd, J=7.3 Hz, 1H), 3.86 (s, 3H), HRMScalcd for [M+Na] + : 583.9386, found 583.9393. The structure is as follows:

[0243] (NTP41).

[0244] (16) Synthesis of NTP36: The process proceeds according to the following reaction:

[0245]

[0246] 100 mg (0.25 mmol) of NTP37 was dissolved in 8 mL of methanol / tetrahydrofuran / water (volume ratio 1:1:1), and 24 mg (0.1 mmol) of lithium hydroxide was added. After stirring at room temperature for 5 h, 1 mL of 1N hydrochloric acid was added to acidify to neutral pH. Extraction was then performed using an ethyl acetate / water system. The ethyl acetate layer was collected, concentrated to remove the solvent, and the product was obtained. Its NMR identification data are as follows:

[0247] 3-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopropanoic acid (NTP 36), yellow solid, 78% yield. ¹H NMR (400 MHz, DMSO-d6) 8.08 (s, 1H), 7.45 (dd, J = 7.9 Hz, 1H), 7.15 (dd, J = 7.9 Hz, 1H), 7.03 (t, J = 7.9 Hz, 1H), 3.85 (s, 3H); HRMS calcd for [M+Na] +: 413.9043, found 413.9047. The structure is as follows:

[0248] (NTP36).

[0249] (17) Synthesis of NTP45 and NTP46: The synthesis proceeds according to the following reaction:

[0250]

[0251] 200 mg (0.49 mmol) of NTP37 was dissolved in 5 mL of DMF, and 0.49 mmol of the corresponding carboxylic acids 2-(2,6-dimethylphenyl)acetic acid (NTP45) and 2-(4-bromophenyl)acetic acid were added. Then, 7 mg (0.05 mmol) of DIC was added, and the mixture was stirred at room temperature for 6 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. Its NMR identification data are as follows:

[0252] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl2-(2,6-dimethylphenyl)acetate (2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl2-(2,6-dimethylphenyl)acetate, NTP 45), white solid, 57% yield; ¹H NMR (400 MHz, DMSO-d6) 8.08 (s, 1H), 7.50–7.40 (m, 2H), 7.19–7.08 (m, 2H), 7.02 (t, J = 7.9 Hz, 1H), 5.96 (s, 2H), 3.98 (s, 2H), 2.56 (s, 6H), HRMS calcd for [M+Na] + : 531.9825, found 531.9824. The structure is as follows:

[0253] (NTP45).

[0254] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl2-(4-bromophenyl)acetate (2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl2-(4-bromophenyl)acetate, NTP46), white solid, 67% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.83 (dd, J = 7.9 Hz, 2H), 7.52 (dd, J = 7.9 Hz, 1H), 7.33 (dd, J = 7.9 Hz, 1H), 7.22 (dd, J = 7.9 Hz, 1H), 7.04 (t, J = 7.9 Hz, 1H), 5.95 (s, 2H), 3.87 (s, 2H), HRMS calcd for [M+Na] + : 581.8617, found 581.8610. The structure is as follows:

[0255] (NTP46).

[0256] (18) Synthesis of NTP47 and NTP48: The synthesis proceeds according to the following reaction:

[0257]

[0258] 500 mg (2.3 mmol) of 4-bromophenylpropionic acid was dissolved in 10 mL of DMF. Then, 2.3 mmol of the corresponding substituted boric acids, 4-isopropylphenylboronic acid (NTP47) and pyridine-4-boronic acid (NTP48), were added. Next, 3.22 g (2.8 mmol) of Pd(PPh3)4 and 642 mg (4.7 mmol) of potassium carbonate were added. After stirring at room temperature for 12 h, the reaction mixture was extracted with ethyl acetate / water. The ethyl acetate layer was concentrated, and the crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase to obtain intermediate P1. Then, using NTP44 as a starting material, esterification was performed using the same method as for synthesizing NTP45 to obtain the final product. Its NMR identification data are as follows:

[0259] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl2-(4'-isopropyl-[1,1'-biphenyl]-4-yl)acetate (2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl 2-(4'-isopropyl-[1,1'-biphenyl]-4-yl)acetate, NTP47), pale yellow solid. , yield 45%; HNMR (400MHz, DMSO-d6) 8.09 (s, 1H), 7.63 (dd, J=7.9Hz, 2H), 7.52 (dd, J=7.9Hz, 2H), 7.40-7.28 (m, 5H) , 7.20 (dd, J=7.9Hz, 1H), 7.07 (t, J=7.9Hz, 1H), 5.95 (s, 2H), 3.76 (s, 2H), 2.77 (m, 1H), 1.67 (d, 7.3Hz, 6H), HRMS calcd for [M+Na] + : 622.0295, found 622.0288. The structure is as follows:

[0260] (NTP47).

