CN110981868B - Imidazopyridine compound, pharmaceutical composition containing compound, preparation method and application thereof - Google Patents

Abstract

The invention relates to the technical field of medicinal chemistry and pharmacotherapeutics, and particularly discloses an imidazo [1,2-a ] pyridine compound and a pharmaceutically acceptable salt or a pharmaceutically acceptable solvate thereof, and further discloses a preparation method, a pharmaceutical composition containing the compound and application of the compound.

Description

Imidazopyridine compound, pharmaceutical composition containing compound, preparation method and application thereof

Technical Field

The invention relates to the field of pharmaceutical chemistry and pharmacotherapeutics, in particular to an imidazopyridine compound, a pharmaceutical composition containing the compound, and a preparation method and application of the compound.

Technical Field

Signal Transduction and Activator of Transcription (STAT) are bifunctional proteins with dual functions of signal transduction and activation of transcription. After being activated by different cytokines and growth factor receptors, the genes undergo phosphorylation and dimerization, translocate from cytoplasm into nucleus, and are combined with DNA to regulate and control the transcription and expression of corresponding target genes. As an important member of the STAT family, STAT3 is responsible for regulating a series of important physiological processes such as cell growth, proliferation, differentiation, and apoptosis. However, studies find that STAT3 has persistent activation and abnormally high expression, can induce cell proliferation, invasion and migration, inhibit apoptosis, promote angiogenesis, and play an important role in the process of tumor generation, development and metastasis. Therefore, the development of targeted STAT3 inhibitors has been a hot spot in the frontier of antitumor drug research.

In addition, STAT3 can interact with other tumor-related molecules on the cell surface, thereby crosslinking, activating and amplifying the related effects of the tumor, and promoting the occurrence, development and metastasis of the tumor to a great extent. Inhibition of the STAT3 signaling pathway may therefore inhibit multiple tumor targets from exerting an effect. The STAT3 inhibitor can simultaneously act on an EGFR (epidermal growth factor receptor) to exert effects, and when the inhibitor is combined with the EGFR inhibitor, the generation of EGFR drug acquired resistance can be delayed, the clinical service life of the EGFR drug acquired resistance can be prolonged, and the STAT3 inhibitor has important clinical significance.

In conclusion, research on specific inhibitors targeting STAT3 has been a research and development hotspot, but the drugs developed so far all have the defects of small drug effect, low selectivity, poor drug forming property and the like in different degrees and different aspects, and limit clinical application and later development of STAT3 inhibitors. While STAT3 is a promising tumor treatment target, the development of a new class of compounds capable of specifically inhibiting STAT3 is urgently needed.

Disclosure of Invention

The invention aims to solve the problems of small drug effect, low selectivity, poor efficiency and the like of a specific inhibitor of STAT3 in the prior art, and provides an imidazo [1,2-a ] pyridine compound.

The second purpose of the invention is to provide a preparation method of the imidazo [1,2-a ] pyridine compound.

The third purpose of the invention is to provide the application of the imidazo [1,2-a ] pyridine compound.

The purpose of the invention is realized by the following technical scheme:

an imidazo [1,2-a ] pyridine compound, which has a structural general formula shown in formula (I):

wherein n is 0, 1 or 2;

R₁、R₂、R₃、R₄each independently selected from hydrogen, halogen, cyano, nitro, amino, hydroxy, trifluoromethyl, substituted or unsubstituted C1-6 alkyl, substituted or unsubstituted C1-6 alkoxy, substituted or unsubstituted C1-6 alkylamino, substituted or unsubstituted C3-8 cycloalkyl, substituted or unsubstituted C3-8 cycloalkoxy, substituted or unsubstituted C3-8 cycloalkylamino, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted 3-to 8-membered heterocyclyl containing 1 to 2 heteroatoms selected from N and O, -COR^a、-CO₂R^a、-CONR^aR^b、-NR^aC(O)R^b、-NR^aSO₂R^b、-SR^a、-SOR^a、-SO₂R^a、-SO₂NR^aR^b、-OC(O)R^a、-OC(O)NR^aR^bThe substitution means that at least 1 site is substituted with the following substituent: halogen, cyano, amino, nitro, hydroxy, trifluoromethyl, C1-3 alkyl, C1-3 alkoxy, C1-3 alkylamino;

wherein said R^aAnd R^bEach independently hydrogen, C1-C6 alkyl, C3-C6 cycloalkyl, aryl, heteroaryl;

R₅is hydrogen, halogen, cyano, nitro, amino, hydroxyl, trifluoromethyl, substituted or unsubstituted C1-8 alkyl, substituted or unsubstituted C1-8 alkoxy, substituted or unsubstituted C1-C8 alkylamino, substituted or unsubstituted C3-8 cycloalkyl, substituted or substituted C3-C8 cycloalkoxy, or a pharmaceutically acceptable salt thereof,Substituted or unsubstituted C1-8 alkylamino, substituted or unsubstituted aryl or heteroaryl, said aryl or heteroaryl being furan, thiophene, pyrrole, oxazole, thiazole, imidazole, pyrazole, benzofuran, benzothiophene, benzoxazole, benzothiazole, phenyl, pyridine, pyridazine, pyrimidine, pyrazine, quinoline or naphthyl, said substitution being at least 1 position substituted by: halogen, cyano, amino, nitro, hydroxy, trifluoromethyl, methylthio, C1-3 alkyl, C1-3 alkoxy, C1-3 alkylamino;

R₆is hydrogen, halogen, cyano, nitro, amino, hydroxyl, trifluoro C1-C3 alkyl, C1-C3 alkoxy, C1-C3 alkylamino.

Further, R₁、R₂、R₃、R₄Each independently selected from H, halogen, cyano, nitro, amino, hydroxyl, trifluoromethyl and C_1-6Alkyl radical, C_1-6Heteroalkylalkoxy, C_1-6Alkylamino radical, C_3-8Cycloalkyl radical, C_3-8Cycloalkoxy, C_3-8Cycloalkylamino, aryl, heteroaryl, 3-to 8-membered heterocyclyl containing 1-2 heteroatoms selected from N and O;

R₅selected from H, halogen, cyano, nitro, amino, hydroxy, trifluoromethyl, C_1-8Alkyl radical, C_1-8Alkoxy radical, C_1-8Alkylamino radical, C_3-8Cycloalkyl radical, C_3-8Cycloalkoxy, C_1-8Alkylamino, aryl, heteroaryl;

R₆represents hydrogen, halogen, cyano, nitro, amino, hydroxy, trifluoromethyl, trifluoroethyl, trifluoropropyl, C_1-3A heteroalkyl group.

More preferably, R₁、R₂、R₃、R₄Each independently selected from: halo, phenyl, C_1-3Alkyl radical, C_1-3Alkoxy, a nitrogen-containing five-or six-membered heterocyclic group; r₁、R₂、R₃、R₄Any one of hydrogen may be C_1-3Alkyl radical, C_1-3Alkoxy substitution; r₅Is aryl, heteroaryl; r₅Any one of hydrogen may be substituted byAnd (3) substituent: halogen radical, C_1-3Alkyl radical, C_1-3Alkoxy, nitro, trifluoromethyl, cyano, methylsulfonyl; r₆Is hydrogen, C_1-3Alkyl or C_1-3An alkoxy group.

Further, R₅The hydrogen substitution at the para position occurs.

Wherein the aryl and heteroaryl in R5 include but are not limited to: furan, thiophene, pyrrole, oxazole, thiazole, imidazole, pyrazole, benzofuran, benzothiophene, benzoxazole, benzothiazole, phenyl, pyridine, pyridazine, pyrimidine, pyrazine, quinoline, naphthyl.

The "pharmaceutically acceptable salts" of the present invention can be synthesized from the parent compound containing an acid or base by conventional chemical methods, and such salts are generally prepared by: prepared by reacting these compounds in free acid or base form with a stoichiometric amount of the appropriate base or acid, in water or an organic solvent or a mixture of the two. "pharmaceutically acceptable salts" include, but are not limited to: inorganic acid salts such as hydrochloride, hydrobromide, nitrate, sulfate, phosphate and the like; organic acid salts such as formate, acetate, propionate, benzoate, maleate, fumaric acid, succinate, tartrate, citrate, and the like; alkyl sulfonates such as methylsulfonate, ethylsulfonate, and the like; aryl sulfonates such as benzenesulfonate, p-toluenesulfonate, and the like.

As the most preferable technical scheme, the imidazo [1,2-a ] pyridine compounds of the invention specifically comprise 24 compounds of formula I-1 to formula I-24;

the invention also provides a preparation method of the compound, which is prepared by two routes, wherein the first route (reaction process) comprises the following steps:

s1, mixing a formula (1) and a formula (2) in a solvent, and reacting to generate an intermediate (3);

s2, generating an intermediate (4) by the intermediate (3) under the catalytic action of a rhodium catalyst;

s3, hydrolyzing the intermediate (4) into acid (5) at room temperature, and then carrying out condensation reaction at room temperature to obtain a target product;

the second route (reaction process) of the preparation method of the above compound comprises the following steps:

s1, catalyzing a compound of a formula (6) by a rhodium catalyst to generate an intermediate (7);

s2, carrying out SUZUKI coupling reaction on the intermediate to generate an intermediate (8);

s3, carrying out room temperature condensation reaction on the intermediate to obtain a target product;

according to the invention, a large number of experimental researches prove that the compound can specifically inhibit STAT 3.

The invention also provides the application of the compound in preparing a medicament for preventing and/or treating in-vivo and in-vitro growth and metastasis of tumors; tumors of the present invention include, but are not limited to: acute lymphocytic leukemia, acute myelocytic leukemia, adrenocortical carcinoma, AIDS-related cancer, AIDS-related lymphoma, anal cancer, extrahepatic-biliary cancer, bladder cancer, bone cancer, brain stem glioma, brain tumor, bronchial adenoma, Burkitt's lymphoma, carcinoid tumor, unknown primary cancer, central nervous system lymphoma, cervical cancer, childhood cancer, germ cell tumor, eye cancer, stomach cancer, kidney cancer, larynx cancer, blood cancer, liver cancer, non-small cell lung cancer, melanoma, prostate tumor, rectal cancer, salivary gland carcinoma, sarcoma, small intestine cancer, soft tissue sarcoma, uterine sarcoma, testicular cancer, and breast cancer.

Experimental research shows that the compound or pharmaceutically acceptable salt and solvate thereof can obviously inhibit proliferation, migration and invasion of various in vitro tumor cells, so that the invention also provides application of the compound in preparing medicines for inhibiting proliferation, migration and invasion of tumor cells.

In addition, research also finds that the compound I-1 can promote tumor cell apoptosis, so the invention also provides application of the compound or pharmaceutically acceptable salts and solvates thereof in preparing medicaments for promoting tumor cell apoptosis.

More preferably, the tumor cell is a breast cancer cell, a lung cancer cell, a gastric adenocarcinoma cell and/or a gastric cancer cell.

The invention researches the inhibition mechanism of the compound on STAT3 through a series of experiments, and results show that the compound can obviously inhibit STAT3 dimerization and combination of STAT3 and DNA, inhibit the tyrosine phosphorylation level of STAT3, inhibit the expression of the downstream target genes BCL-XL, C-myc and Mcl-1 of STAT3 and have concentration dependence. Therefore, the invention also provides application of the compound in preparing a medicament for inhibiting the tyrosine phosphorylation level of STAT 3.

Through in vivo mouse experiments, the influence of the compound on in vivo tumors is investigated, the result is that the compound has no toxicity to the visceral organs of mice, and the compound can obviously reduce the volume and weight of in vivo tumors, namely obviously inhibit the growth and proliferation of the tumors.

The invention also provides a pharmaceutical composition comprising the compound or a pharmaceutically acceptable salt, solvate or solvate thereof, and an EGFR inhibitor in combination; when the inhibitor and the EGFR inhibitor are used together, the generation of acquired drug resistance of EGFR drugs can be delayed, the clinical service life of the EGFR drugs can be prolonged, and the EGFR inhibitor has important clinical significance.

Compared with the prior art, the invention has the following technical effects:

the compound disclosed by the invention has the characteristics of high selectivity, strong drug effect, good drug forming property, safety and the like in inhibiting STAT3 protein, has good application prospect in preparing medicines for treating diseases such as abnormal cell proliferation, morphological change, hyperkinesia and the like with high STAT3 expression and diseases related to angiogenesis or cancer metastasis, and is particularly suitable for treating and preventing tumor growth and metastasis.

Drawings

FIG. 1 photograph of six-well plate for experiment of inhibitory effect of Compound I-1 on Breast, Lung and gastric cancer cell survival; FIG. 2 bar graph of colony counts per well in the inhibition of breast, lung and gastric cancer cell survival assay by Compound I-1;

FIG. 3 is a graph showing the results of a scratch test on the inhibitory effect of Compound I-1 on breast cancer and gastric cancer cell migration;

FIG. 4 is a diagram showing the results of a Transwell experiment on the inhibition effect of compound I-1 on breast cancer cell invasion; FIGS. 4(a) and 4(b) are views of an invasion microscopic field; FIGS. 4(c) and 4(d) are histograms of the invaded cells;

FIG. 5 is a graph showing the results of an Annexin V-FITC/PI double staining method for the ability of compound I-1 to promote apoptosis in breast and gastric cancer cells; the above 3 figures are studied for breast cancer cells; the above 3 figures were studied for gastric cancer cells; i-1-1 μm represents the addition of 1 μm of drug I-1; w1010-3 μm represents 3 μm of the added drug;

FIG. 6 is a WesternBlot experimental result chart of the effect of compound I-1 on the phosphorylation level of STAT3 and the expression of downstream target genes thereof in breast cancer cells;

FIG. 7 is a graph showing the experimental results of a fluorescence confocal method of the inhibitory activity of Compound I-1 on the p-STAT3 nuclear translocation in breast and lung cancer cells;

FIG. 8 is a graph showing the experimental results of the fluorescence confocal method for inhibiting STAT3 dimerization by Compound I-1;

FIG. 9 is a graph showing the results of gel migration (EMSA) method of inhibiting the binding activity of STAT3 to DNA in breast cancer cells by Compound I-1;

FIG. 10 is a graph of the results of a dual luciferase reporter assay for the effect of Compound I-1 on the transcriptional activity of STAT 3;

FIG. 11 is a graph of the growth and proliferation of compound I-1 concentration in a mouse model breast cancer (HCC 70); FIG. 11(a) is the effect on tumor size; FIG. 11(b) is the effect on tumor weight;

FIG. 12 is a morphological diagram of the major organs of a mouse after the compound I-1 acts on the mouse model;

FIG. 13 is a graph of Compound I-1 concentration versus growth proliferation in a human tumor xenograft (PDX) mouse model; FIG. 13(a) is the effect on tumor size; FIG. 13(b) is the effect on tumor weight;

FIG. 14 is a morphological diagram of the major organs of a mouse after the compound I-1 was used as a mouse model of human tumor xenograft (PDX).

Wherein the mouse model used in figures 11 and 12 is a nude mouse; the mouse model used in FIGS. 13 and 14 was a NOD-SCID mouse, and organs were displayed twice using different models to verify their effect on organs.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the technical solutions of the present invention will be described in detail below with reference to specific examples and comparative examples. It is to be understood that the described embodiments are merely exemplary of the invention, and not restrictive of the full scope of the invention. All other embodiments, which can be derived by a person skilled in the art from the examples given herein without any inventive step, are within the scope of the present invention.

Unless otherwise specified, the devices used in the present examples, comparative examples and experimental examples were all conventional experimental devices, the materials and reagents used were commercially available without specific reference, and the experimental methods without specific reference were also conventional experimental methods.

Example 1

The structure of N- (1, 1-dioxybenzene [ b ] thiophene-6-yl) -2- (2-phenylimidazole [1,2-a ] pyridine-3-yl) acetamide (I-1) is shown as follows

Step 1: preparation of 2-phenylimidazo [1,2-a ] pyridines

78), heating and refluxing for 4 hours, detecting by TLC (thin layer chromatography), completely reacting, cooling to room temperature, spin-drying the solvent, extracting and separating by using ethyl acetate and water, combining organic phases, drying by using anhydrous sodium sulfate, spin-drying the solvent, and performing column chromatography (petroleum ether: ethyl acetate 5:1, V/V) gave 0.9 g of a tan solid in: 97 percent.

Step 2: preparation of ethyl 2- (2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

Extraction separation is carried out by dichloromethane and water, organic phases are combined, anhydrous sodium sulfate is dried, solvent is dried by spinning, and column chromatography (petroleum ether: ethyl acetate: 2:1, V/V) is carried out to obtain 505 mg of yellow-brown liquid, wherein the yield is as follows: 70 percent.¹H NMR(400MHz,CDCl₃)δ8.14(d,J＝6.9Hz,1H),7.85(d,J＝7.2Hz,2H),7.67(d,J＝9.1Hz,1H),7.49(t,J＝7.5Hz,2H),7.39(t,J＝7.4Hz,1H),7.24(t,1H),6.88(td,J＝6.8,0.9Hz,1H),4.26–4.22(q,2H),4.06(s,2H),1.29(t,3H).EI-MS:m/z(M+H⁺):280.14.

And step 3: preparation of 2- (2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

The organic phases were taken, combined, dried over anhydrous sodium sulfate and the solvent was dried by spinning to give 360 mg of a white solid in the following yield: 82 percent.¹H NMR(400MHz,CD3OD_SPE)δ8.86(d,J＝6.9Hz,1H),8.11–8.06(m,1H),8.02(d,J＝8.9Hz,1H),7.78–7.73(m,2H),7.69–7.59(m,4H),4.30(s,2H).EI-MS:m/z(M+H⁺):252.15.

And 4, step 4: preparation of (1, 1-dioxybenzo [ b ] thiophen-6-yl) -2- (2-phenylimidazo [1,2-a ] pyridin-3-yl) acetamide (I-1)

Azole (HOAt), and 156 mg of N, N-Diisopropylethylamine (DIPEA) were stirred at room temperature for 4 hours to complete the reaction, extracted with ethyl acetate and water, the organic phase was dried over anhydrous sodium sulfate, the solvent was dried by spin-drying and column chromatography (dichloromethane: methanol 20:1, V/V) gave 82 mg of a pale yellow solid in yield: 49 percent.¹H NMR(400MHz,DMSO)δ10.99(s,1H),8.46(d,J＝6.7Hz,1H),8.17(s,1H),7.84–7.76(m,3H),7.65(d,J＝9.0Hz,1H),7.59(dd,J＝11.5,7.6Hz,2H),7.50(t,J＝7.4Hz,2H),7.39(t,J＝7.2Hz,1H),7.34–7.27(m,2H),6.98(t,J＝6.7Hz,1H),4.36(s,2H)；¹³C NMR(101MHz,DMSO)δ168.6,144.5,143.5,141.8,137.7,134.8，133.2,130.5，129.1,128.3,128.0,127.0，126.1,125.5,125.1,123.8,117.1,115.1,112.4,111.9,32.3.HRMS calcd for C₂₃H₁₇N₃O₃S(M⁺):416.1063,found:416.1062.