[0261] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl2-(4-(pyridin-4-yl)phenyl)acetate (2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxoethyl2-(4-(4-pyridinyl)phenyl)acetate, NTP48), yellow solid, 33% yield; H NMR (400MHz, DMSO-d6) 8.87 (d, J=7.9Hz, 2H), 8.09 (s, 1H), 8.00 (d, J=7.9Hz, 2H), 7.63 (dd, J=7.9Hz, 1H) , 7.41-7.28 (m, 3H), 7.18 (dd, J=7.9Hz, 1H), 7.01 (t, J=7.9Hz, 1H), 5.88 (s, 2H), 3.72 (s, 2H), HRMScalcd for [M+Na] + : 580.9778, found 580.9787. The structure is as follows:

[0262] (NTP48).

[0263] (19) Synthesis of NTP49, NTP50, NTP51, NTP52, and NTP53: The synthesis proceeds according to the following reaction formula:

[0264]

[0265] 100 mg (0.48 mmol) of NTP33 was dissolved in 10 mL of DCM. The corresponding amines (0.48 mmol each): ammonia-methanol solution (NTP49), isopropylamine (NTP50), n-butylamine (NTP51), cyclohexylamine (NTP52), and di-n-propylamine (NTP53) were added. Then, HATU (0.57 mmol) and triethylamine (0.72 mmol) were added. The mixture was stirred at room temperature for 3 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0266] (E)-N-(2-aminoethyl)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopent-4-enamide ((E)-N-(2-aminoethyl)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopent-4-enamide, NTP49), yellow solid, 33% yield; HNMR ( 400MHz, DMSO-d6) 8.05 (s, 1H), 8.01 (t, J=7.3Hz, 2H), 7.65 (dd, J=7.3Hz, 1H), 7.47 (dd, J=7.9Hz, 1H), 7.1 7 (dd, J=7.9Hz, 1H), 7.03 (t, J=7.9Hz, 1H), 4.91 (s, 2H), 4.22 (s, 2H), 3.75 (m, 2H), 2.66 (m, 2H), HRMScalcd for [M+Na] + : 481.9781, found 481.9788. The structure is as follows:

[0267] (NTP49).

[0268] (E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N-isopropyl-3-oxopent-4-enamide ((E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N-isopropyl-3-oxopent-4-enamide, NTP50), yellow solid, yield 42%; ¹H NMR (400 M) Hz, DMSO-d6) 8.14 (s, 2H), 8.05 (s, 1H), 7.56 (dd, J=7.3Hz, 1H), 7.45 (dd, J=7.9Hz, 1H), 7.15 (dd, J=7. 9Hz, 1H), 7.03 (t, J=7.9Hz, 1H), 6.91 (dd, J=7.3Hz, 1H), 3.93-3.78 (m, 3H), 1.19 (d, J=7.3Hz, 6H), HRMS calcd for [M+Na] + : 480.9729, found 480.9722. The structure is as follows:

[0269] (NTP50).

[0270] (E)-N-butyl-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopent-4-enamide ((E)-N-butyl-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxopent-4-enamide, NTP51), yellow solid, 35% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 8.00 (s, 1H), 7.58 (dd, J=7.3Hz, 1H), 7.45 (dd, J=7.9Hz, 1H), 7.15 (dd, J=7.9Hz, 1H), 7.08 (t, J=7.9Hz, 1 H), 6.92 (dd, J=7.3Hz, 1H), 3.93 (s, 2H), 3.22 (t, J=7.3Hz, 2H), 1.56-1.43 (m, 4H), 1.21 (t, J=7.3Hz, 3H), HRMS calcd for [M+Na] + : 495.9985, found 495.9989. The structure is as follows:

[0271] (NTP51).

[0272] (E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N,N-diethyl-3-oxopent-4-enamide ((E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N,N-diethyl-3-oxopent-4-enamide, NTP52), pale yellow solid, 35% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.63 (dd, J=7.3Hz, 1H), 7.44 (dd, J=7.9Hz, 1H), 7.14 (dd, J=7.9Hz, 1H), 7.0 7 (t, J=7.9Hz, 1H), 6.94 (dd, J=7.3Hz, 1H), 3.93 (s, 2H), 3.36 (m, 4H), 1.21 (t, J=7.3Hz, 6H), HRMScalcd for [M+Na] + 494.9985, found 494.9978. The structure is as follows:

[0273] (NTP52).

[0274] (E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxo-N,N-dipropylpent-4-enamide ((E)-5-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-oxo-N,N-dipropylpent-4-enamide, NTP53), pale yellow solid, 35% yield; ¹H NMR (400 MHz, DM) SO-d6) 8.09 (s, 1H), 7.63 (dd, J=7.3Hz, 1H), 7.44 (dd, J=7.9Hz, 1H), 7.14 (dd, J=7.9Hz, 1H), 7.07 (t, J=7.9H z, 1H), 6.94 (dd, J=7.3Hz, 1H), 3.93 (s, 2H), 3.25 (t, J=7.3Hz, 4H), 1.65 (m, 4H), 1.08 (t, J=7.3Hz, 6H), HRMS calcd for [M+Na] + : 523.0298, found 523.0296. The structure is as follows:

[0275] (NTP53).