Example 2

The structure of 2- (7-chloro-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-2) is shown below

Step 1: preparation of 7-chloro-2-phenylimidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (7-chloro-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

(500MHz,CDCl₃)δ8.05(d,J＝7.3Hz,1H),7.80(d,J＝7.4Hz,2H),7.64(d,J＝2.0Hz,1H),7.47(t,J＝7.7Hz,2H),7.38(t,J＝7.4Hz,1H),6.82(dd,J＝7.3,2.0Hz,1H),4.21(q,J＝7.1Hz,2H),4.00(s,2H),1.26(t,J＝7.2Hz,3H).EI-MS:m/z(M+H⁺):314.08

And step 3: preparation of Ethyl 2- (7-chloro-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

MeOD)δ8.82(d,J＝7.2Hz,1H),8.04(s,1H),7.72(d,J＝6.0Hz,2H),7.62(dd,J＝19.0,6.4Hz,4H),4.24(s,2H).EI-MS:m/z(M+H⁺):286.05

And 4, step 4: preparation of 2- (7-chloro-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-2)

DMSO)δ10.93(s,1H),8.52(d,J＝7.4Hz,1H),8.12(s,1H),7.81(d,J＝2.0Hz,1H),7.77–7.73(m,3H),7.58(dd,J＝16.4,7.6Hz,2H),7.50(t,J＝7.7Hz,2H),7.40(t,J＝7.3Hz,1H),7.29(d,J＝6.9Hz,1H),7.06(dd,J＝7.3,2.1Hz,1H),4.35(s,2H)；¹³C NMR(126MHz,DMSO)δ168.3,144.2(d,J＝6.9Hz),141.7,137.6,134.4,133.2,130.5,130.2,129.2,128.3,127.0,126.7,126.1,123.8,115.8(d,J＝12.9Hz),113.4,111.9,32.2.HRMS calcd for C₂₃H₁₆ClN₃O₃S(M⁺):450.0309,found:450.0670.

Example 3

The structure of 2- (7-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-3) is shown below

Step 1: preparation of 7-bromo-2-phenylimidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (7-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

7.47(t,J＝7.5Hz,2H),7.38(t,J＝7.3Hz,1H),6.96(dd,J＝7.2,1.6Hz,1H),4.22(q,J＝7.1Hz,2H),4.01(s,2H),1.27(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):385.03.

And step 3: preparation of Ethyl 2- (7-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

(d,J＝6.9Hz,1H),8.26(s,1H),7.75(d,J＝7.0Hz,3H),7.67(d,J＝6.6Hz,3H),4.27(s,2H).EI-MS:m/z(M+H⁺):330.00.

And 4, step 4: preparation of 2- (7-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-3)

7.96(s,1H),7.75(d,J＝7.2Hz,3H),7.58(dd,J＝16.7,7.5Hz,2H),7.50(t,J＝7.6Hz,2H),7.39(t,J＝7.3Hz,1H),7.29(d,J＝6.9Hz,1H),7.15(d,J＝7.3Hz,1H),4.34(s,2H)；¹³C NMR(126MHz,DMSO)δ168.3,144.6,144.1,141.8,137.6,134.4,133.2,130.5,129.1,128.3(d,J＝7.8Hz),127.0,126.7,126.0,123.8,118.9,118.0,115.8(d,J＝18.7Hz),111.9,32.2.HRMS calcd forC₂₃H₁₆BrN₃O₃S(M⁺):494.0168,found:494.0171.

Example 4

The structure of 2- (7-methyl-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-4) is shown below

Step 1: preparation of 7-methyl-2-phenylimidazo [1,2-a ] pyridine

And the preparation method was the same as in step 1 of example 1, to obtain a yellow solid.

Step 2: preparation of Ethyl 2- (7-methyl-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

(t,J＝7.7Hz,2H),7.42(s,1H),7.37(t,J＝7.4Hz,1H),6.70(dd,J＝7.0,1.5Hz,1H),4.21(q,2H),4.01(s,2H),2.42(s,3H),1.27(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):294.14.

And step 3: preparation of Ethyl 2- (7-methyl-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

J＝6.9Hz,1H),7.75(s,1H),7.73–7.68(m,2H),7.64(d,J＝7.0Hz,2H),7.44(d,J＝6.8Hz,1H),4.29(s,2H),2.65(s,3H).EI-MS:m/z(M+H⁺):266.11.

And 4, step 4: preparation of 2- (7-methyl-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-4)

8.29(d,J＝6.8Hz,1H),8.14(s,1H),7.78(t,J＝7.2Hz,3H),7.61–7.56(m,2H),7.49(t,J＝7.6Hz,2H),7.38(t,J＝7.4Hz,1H),7.29(d,J＝6.9Hz,1H),7.13(d,J＝6.6Hz,1H),6.88(t,J＝6.8Hz,1H),4.31(s,2H),2.56(s,3H)；¹³C NMR(126MHz,DMSO)δ168.6,144.8,142.9,141.8,137.6,134.9,133.2,130.5,129.0,128.4,127.9,127.0,126.5,126.0,123.7(d,J＝17.8Hz),123.2,115.5,112.5,111.9,32.4,17.0.HRMS calcd for C₂₄H₁₉N₃O₃S(M⁺):430.1220,found:430.1203.

Example 5

The structure of 2- (7-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-5) is shown below

Step 1: preparation of 7-methoxy-2-phenylimidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (7-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

10.3,4.9Hz,2H),7.37(d,J＝7.2Hz,1H),6.97(d,J＝1.7Hz,1H),6.58(dd,J＝7.4,2.4Hz,1H),4.21(q,J＝7.1,3.2Hz,2H),3.98(s,2H),3.86(s,3H),1.27(t,3H).EI-MS:m/z(M+H⁺):310.13.

And step 3: preparation of Ethyl 2- (7-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

δ8.64(s,1H),7.69(s,2H),7.64–7.56(m,3H),7.33(s,1H),7.19(s,1H),4.19(s,2H),4.08(s,3H).EI-MS:m/z(M+H⁺):282.10.

And 4, step 4: preparation of 2- (7-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-5)

10.93(s,1H),8.31(d,J＝7.4Hz,1H),8.15(s,1H),7.81–7.74(m,3H),7.58(dd,J＝16.8,7.5Hz,2H),7.48(t,J＝7.4Hz,2H),7.36(t,J＝7.2Hz,1H),7.29(d,J＝6.8Hz,1H),7.01(s,1H),6.69(d,J＝7.0Hz,1H),4.29(s,2H),3.38(s,3H)；¹³C NMR(126MHz,DMSO)δ168.8,157.8,145.9,142.8,141.8,137.6,135.1,133.2,130.5,129.0,128.0,127.7,127.0,126.2,126.0,123.7,113.8,111.9,106.9,94.7,56.0,32.2.HRMS calcd for C₂₄H₁₉N₃O₄S(M⁺):446.1169,found:446.1152.

Example 6

The structure of 2- (8-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-6) is shown below

Step 1: preparation of 8-methoxy-2-phenylimidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (8-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

7.35(t,J＝7.4Hz,1H),6.74(t,J＝7.2Hz,1H),6.49(d,J＝7.6Hz,1H),4.21(q,2H),4.01(d,J＝3.7Hz,5H),1.26(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):310.15.

And step 3: preparation of Ethyl 2- (8-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

7.70(d,J＝3.4Hz,2H),7.63(d,J＝3.7Hz,3H),7.49(s,2H),4.20(d,J＝4.5Hz,5H).EI-MS:m/z(M+H⁺):282.11.

And 4, step 4: preparation of 2- (8-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-6)

7.49(t,J＝7.2Hz,2H),7.37(t,J＝6.9Hz,1H),7.29(d,J＝6.7Hz,1H),6.88(t,J＝6.9Hz,1H),6.73(d,J＝7.3Hz,1H),4.33(s,2H),3.98(s,3H)；¹³C NMR(126MHz,DMSO)δ168.5,148.8,142.4,141.8,138.8,137.6,134.8,133.3,130.5 129.0,128.2,127.9,127.0,126.0,123.8,118.1,116.0,112.5,111.9,102.0,56.2,32.5.HRMS calcd for C₂₄H₁₉N₃O₄S(M⁺):446.1169,found:446.1154.

Example 7

The structure of 2- (6-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-7) is shown below

Step 1: preparation of 6-methoxy-2-phenylimidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (6-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

MHz,CDCl₃)δ7.83(d,J＝7.6Hz,2H),7.68(d,J＝1.9Hz,1H),7.57(d,J＝9.7Hz,1H),7.47(t,J＝7.6Hz,2H),7.37(t,J＝7.4Hz,1H),7.04(dd,J＝9.7,2.3Hz,1H),4.24(q,J＝7.1Hz,2H),4.02(s,2H),3.87(s,3H),1.30(t,J＝3.4Hz,3H).EI-MS:m/z(M+H⁺):310.10.

And step 3: preparation of Ethyl 2- (6-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

The same procedure as in step 3 of example 1 gave a white solid.¹H NMR(500MHz,CDCl₃)δ8.09(s,1H),7.70(d,J＝5.9Hz,3H),7.50(dd,J＝18.8,6.7Hz,4H),4.06(s,2H),3.92(s,3H).EI-MS:m/z(M+H⁺):282.12.