[0276] (20) Synthesis of NTP58 and NTP59: The synthesis is carried out according to the following reaction:

[0277]

[0278] Specifically, 98 mg (0.25 mmol) of NTP60 was dissolved in 2 ml of toluene, and 0.25 mmol of the corresponding amines, n-propylamine (NTP58) and isopropylamine (NTP59), were added respectively. Then, 0.37 mmol of DMAP was added, and the mixture was refluxed at 110 °C with stirring for 5 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. Its NMR identification data are as follows:

[0279] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-oxo-N-propylacetamide (NTP 58), white solid, 35% yield; ¹H NMR (400 MHz, DMSO-d6) 8.90 (s, 1H), 8.09 (s, 1H), 7.46 (dd, J = 7.9 Hz, 1H), 7.16 (dd, J = 7.9 Hz, 1H), 7.10 (t, J = 7.9 Hz, 1H), 3.17 (t, J = 7.3 Hz, 2H), 2.45 (m, 2H), 1.25 (t, J = 7.3 Hz, 3H); HRMS calcd for [M+Na] + : 440.9516, found 440.9520. The structure is as follows:

[0280] (NTP58).

[0281] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N-isopropyl-2-oxoacetamide (NTP 59), white solid, 43% yield; ¹H NMR (400 MHz, DMSO-d6) 8.80 (s, 1H), 8.09 (s, 1H), 7.46 (dd, J = 7.9 Hz, 1H), 7.16 (dd, J = 7.9 Hz, 1H), 7.10 (t, J = 7.9 Hz, 1H), 4.45 (m, 1H), 1.43 (d, J = 7.3 Hz, 6H); HRMS calcd for [M+Na] +: 440.9516, found 440.9512. The structure is as follows:

[0282] (NTP59).

[0283] (21) Synthesis of NTP68 and NTP69: The synthesis proceeds according to the following reaction:

[0284]

[0285] 200 mg (0.47 mmol) of NTP was dissolved in 15 mL of DMF, and 0.47 mmol of the corresponding boric acids, 4-isopropylphenylboronic acid (NTP68) and pyridine-4-boronic acid (NTP69), were added. Then, 812 mg (0.70 mmol) of Pd(PPh3)4 and 129 mg (0.94 mmol) of potassium carbonate were added. The mixture was stirred at room temperature for 3 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase to obtain the product 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-y l)-2-(4-isopropylphenyl)ethan-1-one ((5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-(4-isopropylphenyl)ethyl ketone, NTP 68), white solid, 52% yield; ¹H NMR (400 MHz, DMSO-d6) 8.09 (s, 1H), 7.49–7.39 (m, 3H), 7.18–7.05 (m, 4H), 4.65 (s, 2H), 2.86 (m, 1H), 1.23 (d, J = 7.3 Hz, 6H); HRMS calcd for [M+Na] + 487.9927, found 487.9932. The structure is as follows:

[0286] (NTP68).

[0287] 1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-(pyridin-4-yl)ethan-1-one (1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-(4-pyridyl)ethyl ketone, NTP 69), yellow solid, 43% yield; ¹H NMR (400 MHz, DMSO-d6) 8.56 (dd, J = 7.9 Hz, 2H), 8.09 (s, 1H), 7.49 (dd, J = 7.9 Hz, 1H), 7.20–7.14 (m, 3H), 7.09 (dd, J = 7.9 Hz, 1H), 5.46 (s, 2H); HRMS calcd for [M+Na] + : 446.9410, found 446.9415. The structure is as follows:

[0288] (NTP69).

[0289] (22) Synthesis of NTP70, NTP71, and NTP72: The synthesis proceeds according to the following reaction formula:

[0290]

[0291] 200 mg (0.43 mmol) of NTP68 was dissolved in 10 mL of ethanol, and 0.43 mmol of the corresponding aldehydes were added: 2-(furan-2-yl)acetaldehyde (NTP70), 2-(2,4,6-trifluorophenyl)acetaldehyde (NTP71), and 2-(4-methoxyphenyl)acetaldehyde (NTP72). Then, 34 mg (0.86 mmol) of sodium hydroxide was added, and the reaction mixture was stirred at room temperature for 4 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. The NMR identification data are as follows:

[0292] (E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-4-(furan-2-yl)-2-(4-isopropylphenyl)but-2-en-1-one ((E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-4-(furan-2-yl)-2-(4-isopropylphenyl)but-2-en-1-one, NTP70), yellow solid, 43% yield; H NMR (400MHz, DMSO-d6) 8.03 (s, 1H), 7.49-7.31 (m, 6H), 7.20 (dd, J=7.9Hz, 1H), 7.09 (dd, J=7.9Hz, 1H), 6.54 (t, J=7.3 HRMS calcd for [M+Na] + : 580.0189, found 580.0191. The structure is as follows:

[0293] (NTP70).

[0294] (E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-(4-isopropylphenyl)-4-(2,4,6-trifluorophenyl)but-2-en-1-one((E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-(4-isopropylphenyl)-4-(2,4,6-trifluorophenyl)but-2-en-1-one, NT P71), yellow solid, yield 54%; ¹H NMR (400 MHz, DMSO-d⁶): 8.03 (s, 1H), 7.49–7.31 (m, 6H), 7.22 (dd, J = 7.9 Hz, 1H), 7.16 (dd, J = 7.3 Hz, 1H), 7.08 (dd, J = 7.9 Hz, 1H), 6.45 (t, J = 7.9 Hz, 2H), 5.00 (m, 1H), 3.33 (d, J = 7.3 Hz, 1H), 1.20 (d, J = 7.3 Hz, 6H); HRMS calcd for [M+Na] + : 644.0114, found 644.0122. The structure is as follows:

[0295] (NTP71).

[0296] (E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-(4-isopropylphenyl)-4-(4-methoxyphenyl)but-2-en-1-one ((E)-1-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-2-(4-isopropylphenyl)-4-(4-methoxyphenyl)but-2-en-1-one, NTP72), yellow solid Body, yield 53%; HNMR (400MHz, DMSO-d6) 8.03 (s, 1H), 7.49-7.31 (m, 6H), 7.22 (dd, J=7.9Hz, 1H), 7.16-7.12 (m, 4H), 7.08 (d HRMS calcd for [M+Na] + : 620.0502, found 620.0500. The structure is as follows:

[0297] (NTP72).

[0298] (23) Synthesis of NTP65 and NTP66: The synthesis proceeds according to the following reaction:

[0299]

[0300] Specifically, 100 mg (0.24 mmol) of NTP62 was dissolved in 10 mL of toluene, and 0.24 mmol of the corresponding amine, either isopropylamine (NTP65) or 4-methoxy-2,6-dimethylaniline (NTP66), was added. Then, DMAP (0.36 mmol) was added, and the mixture was refluxed at 110 °C for 12 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. Its NMR identification data are as follows:

[0301] 4-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N-isopropyl-4-oxobutanamide (4-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N-isopropyl-4-oxobutanamide, NTP65), pale yellow solid, 41% yield; ¹H NMR (400 MHz, DMSO) -d6) 8.15 (s, 1H), 8.06 (s, 1H), 7.49 (dd, J=7.9Hz, 1H), 7.22 (dd, J=7.9Hz, 1H), 7.19 (dd, J=7.9Hz, 1H), 7.10 (dd, J=7.9Hz, 1H), 3.81-3.65 (m, 3H), 2.58 (t, J=7.3Hz, 2H), 1.23 (d, J=7.3Hz, 6H); HRMS calcd for [M+Na] + 468.9829, found 468.9834. The structure is as follows:

[0302] (NTP65).

[0303] 4-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N-(4-methoxy-2,6-dimethylphenyl)-4-oxobutanamide (4-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-N-(4-methoxy-2,6-dimethylphenyl)-4-oxobutanamide, NTP66), pale yellow solid, yield 42%; ¹H NMR ( 400MHz, DMSO-d6) 9.74 (s, 1H), 8.06 (s, 1H), 7.49 (dd, J=7.9Hz, 1H), 7.22 (dd, J=7.9Hz, 1H), 7.19 (dd, J=7.9Hz, HRMS calcd for [M+Na] + : 561.0091, found 561.0094. The structure is as follows:

[0304] (NTP66).

[0305] (24) Synthesis of NTP64: The process proceeds according to the following reaction:

[0306]

[0307] Specifically, 100 mg (0.25 mmol) of NTP63 was dissolved in 10 mL of methanol solution, and then 2 mL of ammonia in methanol solution (7 mol / L) was added. The mixture was stirred at room temperature for 12 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. Its NMR identification data are as follows:

[0308] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-5-methyl-1H-pyrrole (2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-5-methyl-1H-pyrrole, NTP64), white solid, 51% yield; ¹H NMR (400 MHz, DMSO-d6) 11.34 (s, 1H), 8.34 (s, 1H), 7.44 (dd, J=7.9 Hz, 1H), 7.23 (dd, J=7.9 Hz, 1H), 7.14 (dd, J=7.9 Hz, 1H), 6.31 (dd, J=7.9 Hz, 1H), 5.88 (dd, J=7.9 Hz, 1H), 2.34 (s, 3H); HRMS calcd for [M+Na] + : 406.9461, found 406.9467. The structure is as follows:

[0309] (NTP64).