And 4, step 4: preparation of 2- (6-methoxy-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-7)

1H),8.17(d,J＝27.5Hz,2H),7.81(dd,J＝21.5,8.0Hz,3H),7.61–7.55(m,3H),7.48(t,J＝7.5Hz,2H),7.36(t,J＝7.0Hz,1H),7.28(d,J＝6.8Hz,1H),7.12(d,J＝9.7Hz,1H),4.39(s,2H),3.84(s,3H)；¹³C NMR(126MHz,DMSO)δ168.7,149.0,141.9,141.4,137.6,134.8,133.3,130.5,129.0,128.1,127.9,127.0,126.0,123.7,119.9,117.34,116.4,111.8,107.5,57.0,32.4.HRMS calcd for C₂₄H₁₉N₃O₄S(M⁺):446.1169,found:446.1164.

Example 8

The structure of 2- (2- (4-fluorophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-8) is shown below

Step 1: preparation of 2- (4-fluorophenyl) -7-methoxyimidazo [1,2-a ] pyridine

A brown solid was obtained.

Step 2: preparation of Ethyl 2- (2- (4-fluorophenyl-7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetate

¹H NMR(400MHz,CDCl₃)δ7.98(dd,J＝14.3,6.8Hz,1H),7.90–7.77(m,2H),7.17(dt,J＝17.3,8.7Hz,2H),6.99(dd,J＝11.2,9.2Hz,1H),6.61(dd,J＝10.4,3.0Hz,1H),4.24(q,2H),3.90(d,J＝30.0Hz,5H),1.30(t,J＝6.9Hz,3H).EI-MS:m/z(M+H⁺):328.12.

And step 3: preparation of Ethyl 2- (2- (4-fluorophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetic acid

And the preparation method was the same as in step 3 of example 1, to obtain a white solid.¹H NMR(500MHz,MeOD)δ8.63(d,J＝7.5Hz,1H),7.74(dd,J＝8.3,5.2Hz,2H),7.38(t,J＝8.5Hz,2H),7.30(s,1H),7.20(d,J＝7.3Hz,1H),4.23(s,2H),4.09(s,3H).EI-MS:m/z(M+H⁺):300.09.

And 4, step 4: preparation of 2- (2- (4-fluorophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-8)

δ10.95(s,1H),8.31(d,J＝7.5Hz,1H),8.14(s,1H),7.78(t,J＝6.6Hz,2H),7.58(dd,J＝17.1,7.6Hz,2H),7.30(dd,J＝15.2,7.7Hz,3H),7.00(d,J＝2.0Hz,1H),6.69(dd,J＝7.5,2.3Hz,1H),5.76(s,1H),4.26(s,2H),3.86(s,3H)；¹³C NMR(126MHz,DMSO)δ168.7,163.0,157.8,145.8,141.9(d,J＝17.7Hz),137.6,133.2,130.5,129.9(d,J＝8.2Hz),127.0,126.1(d,J＝13.3Hz),123.8,116.0,115.8,113.7,111.9,106.9,94.7,56.0,55.3,32.1.HRMS calcd forC₂₄H₁₈FN₃O₄S(M⁺):464.1075,found:464.1069.

Example 9

The structure of 2- (2- (4-trifluoromethylphenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-9) is shown below

Step 1: preparation of 2- (4-trifluoromethylphenyl) -7-methoxyimidazo [1,2-a ] pyridine

1, a brown solid was obtained.

Step 2: preparation of Ethyl 2- (2- (4-trifluoromethylphenyl-7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetate

A colored liquid.¹H NMR(500MHz,CDCl₃)δ8.04(d,J＝7.5Hz,1H),7.99(d,J＝8.1Hz,2H),7.69(d,J＝8.1Hz,2H),7.08(s,1H),6.65(dd,J＝7.5,2.4Hz,1H),4.23(q,2H),3.99(s,2H),3.86(s,3H),1.29(t,3H).EI-MS:m/z(M+H⁺):378.12.

And step 3: preparation of Ethyl 2- (2- (4-trifluoromethylphenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetic acid

The reagents and preparation were the same as in step 3 of example 1 to give a white solid.¹H NMR(400MHz,MeOD)δ8.67(s,1H),7.95(s,4H),7.36(s,1H),7.23(s,1H),4.26(s,2H),4.11(s,3H).EI-MS:m/z(M+H⁺):350.09.

And 4, step 4: preparation of 2- (2- (4-trifluoromethylphenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-9)

δ10.93(s,1H),8.34(d,J＝7.5Hz,1H),8.14(s,1H),7.98(d,J＝8.1Hz,2H),7.84(d,J＝8.2Hz,2H),7.76(s,1H),7.58(dd,J＝16.3,7.5Hz,2H),7.29(d,J＝6.9Hz,1H),7.04(d,J＝2.0Hz,1H),6.72(d,J＝7.4Hz,1H),4.32(s,2H),3.87(s,3H)；¹³C NMR(126MHz,DMSO)δ168.5,158.1,146.1,141.7,141.2,139.1,137.6,133.2,130.5,128.5,127.8,127.0,126.3,126.2–125.8,123.8,115.2,112.0,107.4,94.8,56.1,32.1.HRMS calcd for C₂₅H₁₈F₃N₃O₄S(M⁺):514.1043,found:514.1057.

Example 10

The structure of 2- (2- (4-cyanophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-10) is shown below

Step 1: preparation of 2- (4-cyanophenyl) -7-methoxyimidazo [1,2-a ] pyridine

Step 1, a white solid was obtained.

Step 2: preparation of Ethyl 2- (2- (4-cyanophenyl-7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetate

Hz,3H),7.78(d,1H),7.69(t,J＝8.8Hz,1H),7.31–7.27(m,1H),6.98–6.87(m,1H),4.25(q,2H),4.04(s,2H),1.30(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):335.13.

And step 3: preparation of Ethyl 2- (2- (4-cyanophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetic acid

The materials, reagents and preparation were the same as in step 3 of example 1 to give a white solid.¹H NMR(400MHz,MeOD)δ8.65(s,1H),7.97(d,J＝26.4Hz,4H),7.26(d,J＝34.0Hz,2H),4.15(d,J＝48.7Hz,5H).EI-MS:m/z(M+H⁺):307.10.

And 4, step 4: preparation of 2- (2- (4-cyanophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-10)

(s,1H),8.35(d,J＝7.4Hz,1H),8.13(s,1H),7.95(dd,J＝19.3,8.1Hz,4H),7.77(d,J＝7.9Hz,1H),7.58(dd,J＝16.4,7.5Hz,2H),7.29(d,J＝6.8Hz,1H),7.03(s,1H),6.73(d,J＝6.1Hz,1H),4.33(s,2H),3.87(s,3H)；¹³C NMR(126MHz,DMSO)δ167.3,152.1,143.2,141.7,141.5,138.1,137.9,135.2,133.5,129.6,128.1,127.0,125.9,125.8–125.2,123.8,118.5,111.0,107.8,93.7,57.2,31.8.HRMS calcd for C₂₅H₁₈N₄O₄S(M⁺):471.1122,found:471.1157.

Example 11

The structure of 2- (2- (4-nitrophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-11) is shown below

Step 1: preparation of 2- (4-nitrophenyl) -7-methoxyimidazo [1,2-a ] pyridine

Step 1, a white solid was obtained.

Step 2: preparation of Ethyl 2- (2- (4-nitrophenyl-7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetate

(dd,J＝11.7,8.1Hz,3H),6.94(d,J＝2.1Hz,1H),6.65(dd,J＝7.5,2.3Hz,1H),4.25(q,J＝7.1Hz,2H),4.01(s,2H),3.90(s,3H),1.30(t,3H).EI-MS:m/z(M+H⁺):355.12.

And step 3: preparation of Ethyl 2- (2- (4-nitrophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetic acid

The reagents and preparation were the same as in step 3 of example 1 to give a white solid.¹H NMR(500MHz,MeOD)δ8.66(d,J＝6.3Hz,1H),8.47(d,J＝6.7Hz,2H),7.99(d,J＝6.5Hz,2H),7.34(s,1H),7.22(d,J＝5.5Hz,1H),4.26(s,2H),4.10(s,3H).EI-MS:m/z(M+H⁺):327.09.

And 4, step 4: preparation of 2- (2- (4-nitrophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-11)

1H),8.35(dd,J＝14.0,8.3Hz,3H),8.14(s,1H),8.07(d,J＝8.9Hz,2H),7.79(dd,J＝8.2,1.9Hz,1H),7.58(dd,J＝11.3,7.6Hz,2H),7.28(d,J＝6.9Hz,1H),7.04(d,J＝2.4Hz,1H),6.74(dd,J＝7.5,2.5Hz,1H),4.37(s,2H),3.88(s,3H)；¹³C NMR(126MHz,DMSO)δ168.3,158.3,146.6,146.3,141.8,140.5,137.6,133.2,130.5,128.6,127.0,126.4,126.1,124.3,123.9,116.3,112.0,107.7,94.8,56.1,32.2.HRMS calcd for C₂₄H₁₈N₄O₆S(M⁺):491.1020,found:491.1017.

Example 12

The structure of 2- (2- (4-methylthiophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-12) is shown below

Step 1: preparation of 2- (4-methylthiophenyl) -7-methoxyimidazo [1,2-a ] pyridine

Step 1, a white solid was obtained.