[0310] (25) Synthesis of NTP61: The process proceeds according to the following reaction:

[0311]

[0312] Specifically, 100 mg (0.25 mmol) of NTP60 was dissolved in 10 ml of acetonitrile, and then 22 mg (0.25 mmol) of 3-aminobutane-2-one was added. The mixture was stirred at 80 °C for 12 h. The reaction solution was extracted with ethyl acetate / water, and the ethyl acetate layer was concentrated. The crude product was purified by column chromatography using petroleum ether / ethyl acetate as the mobile phase. Its NMR identification data are as follows:

[0313] 2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-methoxy-4,5-dimethyl-2,3-dihydro-1H-pyrrole (2-(5-((2,3-dichlorophenyl)thio)-4-nitrothiophen-2-yl)-3-methoxy-4,5-dimethyl-2,3-dihydro-1H-pyrrole, NTP61) White solid, yield 26%; ¹H NMR (400 MHz, DMSO-d⁶) 7.48 (dd, J = 7.9 Hz, 1H), 7.23–7.15 (m, 3H), 7.03 (dd, J = 7.9 Hz, 1H), 4.31 (d, J = 7.3 Hz, 1H), 4.16 (d, J = 7.3 Hz, 1H), 3.76 (s, 3H), 2.26 (s, 3H), 1.78 (s, 3H); HRMS calcd for [M+Na] + : 452.9879, found 452.9883. The structure is as follows:

[0314] (NTP61).

[0315] Example 3

[0316] The activities of each compound prepared in Example 2 were evaluated. The activity evaluation employed a reporter gene assay based on THP1-Luc cells. The specific steps were as follows: THP1-Luc cells were seeded at a density of 100,000 cells / well in 12-well plates. Then, 1 μM or 10 μM of the compound (H151 as a control) was added for pre-incubation for 2 hours, followed by the addition of 100 nM S-cddA, and incubation for a total of 12 hours. After incubation, the cell suspension was transferred to a 1.5 mL centrifuge tube and centrifuged at 1500 rpm for 3 minutes at room temperature. The supernatant was discarded. After washing with 1 mL of PBS buffer, the centrifugation and supernatant discarding steps were repeated. Subsequent steps were performed using a René luciferase reporter gene assay kit (Beyotime).

[0317] (1) Cell lysis: Add 300 μL of cell lysis buffer to the cell pellet and lyse at room temperature for 30 min. Then, centrifuge at 12000 rpm for 3 min at room temperature to obtain the supernatant cell lysis buffer.

[0318] (2) Preparation of diluent for Renal luciferase detection: The diluent is prepared during the previous lysis step. The diluent is prepared at a volume ratio of 1:100 between the Renal luciferase detection substrate and the working solution. The process is carried out on ice, and subsequent steps must be completed under light-protected conditions.

[0319] (3) Detection of reporter genes: Take a white 96-well plate and add 80 μL of the cell lysis buffer to be tested to each well (for the blank group, add an equal volume of lysis buffer not used for cell lysis). Then, quickly add 100 μL of René luciferase detection diluent using a pipette, ensuring that the diluent addition step is controlled within 3 minutes. Gently shake the 96-well plate and immediately place it in a multi-mode microplate reader to test the chemiluminescence reading.

[0320] (4) Data processing: The expression level of the reporter gene was characterized by chemiluminescence intensity, which reflects the relative expression level of IFN-β1. The relative light unit (RLU) was obtained by subtracting the blank group reading from the readings of the control group and experimental group. The RLU value of the control group was corrected to 100%. The corrected readings of the experimental group were the fold increase of the target gene expression relative to the control group.

[0321] The inhibition rates (%) of each compound are shown in Table 1 below. The results show that each compound has different inhibitory activities against IFN-β1, among which compounds NTP14 and NTP16 have the most significant activities. At a concentration of 10 μM, they can show significantly higher inhibitory efficacy than NTP1 (P005091), and can achieve complete inhibition of the IFN-β1 response, making them potentially highly active molecules.