Step 2: preparation of Ethyl 2- (2- (4-methylthiophenyl-7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetate

A liquid.¹H NMR(500MHz,CDCl₃)δ8.04(dd,J＝15.9,8.6Hz,5H),6.98(d,J＝1.6Hz,1H),6.65(dd,J＝7.5,2.3Hz,1H),4.24(q,J＝7.1Hz,2H),3.99(s,2H),3.89(s,3H),3.10(s,3H),1.29(t,J＝7.0Hz,3H).EI-MS:m/z(M+H⁺):388.11.

And step 3: preparation of Ethyl 2- (2- (4-methylthiophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) acetic acid

The formulation and preparation were the same as in step 3 of example 1 to give a white solid.¹H NMR(500MHz,CDCl₃)δ8.11(d,J＝7.5Hz,1H),7.90(q,J＝8.5Hz,4H),6.80(d,J＝2.3Hz,1H),6.63(dd,J＝7.5,2.4Hz,1H),3.84–3.76(m,5H),3.02(s,3H).EI-MS:m/z(M+H⁺):360.08.

And 4, step 4: preparation of 2- (2- (4-methylthiophenyl) -7-methoxyimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-13)

The procedure was as in step 4 of example 1 to give a white solid.¹H NMR(500MHz,DMSO)δ11.00(s,1H),8.36(d,J＝7.6Hz,1H),8.14(s,1H),8.09–7.97(m,4H),7.81–7.76(m,1H),7.58(dd,J＝14.8,7.5Hz,2H),7.29(d,J＝6.7Hz,1H),7.04(d,J＝2.3Hz,1H),6.73(dd,J＝7.5,1.3Hz,1H),4.34(s,2H),3.87(s,3H),3.25(s,3H)；¹³C NMR(126MHz,DMSO)δ165.6,157.3,147.2,146.3,142.7,141.6,136.6,133.2,132.1,127.5,127.0,126.9,125.7,123.7,122.9,116.3,112.8,108.9,95.7,57.2,48.1，32.2.HRMS calcd for C₂₅H₂₁N₃O₆S₂(M⁺):524.0945,found:524.0971.

Example 13

The structure of 2- (2- (7-methoxy-2- (thiophene-2-yl) imidazole [1,2-a ] pyridine-3-yl) -N- (1, 1-dioxybenzene [ b ] thiophene-6-yl) acetamide (I-13) is shown as follows

Step 1: preparation of 7-methoxy-2- (thien-2-yl) imidazo [1,2-a ] pyridine

White solid.

Step 2: preparation of Ethyl 2- (7-methoxy-2- (thiophen-2-yl) imidazo [1,2-a ] pyridin-3-yl) acetate

(d,J＝39.3Hz,1H),7.16–7.08(m,1H),6.94(d,J＝7.5Hz,1H),6.58(d,J＝7.5Hz,1H),4.21(d,J＝23.6Hz,2H),4.06(s,2H),3.86(s,3H),1.27(d,J＝7.2Hz,3H).EI-MS:m/z(M+H⁺):316.09.

And step 3: preparation of Ethyl 2- (7-methoxy-2- (thiophen-2-yl) imidazo [1,2-a ] pyridin-3-yl) acetic acid

1H),7.79(s,1H),7.65(s,1H),7.31(s,2H),7.18(s,1H),4.33(s,2H),4.09(s,3H).EI-MS:m/z(M+H⁺):288.06.

And 4, step 4: preparation of 2- (7-methoxy-2- (thiophen-2-yl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-13)

10.95(s,1H),8.33(d,J＝7.4Hz,1H),8.11(s,1H),7.78(d,J＝8.1Hz,1H),7.56(dd,J＝14.2,4.7Hz,3H),7.40(s,1H),7.27(d,J＝5.9Hz,1H),7.15(s,1H),6.99(s,1H),6.67(d,J＝7.4Hz,1H),4.36(s,2H),3.86(s,3H)；¹³C NMR(126MHz,DMSO)δ168.4,158.0,145.8,141.8,138.4,137.6,137.4,133.2,130.5,128.4,127.0,126.5–125.9,124.0,123.7,113.1,111.8,106.9,94.5,56.1,31.9.HRMS calcd for C₂₂H₁₇N₃O₄S₂(M⁺):452.0790,found:452.0781.

Example 14

The structure of 2- (2- (7-methoxy-2- (naphthyl-2-yl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-14) is shown below

Step 1: preparation of 7-methoxy-2- (naphthyl-2-yl) imidazo [1,2-a ] pyridine

To a white solid.

Step 2: preparation of Ethyl 2- (7-methoxy-2- (naphthyl-2-yl) imidazo [1,2-a ] pyridin-3-yl) acetate

And step 3: preparation of Ethyl 2- (7-methoxy-2- (naphthyl-2-yl) imidazo [1,2-a ] pyridin-3-yl) acetic acid

And the preparation method is the same as the step 3 in the example 1, and a white solid is obtained.

And 4, step 4: preparation of 2- (7-methoxy-2- (naphthyl-2-yl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-14)

δ11.01(s,1H),8.36(d,J＝7.5Hz,1H),8.24(s,1H),8.17(s,1H),8.01(d,J＝8.6Hz,1H),7.99–7.91(m,3H),7.81(dd,J＝8.2,1.7Hz,1H),7.59(dd,J＝12.6,7.6Hz,2H),7.56–7.50(m,2H),7.29(d,J＝6.9Hz,1H),7.04(d,J＝2.4Hz,1H),6.71(dd,J＝7.5,2.4Hz,1H),4.41(d,J＝25.1Hz,2H),3.88(s,3H)；¹³C NMR(126MHz,DMSO)δ168.8,157.9,146.0,142.6,141.9,137.7,133.4(d,J＝24.6Hz),132.6,130.5,128.5(d,J＝15.5Hz),128.0,127.1,126.8,126.4(d,J＝7.7Hz),126.2,126.1,123.8,114.4,111.9,106.9,94.7,56.1,32.3.HRMS calcd for C₂₈H₂₁N₃O₄S(M⁺):496.1314,found:496.1312.

Example 15

The structure of 2- (6-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-15) is shown below

Step 1: preparation of 6-bromo-2-phenylimidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (6-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

7.51(t,J＝7.6Hz,2H),7.42(t,J＝7.4Hz,1H),7.31(dd,J＝9.5,1.8Hz,1H),4.28(q,2H),4.05(s,2H),1.33(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):358.03.

And step 3: preparation of Ethyl 2- (6-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

Step 3 in example 1 gave a white solid.¹H NMR(500MHz,CDCl₃)δ9.08(s,1H),7.95(dd,J＝48.2,41.2Hz,3H),7.61(d,J＝43.9Hz,4H),4.18(d,J＝16.4Hz,2H).EI-MS:m/z(M+H⁺):330.00.

And 4, step 4: preparation of 2- (6-bromo-2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-15)

8.86(s,1H),8.13(s,1H),7.79–7.73(m,3H),7.64–7.56(m,3H),7.50(t,J＝7.6Hz,2H),7.45–7.37(m,2H),7.29(d,J＝6.9Hz,1H),4.37(s,2H)；¹³C NMR(126MHz,DMSO)δ168.4,144.1,143.0,141.8,137.7,134.3,133.2,130.5,129.2,128.3,128.0,127.1,126.0,125.8,123.7,118.1,116.1,111.9,106.4,32.2.HRMS calcd for C₂₃H₁₆BrN₃O₃S(M⁺):494.0168,found:494.0178.

Example 16

The structure of 2- (2- (4-methyl) -6-methylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-16) is shown below

10.92(s,1H),8.26(s,1H),8.14(s,1H),7.78(dd,J＝8.2,1.9Hz,1H),7.63(d,J＝8.1Hz,2H),7.61–7.56(m,2H),7.52(d,J＝9.1Hz,1H),7.28(d,J＝7.0Hz,3H),7.16(dd,J＝9.2,1.5Hz,1H),4.28(s,2H),2.34(d,J＝8.6Hz,6H)；¹³C NMR(126MHz,DMSO)δ168.7,143.4(d,J＝15.1Hz),141.9,137.6,137.1,133.3,132.2,130.5,129.6,128.1,127.8,127.1,126.0,123.7,122.8,121.4,116.4,114.4,111.8,32.3,21.2,18.2.HRMS calcd for C₂₅H₂₁N₃O₃S(M⁺):444.1376,found:444.1381.

Example 17

The structure of 2- (2- (4-cyanophenyl) -6-methylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-11) is shown below

Step 1: preparation of 2- (4-cyanophenyl) -6-methylimidazol [1,2-a ] pyridine

A white solid was obtained.

Step 2: preparation of Ethyl 2- (2- (4-cyanophenyl-6-methylimidazo [1,2-a ] pyridin-3-yl) acetate

¹H NMR(400MHz,CDCl₃)δ8.02–7.97(m,2H),7.93(s,1H),7.79–7.71(m,2H),7.59(d,J＝9.2Hz,1H),7.14(dd,J＝9.2,1.6Hz,1H),4.26(q,2H),4.01(s,2H),2.39(d,J＝0.6Hz,3H),1.31(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):319.13.

And step 3: preparation of Ethyl 2- (2- (4-cyanophenyl) -6-methylimidazo [1,2-a ] pyridin-3-yl) acetic acid

And the preparation method was the same as in step 3 of example 1, to obtain a white solid.¹H NMR(500MHz,MeOD)δ8.68(s,1H),8.05–7.89(m,6H),4.30(s,2H),2.55(s,3H).EI-MS:m/z(M+H⁺):291.10.