[0322] Table 1 compound Inhibition rate (1μM) Inhibition rate (10 μM) compound Inhibition rate (1μM) Inhibition rate (10 μM) H-151 35.64 86.46 NTP-37 22.32 75.56 NTP-1 -4.49 79.52 NTP-38 17.85 69.03 NTP-2 -6.14 11.89 NTP-39 3.38 68.87 NTP-3 -8.65 16.00 NTP-40 11.51 76.71 NTP-4 -1.84 19.10 NTP-41 13.28 75.52 NTP-5 -6.97 28.06 NTP-42 23.34 77.83 NTP-6 -6.87 28.99 NTP-43 4.67 76.59 NTP-7 -7.69 -0.16 NTP-44 3.23 83.28 NTP-8 -2.70 13.49 NTP-45 16.78 76.86 NTP-9 3.65 13.77 NTP-46 6.98 70.06 NTP-10 -0.48 15.15 NTP-47 14.45 73.80 NTP-11 -2.35 2.50 NTP-48 12.76 70.92 NTP-12 -5.34 -7.49 NTP-49 -3.54 74.50 NTP-13 -2.33 -0.49 NTP-50 4.34 82.56 NTP-14 21.87 109.79 NTP-51 5.09 78.70 NTP-15 2.78 56.11 NTP-52 11.11 77.68 NTP-16 24.81 105.98 NTP-53 7.80 75.63 NTP-17 2.61 21.48 NTP-54 14.78 83.41 NTP-18 22.93 71.54 NTP-55 13.32 77.84 NTP-19 31.13 94.71 NTP-56 8.89 75.09 NTP-20 5.48 11.27 NTP-57 12.22 87.43 NTP-21 1.79 -8.43 NTP-58 2.13 75.45 NTP-22 10.99 70.76 NTP-59 -4.55 81.43 NTP-23 5.34 75.22 NTP-60 -0.67 70.87 NTP-24 3.83 75.80 NTP-61 12.56 75.40 NTP-25 -0.98 75.19 NTP-62 9.99 73.89 NTP-26 0.58 82.36 NTP-63 14.22 81.15 NTP-27 12.11 76.39 NTP-64 -5.51 78.98 NTP-28 7.58 76.39 NTP-65 4.78 72.87 NTP-29 -2.21 83.81 NTP-66 1.14 88.89 NTP-30 10.09 74.92 NTP-67 10.09 84.62 NTP-31 7.67 71.28 NTP-68 4.67 81.17 NTP-32 5.48 83.90 NTP-69 -0.83 76.76 NTP-33 -0.92 74.45 NTP-70 4.32 75.53 NTP-34 -4.67 71.08 NTP-71 6.78 83.34 NTP-35 7.87 74.20 NTP-72 -2.98 73.46 NTP-36 21.25 85.98

[0323] Example 4

[0324] In the aforementioned examples, the bioactivity of NTP1 was evaluated in various cell types, revealing its good inhibitory ability against type I interferon. Therefore, an acute enteritis model (DSS model) was established using C57BL / 6 mice to evaluate the anti-inflammatory capacity of NTP1. Specifically, 6-week-old C57BL / 6 mice were given 3% dextran sulfate sodium salt (DSS) for 10 consecutive days to induce acute enteritis. During this period, 10 mg / kg of NTP1 was injected intraperitoneally daily, and the weight changes of each mouse were recorded. See the schematic diagram of the experimental procedure. Figure 7 On day 11, the mice were sacrificed, their colons were removed and their length measured, blood was collected, and serum was extracted. The results showed that, compared to the control group, the mice in the DSS model group had significantly lower body weight and significantly shorter colon length, while the mice injected with NTP1 showed some recovery in body weight and colon length. These differences were statistically significant (see [link to relevant documentation]). Figure 8-10 ).

[0325] To assess the effects of the experimental intervention on serum inflammatory factors in model animals, enzyme-linked immunosorbent assay (ELISA) was used to quantitatively analyze serum samples from each group. Compared with the blank control group, the serum concentrations of IFN-β1, CXCL10, and IL-6 were increased in the disease model group (DSS group) (see [link to data]). Figure 11 This result confirms that the modeling operation successfully induced the expected innate immune activation or inflammatory response. Within the DSS group, compared with the untreated model control group, the serum inflammatory factor levels in the NTP1-treated group were significantly reduced (p < 0.05). H&E staining of mouse colon sections showed that the colon tissue of the DSS model group mice exhibited typical pathological features, and the semi-quantitative H&E score based on these features was increased. After NTP1 intervention, the above-mentioned pathological damage in the treated group was significantly alleviated, and the histological score was significantly reduced (p < 0.05). Immunohistochemical scoring analysis of colon tissue revealed that the scores of key inflammatory factors (P-IRF3, P-STAT3) were upregulated in the DSS group, while the scores were significantly reduced after administration (see [link to relevant documentation]). Figure 12-13 In summary, these results indicate that NTP1 can effectively inhibit abnormal immune activation induced by an acute intestinal inflammation model, demonstrating potential immunomodulatory or anti-inflammatory effects.

[0326] Subsequently, NTP14 and NTP16, which exhibited significant activity, were used as representative compounds for in-depth activity experiments. NTP14 and NTP16 are two small molecule compounds modified by replacing the sulfur linkages between the rings of NTP1 with oxygen linkages. Specifically, the acetyl side chain of NTP16 was replaced with methyl formate. This example explores their regulatory effects on the transcriptional and activation levels of key proteins in the IFNβ1 and CXCL10 gene pathways, providing a basis for the discovery of small molecules involved in immune regulation.