And 4, step 4: preparation of 2- (2- (4-cyanophenyl) -6-methylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-17)

1H),8.32(s,1H),8.15(s,1H),8.00–7.92(m,4H),7.79(d,J＝8.2Hz,1H),7.59(dd,J＝11.7,7.8Hz,3H),7.29(d,J＝6.9Hz,1H),7.22(d,J＝9.1Hz,1H),4.35(s,2H),2.33(s,3H)；¹³C NMR(126MHz,DMSO)δ168.2,143.8,141.2,141.3,139.6,137.6,133.3,133.0,130.5,128.7(d,J＝12.9Hz),127.0,126.1,123.8,122.9,122.2,119.4,116.7,116.4,112.0,110.1,32.2,18.2.HRMS calcd for C₂₅H₁₈N₄O₃S(M⁺):455.1172,found:455.1163.

Example 18

The structure of 2- (2- (4-chloro-phenyl) -6-methylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-18) is shown below

Step 1: preparation of 2- (4-chloro-phenyl) -6-methylimidazol [1,2-a ] pyridine

Step 1, a white solid was obtained.

Step 2: preparation of Ethyl 2- (2- (4-chloro-phenyl-6-methylimidazo [1,2-a ] pyridin-3-yl) acetate

¹H NMR(500MHz,CDCl₃)δ7.90(s,1H),7.82–7.75(m,2H),7.60–7.55(m,1H),7.47–7.39(m,2H),7.09(dd,J＝8.4,4.5Hz,1H),4.22(q,2H),3.98(s,2H),2.38(s,3H),1.28(t,J＝7.0Hz,3H).EI-MS:m/z(M+H⁺):328.10.

And step 3: preparation of Ethyl 2- (2- (4-chloro-phenyl) -6-methylimidazo [1,2-a ] pyridin-3-yl) acetic acid

MeOD)δ8.65(s,1H),7.92(s,2H),7.71(s,2H),7.66(s,2H),4.28(d,J＝16.9Hz,5H).EI-MS:m/z(M+H⁺):300.07.

And 4, step 4: preparation of 2- (2- (4-chloro-phenyl) -6-methylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-18)

1H),8.29(s,1H),8.15(s,1H),7.79–7.75(m,3H),7.61–7.53(m,5H),7.29(d,J＝6.9Hz,1H),7.20(d,J＝9.2Hz,1H),4.29(s,2H),2.33(s,3H)；¹³C NMR(101MHz,DMSO)δ168.4,143.4,141.8,137.6,133.6,133.2,132.7,130.5,129.8,129.1,128.5,127.0,126.1,123.8,122.9,122.0,116.4,115.2,112.0,32.2,18.2.HRMS calcd for C₂₄H₁₈ClN₃O₃S(M⁺):464.0831,found:464.0811.

Example 19

The structure of 2- (2- (4-methylphenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzene [ b ] thiophen-6-yl) acetamide (I-19) is shown below

Step 1: preparation of 2- (4-methylphenyl) imidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (2- (4-methylphenylimidazo [1,2-a ] pyridin-3-yl) acetate

MHz,CDCl₃)δ8.12(d,J＝6.6Hz,1H),7.73(d,J＝7.1Hz,2H),7.68(d,J＝9.0Hz,1H),7.29(d,J＝7.4Hz,2H),7.23(t,J＝7.8Hz,1H),6.86(t,J＝6.6Hz,1H),4.22(q,2H),4.03(s,2H),2.41(s,3H),1.27(t,J＝7.0Hz,3H).EI-MS:m/z(M+H⁺):294.14.

And step 3: preparation of Ethyl 2- (2- (4-methylphenyl) imidazo [1,2-a ] pyridin-3-yl) acetic acid

Procedure in example 1Step 3, a white solid was obtained.¹H NMR(500MHz,MeOD)δ8.83(s,1H),8.02(d,J＝16.8Hz,2H),7.60(d,J＝16.6Hz,3H),7.45(d,J＝6.7Hz,2H),4.27(s,2H),2.45(s,3H).EI-MS:m/z(M+H⁺):266.11.

And 4, step 4: preparation of 2- (2- (4-methylphenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-19)

8.67(d,J＝3.4Hz,1H),8.47(dd,J＝10.3,3.1Hz,2H),8.15(s,1H),7.81(dd,J＝8.2,1.6Hz,1H),7.67(d,J＝8.0Hz,2H),7.59(dd,J＝8.1,4.0Hz,2H),7.46(d,J＝4.0Hz,1H),7.30–7.28(m,2H),6.96(t,J＝6.5Hz,1H),4.34(s,2H),2.35(s,3H)；¹³C NMR(126MHz,DMSO)δ167.8,143.4(d,J＝15.1Hz),142.1,137.6,136.2,134.3,132.7,130.4,129.6,128.9,127.8,126.9,126.5,124.6,123.8,121.4,115.5,114.7,112.8,33.3,21.2.HRMS calcd for C₂₄H₁₉N₃O₃S(M⁺):430.1220,found:430.1204.

Example 20

The structure of 2- (2- (4-methylthiophenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-20) is shown below

Step 1: preparation of 2- (4-methylthiophenyl) imidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (2- (4-methylthiophenylimidazo [1,2-a ] pyridin-3-yl) acetate

(500MHz,CDCl₃)δ8.19(d,J＝6.9Hz,1H),8.10(d,J＝8.3Hz,2H),8.05(d,J＝8.4Hz,2H),7.70(d,J＝9.1Hz,1H),7.31–7.27(m,1H),6.94(t,J＝6.8Hz,1H),4.25(q,2H),4.06(s,2H),3.11(s,3H),1.31(t,J＝4.2Hz,3H).EI-MS:m/z(M+H⁺):358.10.

And step 3: preparation of Ethyl 2- (2- (4-methylthiophenyl) imidazo [1,2-a ] pyridin-3-yl) acetic acid

The procedure was as in step 3 of example 1 to give a white solid.

And 4, step 4: preparation of 2- (2- (4-methylthiophenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-20)

8.50(s,1H),8.14(s,1H),8.05(q,J＝8.7Hz,4H),7.78(dd,J＝8.2,1.9Hz,1H),7.67(d,J＝9.1Hz,1H),7.59(dd,J＝11.6,7.6Hz,2H),7.38–7.33(m,1H),7.29(d,J＝6.9Hz,1H),7.02(t,J＝6.8Hz,1H),4.40(s,2H),3.26(s,3H)；¹³C NMR(126MHz,DMSO)δ162.7,143.7(d,J＝15.1Hz),142.1,136.7,136.2,133.9,132.6,130.9,129.6,129.3,127.6,126.9,126.2,124.6,122.8,122.4,116.7,114.8,113.7,48.1，33.9.HRMS calcd for C₂₄H₁₉N₃O₅S₂(M⁺):494.0839,found:494.0807.

Example 21

The structure of 2- (2- (4-nitrophenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzo [ b ] thiophen-6-yl) acetamide (I-21) is shown below

Step 1: preparation of 2- (4-nitrophenyl) imidazo [1,2-a ] pyridine

Step 2: preparation of Ethyl 2- (2- (4-nitrophenylimidazo [1,2-a ] pyridin-3-yl) acetate

–8.04(m,2H),7.70(d,J＝9.1Hz,1H),7.31(dd,J＝8.5,7.5Hz,1H),6.95(t,J＝6.8Hz,1H),4.26(q,J＝7.1Hz,2H),4.07(s,2H),1.31(t,3H).EI-MS:m/z(M+H⁺):325.11.

And step 3: preparation of Ethyl 2- (2- (4-nitrophenyl) imidazo [1,2-a ] pyridin-3-yl) acetic acid

And 4, step 4: preparation of 2- (2- (4-nitrophenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-21)

J＝6.9Hz,1H),8.35(d,J＝8.9Hz,2H),8.14(s,1H),8.09(d,J＝8.9Hz,2H),7.80–7.76(m,1H),7.68(d,J＝9.1Hz,1H),7.59(dd,J＝11.5,7.6Hz,2H),7.37(s,1H),7.29(d,J＝6.9Hz,1H),7.03(t,J＝6.4Hz,1H),4.42(s,2H)；¹³C NMR(126MHz,DMSO)δ166.8,142.7(d,J＝15.1Hz),142.1,137.4,136.6,134.2,132.6,131.8,129.5,129.3,127.9,127.7,126.4,124.8,123.8,122.4,115.7,115.2,113.8,33.1.HRMS calcd for C₂₃H₁₆N₄O₅S(M⁺):460.0812,found:460.0815.

Example 22

The structure of 2- ((2, 7-diphenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxy-benzo [ b ] thiophen-6-yl) acetamide (I-22) is shown as follows

Step 1: preparation of Ethyl 2- (2, 7-diphenylimidazo [1,2-a ] pyridin-3-yl) acetate

Under the protection of nitrogen, 110, heating and refluxing are carried out overnight. TLC detection, reaction completed, extraction with ethyl acetate and brine, combination of organics, drying over anhydrous sodium sulfate, spin drying of organic phase followed by column chromatography (petroleum ether: ethyl acetate ═ 2:1, V/V) to afford 300 mg of red solid, yield: 75 percent.¹H NMR(400MHz,CDCl₃)δ8.20(d,J＝7.2Hz,1H),7.89(d,J＝8.6Hz,3H),7.73–7.66(m,2H),7.51(td,J＝7.5,3.9Hz,4H),7.42(t,J＝7.4Hz,2H),7.18(dd,J＝7.2,1.8Hz,1H),4.26(q,J＝7.1Hz,2H),4.08(s,2H),1.31(s,3H).EI-MS:m/z(M+H⁺):356.15.