[0327] In THP-1 ISG reporter cells, after S-cddA (400 nM) activates the pathway, different compounds inhibit relative luciferase activity in a concentration-dependent manner (0.5-20 μM), and the inhibitory efficacy of each compound varies. For example, NTP14 and NTP16 show significant inhibitory effects at specific concentrations (see 14-15).

[0328] In BJ cells, after diABZI activation of the pathway, NTP14 can inhibit IFNβ1 and CXCL10 mRNA expression in a concentration-dependent manner, IC50... 50 The concentrations reached 0.2865 μM and 0.3420 μM, respectively; under NTP16 action, IFNβ1 and CXCL10 mRNA inhibited IC50 by 0.2865 μM and 0.3420 μM, respectively. 50The concentrations were 0.4786 μM and 0.2212 μM, respectively, demonstrating their inhibitory effect on transcriptional activation of this cellular pathway. In DC2.4 cells, after S-cddA activation, NTP14 inhibited the IC50 of IFNβ1 and CXCL10 mRNA. 50 Low concentrations of 0.0445 μM and 0.0830 μM; NTP16 exhibits stronger inhibition performance, corresponding to IC... 50 The concentrations were 0.0198 μM and 0.0064 μM, indicating that the compounds had different effects on transcriptional regulation of different cell lines, and that NTP16 had a more significant inhibitory effect in DC2.4 cells (see [link to relevant documentation]). Figure 16-17 ).

[0329] In the aforementioned experiments, the bioactivity of NTP14 and NTP16 was evaluated in BJ cells, and both showed good inhibitory activity against type I interferon. Consistent with the experimental animal disease model used to evaluate NTP1, a DSS-induced colitis mouse model was used to investigate its regulatory effects on intestinal tissue damage, expression of inflammation-related genes, and activation of pathway proteins, in order to clarify the potential value of the compound in the intervention of intestinal inflammation.

[0330] Results of colon morphology and length experiments in mice showed that in the DSS-induced colitis model (DSS+NS group), the colon of mice was significantly shortened and morphologically abnormal; while after intervention with NTP14 and NTP16 (DSS+NTP14, DSS+NTP16 groups), the colon length was restored and the morphology was closer to normal (Ctrl, NTP14, NTP16 groups). Statistical analysis showed that the compound significantly improved DSS-induced colon shortening, and the difference was statistically significant. In addition, the body weight of mice in the DSS+NS group decreased significantly over time; the trend of body weight loss in the DSS+NTP14 and DSS+NTP16 groups was slowed down, and the difference in body weight compared with the DSS+NS group was statistically significant, indicating that the compound can alleviate DSS-induced weight loss (see...). Figure 18-20 Histopathological results showed that inflammatory lesions were observed in the colonic tissue in the DSS+NS group; after treatment with NTP14 and NTP16, histopathological damage was reduced, glandular structure was more intact, and H&E score was significantly reduced (p<0.0001), verifying the protective effect of the compound on intestinal tissue (see [link]). Figure 21-22 Quantitative analysis of serum samples from each group using ELISA showed that both NTP14 and NTP16 could reduce the elevated serum IFN-β1, CXCL10, and IL-6 concentrations caused by acute inflammatory bowel disease (see [link to ELISA]). Figure 23 Furthermore, the above experiments were repeated using other compounds from Example 2, exhibiting similar therapeutic activity (results not shown).

[0331] In summary, based on topological analysis of the P005091 structure, this invention designed and synthesized a series of structurally modified compounds of P005091, providing a foundation for analyzing the structure-activity relationship of their inhibitory activity. The activity of the modified molecules was evaluated using qPCR and Renal luciferase reporter gene assays, laying the groundwork for further structural optimization. Overall, the nitrothiophene phenyloxythioether compounds synthesized in this invention can inhibit the transcription of inflammatory factors such as type I interferon, and exert an in vivo intestinal protective effect by improving colon morphology, body weight, and histopathological damage in DSS-induced colitis mice. This invention provides a basis for elucidating the mechanism of action of small molecule compounds in intervening intestinal inflammation and also provides potential targets and candidate molecules for the development of treatment strategies for diseases such as inflammatory bowel disease.

[0332] The above detailed embodiments provide a specific description of the analytical methods involved in this invention. It should be noted that the above description is only intended to help those skilled in the art better understand the methods and ideas of this invention, and is not intended to limit the scope of the invention. Without departing from the principles of this invention, those skilled in the art can make appropriate adjustments or modifications to this invention, and such adjustments and modifications should also fall within the protection scope of this invention.

Claims

1. A nitthiophene phenyl oxysulfide compound or a pharmaceutically acceptable salt, solvate, or prodrug thereof, comprising its tautomers, cis or trans isomers, meso compounds, racemic compounds, enantiomers, diastereomers, or mixtures thereof, wherein the structure of the nitthiophene phenyl oxysulfide compound is shown in Formula (I): （I）； Where R 1 It is selected from one of hydrogen, halogen, alkyl, alkoxy, haloalkyl, and amino; R 2 It is selected from one of hydrogen, halogen, alkyl, alkoxy, haloalkyl, and amino; R 3 Selected from one of hydrogen, alkyl, alkoxy, and alkylamino; R 4 Selected from hydrogen and halogen; R 5 Selected from , , , , , , , , One of them; Among them G 1 Selected from hydrogen, hydroxyl, alkyl, alkenyl, alkynyl, alkoxy, -NH-(CH2)4, -NHC2H4NH2, -NHC2H4NHCOOC(CH3)3, 2-acrylamidomethyl, , , One of the following; the alkyl, alkenyl or alkynyl group is either unsubstituted or substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl and heteroaryl; J 1 It is selected from one of alkyl, phenyl, benzoylmethyl, acetylmethyl, carboxymethyl, and (N,N-dimethylcarbamoyl)methyl. K 1 Selected from hydrogen, carboxyformamide group, One of them, K 2 It is selected from alkyl, hydroxy, alkoxy, and amino; the alkyl or amino group is not substituted or is substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl. Q 1 Selected from , , , One of them; T 1 It is selected from alkyl, amino, N-(2-aminoethyl)amino, -NH-nBu; the alkyl or amino group is not substituted or is substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl and heteroaryl; U 1 It is selected from one of alkyl, halogen, alkoxy, and amino; the alkyl or amino group is not substituted or is substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl. W 1 Selected from alkyl, , One of the following; the alkyl and amino groups are either unsubstituted or substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl; X 1 Selected from alkyl, alkoxy, amino, One of the following; wherein the alkyl, alkoxy, and amino groups are unsubstituted or substituted by one or more substituents selected from halogen, alkyl, alkoxy, haloalkyl, amino, cyano, hydroxy, hydroxyalkyl, cycloalkyl, heterocyclic, aryl, and heteroaryl; Y 1 Selected from , , One of them; R 6 Selected from H, alkyl, and alkylene groups; R 5 and R 6 These are either independently substituted substituents or connected to form a five-membered ring or a six-membered ring; X is selected from O and S; The compound of formula (I) is not one of the following compounds: 。 2. The nitrothiophene phenyl oxysulfide compound according to claim 1, or its pharmaceutically acceptable salt, solvate, or prodrug, characterized in that, The R 5 Selected from , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , One of them.

3. The nitrothiophene phenyl oxysulfide compound according to claim 1, or its pharmaceutically acceptable salt, solvate, or prodrug, characterized in that, The alkyl group is C 1-6 Alkyl; the alkylene group is C10. 1-6 Alkylene.

4. The nitrothiophene phenyl oxysulfide compound according to claim 1, or its pharmaceutically acceptable salt, solvate, or prodrug, characterized in that, The alkenyl group is C 2-6 alkenyl group; the alkenyl group is C 2-6 Alynyl group.

5. The nitrothiophene phenyl oxysulfide compound according to claim 1, or its pharmaceutically acceptable salt, solvate, or prodrug, characterized in that, The alkoxy group is C 1-6 alkoxy; the haloalkyl group is C 1-6 Halogenated alkyl; the hydroxyalkyl group is C10. 1-6 Hydroxyalkyl; the cycloalkyl group is a 3- to 6-membered cycloalkyl group.

6. The nitrothiophene phenyl oxysulfide compound according to claim 1, or its pharmaceutically acceptable salt, solvate, or prodrug, characterized in that, The aryl group is a 6- to 10-membered aryl group; the heteroaryl group is a 5- to 10-membered heteroaryl group.

7. The nitrothiophene phenyl oxysulfide compound according to claim 1, or its pharmaceutically acceptable salt, solvate, or prodrug, characterized in that, The nitthiopheneylphenyloxysulfide compounds are selected from one or more of the following structural formulas: 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 、 。 8. A pharmaceutical composition, characterized in that, It includes the nitrothiophene phenyl oxythioether compound according to any one of claims 1-7 or its pharmaceutically acceptable salt, solvate or prodrug, comprising its tautomer, cis or trans isomer, meso compound, racemic compound, enantiomer, diastereomer or mixture, and one or more active ingredients for treating inflammatory bowel disease.

9. The nitrothiophene phenyl oxythioether compound or a pharmaceutically acceptable salt, solvate or prodrug according to any one of claims 1-7, comprising the use of its tautomers, cis or trans isomers, meso compounds, racemic compounds, enantiomers, diastereomers or mixtures thereof in the preparation of a product for inhibiting inflammatory bowel disease.

10. The application according to claim 9, characterized in that, The inflammatory bowel disease is selected from one or more of Crohn's disease and ulcerative colitis.

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