Step 2: preparation of Ethyl 2- (2, 7-diphenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

1H),8.15(s,1H),7.91(s,3H),7.77(s,2H),7.66(d,J＝6.8Hz,3H),7.58(dd,J＝11.7,6.6Hz,3H),4.29(s,2H).EI-MS:m/z(M+H⁺):328.12.

And step 3: preparation of 2- ((2, 7-Diphenyl) imidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-22)

J＝7.2Hz,1H),8.30(d,J＝4.6Hz,1H),8.16(s,1H),7.96(s,1H),7.87–7.81(m,3H),7.59(q,J＝6.7Hz,2H),7.52(q,J＝8.0Hz,2H),7.41–7.38(m,2H),7.29(d,J＝6.8Hz,1H),7.18(d,J＝8.2Hz,1H),6.88(d,1H),6.69(d,J＝8.2Hz,1H),6.10(s,1H),4.40(s,2H)；¹³C NMR(101MHz,DMSO)δ168.6,144.9,144.2,141.8,138.3,137.7,136.6,134.8,133.2,130.5,129.5,129.1,128.7,128.2(d,J＝19.8Hz),127.0(d,J＝5.1Hz),126.1,125.7,123.8,115.5,113.3,111.8(d,J＝24.7Hz),32.3.HRMS calcd for C₂₉H₂₁N₃O₃S(M⁺):492.1356,found:492.1360.

Example 23

The structure of 2- (7- (4-methoxyphenyl) -2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzene [ b ] thiophen-6-yl) acetamide (I-23) is shown below

Step 1: preparation of Ethyl 2- (7- (4-methoxyphenyl) -2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

(t,J＝7.3Hz,1H),7.16(dd,J＝7.2,1.8Hz,1H),7.04(d,J＝8.8Hz,2H),4.26(q,J＝7.1Hz,2H),4.08(s,2H),3.88(s,3H),1.32(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):386.16.

Step 2: preparation of Ethyl 2- (7- (4-methoxyphenyl) -2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

MeOD)δ8.82(s,1H),8.09(s,1H),7.91(d,J＝8.2Hz,4H),7.77(d,J＝6.4Hz,2H),7.70–7.66(m,2H),7.15(d,J＝8.2Hz,2H),4.29(s,2H),3.90(s,3H).EI-MS:m/z(M+H⁺):358.13.

And step 3: preparation of 2- (7- (4-methoxyphenyl) -2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-23)

δ10.98(s,1H),8.49(d,J＝7.2Hz,1H),8.15(s,1H),7.88(d,J＝1.0Hz,1H),7.83(s,1H),7.82–7.75(m,4H),7.63–7.56(m,2H),7.50(t,J＝7.6Hz,2H),7.39(t,J＝7.4Hz,1H),7.34(dd,J＝7.3,1.8Hz,1H),7.29(d,J＝6.9Hz,1H),7.08(d,J＝8.8Hz,2H),4.37(s,2H),3.83(s,3H)；¹³CNMR(126MHz,DMSO)δ168.6,159.9,145.1,144.0,141.8,137.7,136.4,134.8,133.3,130.6,129.1,128.2,128.0,127.1,126.0,125.5,123.8,114.9(d,J＝10.6Hz),112.1,111.9,111.5,55.7,32.3.HRMS calcd for C₃₀H₂₃N₃O₄S(M⁺):522.1482,found:522.1475.

Example 24

The structure of 2- (7- (1-methyl-1H-pyrazol-4-yl) -2-phenylimidazole [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxybenzene [ b ] thiophen-6-yl) acetamide (I-24) is shown as follows

Step 1: preparation of Ethyl 2- (7- (1-methyl-1H-pyrazol-4-yl) -2-phenylimidazo [1,2-a ] pyridin-3-yl) acetate

(t,J＝7.6Hz,2H),7.39(d,J＝7.3Hz,1H),6.96(dd,J＝7.1,1.7Hz,1H),4.22(q,J＝7.1Hz,2H),4.01(s,2H),3.93(s,3H),1.28(t,J＝7.1Hz,3H).EI-MS:m/z(M+H⁺):360.16.

Step 2: preparation of Ethyl 2- (7- (1-methyl-1H-pyrazol-4-yl) -2-phenylimidazo [1,2-a ] pyridin-3-yl) acetic acid

MeOD)δ8.75(s,1H),8.44(s,1H),8.18(s,1H),8.04(s,1H),7.82(s,1H),7.73(d,J＝6.3Hz,2H),7.66(d,J＝6.0Hz,3H),4.31(s,2H),4.01(s,3H).EI-MS:m/z(M+H⁺):332.13.

And step 3: preparation of 2- (7- (1-methyl-1H-pyrazol-4-yl) -2-phenylimidazo [1,2-a ] pyridin-3-yl) -N- (1, 1-dioxobenzo [ b ] thiophen-6-yl) acetamide (I-24)

DMSO)δ10.94(s,1H),8.44(d,J＝7.1Hz,1H),8.34(s,1H),8.14(s,1H),8.07(s,1H),7.83–7.76(m,4H),7.59(dd,J＝11.2,7.6Hz,2H),7.50(t,J＝7.6Hz,2H),7.39(d,J＝7.3Hz,1H),7.30–7.22(m,2H),4.34(s,2H),3.90(s,3H)；¹³C NMR(101MHz,DMSO)δ168.6,145.0,143.5,141.8,137.7,137.0,134.8,133.9,130.5,129.8,129.1,128.2,128.0,127.0,126.0,125.6,123.8,120.7,114.9,111.9,111.1,110.2,32.2,19.0.HRMS calcd for C₂₇H₂₁N5O₃S(M⁺):496.1438,found:496.1426.

Biological activity test part:

the compounds of the general formula inhibit proliferation, survival, migration and invasion of breast and gastric cancer cells and promote apoptosis thereof

1. Compounds for inhibiting proliferation of breast cancer, lung cancer, gastric adenocarcinoma cell and gastric cancer cell

The cell activity inhibition effect of the compound is tested by adopting a cell activity measuring method, and the experimental method is as follows: (1) cells in logarithmic growth phase were taken, according to experimental wells: drug + cells + media + CCK8, positive control wells: drug solvent (SH4-54) + cells + medium + CCK8, blank well: medium + CCK8, absorbance at 450nm was measured using a microplate reader. Typical OD values are between 0.5 and 1.5, typically between 0.8 and 1.5. The cell activity was calculated according to the following formula: cell viability (%) - (experimental well OD value-blank well OD value)/(negative control well OD value-blank well OD value) × 100%. Wherein, Table 1 shows the concentration of the drug (IC) at which the cell viability was half inhibited₅₀)。

TABLE 1

As can be seen from the results in Table 1, compounds I-1 to I-24 were able to inhibit the proliferation of lung cancer cells and gastric cancer cells in vitro; here, the STAT inhibitor SH4-54 is taken as a positive exclusive reference, and the effects of the compounds I-10, I-12, I-20 and I-21 on inhibiting the in vitro proliferation of the lung cancer cells and the gastric cancer cells are equivalent to the effect of the inhibitor SH4-54, and the effects of other compounds on inhibiting the in vitro proliferation of the lung cancer cells and the gastric cancer cells are basically better than the effect of the inhibitor SH 4-54.

In addition, the compounds I-1, I-3, I-4, I-5 and I-9 can also obviously inhibit the in vitro proliferation of breast cancer cells and human gastric adenocarcinoma cells; although the compound I-8 has no obvious inhibitory effect on breast cancer cells, the compound I-8 can obviously inhibit the in vitro proliferation of gastric adenocarcinoma cells.

2. The inhibition of breast, lung and gastric cancer cell survival by compounds of the general formula is illustrated below by way of example of compound I-1: the inhibition effect of the compound I-1 on breast cancer, lung cancer and gastric cancer cell survival is tested by adopting a cell plate cloning experiment method, and the specific operation is as follows: (1) plate paving: taking MDA-MB-231, MDA-MB-468, HCC70, A549, MGC-803 and AGS cells in logarithmic growth phase, abandoning the culture medium, washing with PBS, digesting with pancreatin, terminating digestion of the culture medium, suspending the cells into single cell suspension by the culture medium after centrifugation, counting the cell suspension by a counting plate, preparing into 500 single cell suspensions of 250 cells/ml, inoculating the cells into a six-well plate according to 2 ml/well, placing the six-well plate at 37 ℃ and 5% CO₂The incubator is used for 24 h. (2) Adding medicine: after 24h, different concentrations of I-1 were added, and a Blank (no treatment, Blank), control (DMSO, C) and experimental wells (drug addition) were set, with 3 duplicate wells for each drug and concentration. Culturing in 5% CO2 incubator at 37 deg.C for 10-15 days. (3) Fixing and dyeing: after 10-15 days, cell clone colonies can be seen by naked eyes, the culture medium is discarded, PBS is washed once, 4% paraformaldehyde is added, about 1 ml/hole is fixed for 15min, then paraformaldehyde is discarded, PBS is washed for 2 times, crystal violet with the concentration of about 500 ul/hole is added, light-shielding dyeing is carried out for 30min, then crystal violet is discarded, PBS is washed until the background is clean, and the cell clone colonies are dried at room temperature. (4) The number of colonies per well was photographed and counted, and statistically analyzed using GraphPad Prism software.

As shown in FIGS. 1 and 2, when the concentration of compound I-1 was 0.03. mu.m, 0.1. mu.m, 0.3. mu.m, and 1. mu.m, respectively, there were fewer and fewer colonies of lung cancer cells, gastric adenocarcinoma cells, and gastric cancer cells, indicating that the inhibition of the survival of lung cancer cells, gastric adenocarcinoma cells, and gastric cancer cells by compound I-1 was more and more significant and concentration-dependent. In addition, when the concentration of the compound I-1 is respectively 0.1 μm, 0.3 μm and 1 μm, the clone colonies of the three breast cancer line cells (MDA-MB-231, MDA-MB-468 and HCC70) are less and less, which shows that the survival of the three breast cancer line cells (MDA-MB-231, MDA-MB-468 and HCC70) is inhibited more and more obviously by the compound I-1 and is in concentration dependence.

3. Compound I-1 inhibits cell migration in breast and gastric cancers

The inhibition of breast cancer and gastric cancer cell migration by compound I-1 was tested using the cell scratch assay as follows: (1) plate paving: taking MDA-MB-231, MDA-MB-468, HCC70, MGC-803 and AGS cells in logarithmic growth phase (cell number is about 80-90%), discarding the culture medium, washing with PBS, digesting with pancreatin, terminating digestion of the culture medium, re-suspending with the culture medium to obtain single cell suspension after centrifugation, counting with a counting plate, preparing into 50-100 ten thousand cell suspensions, inoculating the cells into a six-well plate according to 2 ml/well, placing at 37 deg.C and 5% CO₂The incubator is used for 24 h. (2) Scratching and adding medicine: after 24h, when the cell density is more than 90%, scratching the cell with a tip compared with a ruler, then removing the culture medium, washing the cell with PBS for 2 times, adding a fresh culture medium, taking a picture under a microscope, adding the compound I-1 with different concentrations, and setting a control group (adding a drug solvent and marking as a group 0) and an experimental well (adding the compound I-1 with different concentrations). Placing at 37 ℃ and 5% CO₂Culturing in an incubator for 24-96 h. (3) Photographing and counting: after 24-96h, the medium was discarded, washed 2 times with PBS, and photographed under a microscope.

The test result is shown in FIG. 3, FIG. 3(a) shows the effect of compound I-1 on breast cancer cells, wherein FIG. 3(a) shows the effect of compound I-1 on MDA-MB-468, MDA-MB-231 and HCC70, respectively, and after 96h, the result shows that the higher and higher concentration of compound I-1 has more obvious inhibition effect on the migration of MDA-MB-468, MDA-MB-231 and HCC 70; FIG. 3(b) shows the effect of compound I-1 on MGC-803 and AGS, respectively, and after 24h, the results show that the higher the concentration of compound I-1, the more significant the inhibition effect on MGC-803 and AGS migration.

4. Compound I-1 inhibits breast cancer cell invasion

The inhibitory effect of the compound I-1 on breast cancer cell invasion is tested by adopting a Transwell experiment method, which comprises the following steps: (1) transwell chamber pre-incubation: taking out the used nest to a new 24-well plate, adding blank cultureSoaking in medium (without serum or double antibody), and standing in 37 deg.C incubator for 1 hr. (2) Plate paving: collecting MDA-MB-231 and HCC70 cells in logarithmic growth phase, discarding culture medium, washing with PBS, digesting with pancreatin, terminating digestion of culture medium, re-suspending with 2% FBS culture medium after centrifugation to obtain single cell suspension, counting with counting plate, preparing 10-30 ten thousand/ml cell suspension, inoculating cells into upper layer of transwell chamber at 300 μ L/hole, adding 2% FBS culture medium at 500 μ L/hole into lower layer, standing at room temperature for 30min, placing at 37 deg.C, and placing at 5% CO₂The incubator is used for 24 h. (3) Adding medicine: after 24h, the upper and lower layers of the chamber were aspirated, 500. mu.L of 20% FBS medium was added to the lower layer, and 300. mu.L of 2% FBS medium containing Compound I-1 at various concentrations was added to the upper layer, and the control group and the experimental group were set. Culturing in a 5% CO2 incubator at 37 deg.C for 12 h. (4) Fixing and dyeing: after 12h, the upper and lower layers of culture medium are discarded, PBS is used for washing once, the upper layer cells of the chamber are scraped by a cotton swab, the upper layer is washed by PBS for 2-3 times, residual cells are washed, then 4% paraformaldehyde is used for fixing for 15min, PBS is used for washing for 2 times, and crystal violet is added for shading and dyeing for 30 min. The residual crystal violet was washed off with PBS and the chamber was air dried at room temperature. (5) Photographing and counting: after drying, 5 fields of view were randomly selected under the microscope for photographing and counted with Image J.

As shown in FIG. 4, it can be seen from FIG. 4(a), FIG. 4(b) and FIG. 4(c) that the inhibitory effect on MDA-MB-231 and HCC70 in breast cancer cell lines becomes more and more significant as the concentration of compound I-1 becomes higher, i.e., compound I-1 can inhibit the invasion of breast cancer cells, and the inhibition becomes stronger with the increase of the concentration, and has statistical difference.

5. Compound I-1 promotes apoptosis in breast and gastric cancer cells

The ability of the compound I-1 to promote apoptosis of breast cancer and gastric cancer cells is detected by adopting Annexin V-FITC/PI double staining method, and the experimental method is as follows: (1) plate paving: collecting MDA-MB-231 and MGC-803 cells in logarithmic growth phase, discarding culture medium, washing with PBS once, digesting with pancreatin, terminating digestion of culture medium, centrifuging, re-suspending with culture medium to obtain single cell suspension, counting, preparing into 50-80 ten thousand/ml cell suspension, inoculating cells into six-well plate at 2 ml/well, standing at 37 deg.C and 5% CO₂The incubator is used for 24 h. (2) Adding medicine: adding different concentrations of compound I-1 into the cells,and treating for 48 h. (3) And (3) detection: the detection is carried out by adopting a Bebo Annexin V-FITC apoptosis detection kit (BB-4101-1). The medium was discarded, washed with PBS, digested with pancreatin without EDTA, collected and centrifuged, washed 2 times with cold PBS and the PBS was aspirated off. Suspending cells with 400 μ L of 1 × Annexin V binding solution, adding 5 μ L of Annexin V-FITC staining solution into the suspension, gently mixing, incubating on 2 ice in the dark for 15min, adding 10 μ L of PI staining solution, gently mixing, and incubating on ice in the dark for 5 min. Immediately detected with a flow cytometer. And then processed with FlowJo 7.6 software.

As shown in FIG. 5, it is understood from FIG. 5 that Compound I-1 promotes apoptosis, especially early apoptosis, in breast cancer and gastric cancer cells, and has concentration dependence.

The following example of compound I-1 illustrates the mechanism of the compounds of the general formula of the present invention for inhibiting the growth of cancer cells

1. The compound I-1 inhibits the phosphorylation level of cancer cell STAT3 and the expression of downstream target genes thereof, and the experimental method is as follows: (1) protein sample extraction: taking MDA-MB-231, MDA-MB-468 and HCC70 cells which have been treated by different concentrations of compound I-1 for corresponding time, placing on ice, removing the culture medium, washing with precooled PBS once, removing PBS, adding RIPA lysate (containing protease and phosphatase inhibitor), shaking and cracking on ice for 15min, scraping the cells with cell scraper, placing the cell culture plate obliquely for 5min, transferring the cell lysate to a 1.5mL centrifuge tube, vortexing for 20s, standing on ice for 5min, 15000rpm, and centrifuging at 4 ℃ for 15 min. The supernatant was pipetted into a new 1.5mL centrifuge tube and quantified by BCA. After the concentration is determined, the protein sample is diluted into a protein sample with the same total amount of protein, 5x Loading buffer is added, denaturation is carried out for 5min at 100 ℃, and temporary storage is carried out at-80 ℃. (2) Western Blot: adding the protein samples into the gel according to the equal protein amount of each hole, performing electrophoresis, and rotating the membrane on ice after the electrophoresis is finished. After the membrane transfer was completed, the membrane was soaked in 5% BSA and placed on a shaker for 1h at room temperature. The membrane was then cut and transferred to pY705-STAT3, pS727-STAT3, STAT3, β -actin, BCL-XL, C-myc and Mcl-1 primary antibodies for overnight incubation at 4 ℃. After the primary antibody incubation was completed, the cells were washed 3 times with TBST for 10 min/time. The membrane was then transferred to a secondary murine or rabbit antibody and incubated for 1h at room temperature. After incubation with secondary antibody, wash with TBST 3 times, 10 min/time. The developer (solution a: solution B ═ 1:1) was prepared in situ, and developed using a chemiluminescence developing apparatus.

Respectively detecting 705 tyrosine phosphorylation level, 727 serine phosphorylation level and expression of STAT3 downstream target genes BCL-XL, C-myc and Mcl-1 of STAT3 in different cancer cells by a protein immunoblotting method; meanwhile, the results are shown in FIG. 6 by taking beta-actin as an internal reference, and as can be seen from FIG. 6, the compound I-1 can remarkably inhibit 705 tyrosine phosphorylation level of STAT3 in a short time (3h), has concentration dependence, but does not influence 727 serine phosphorylation level. Meanwhile, the expression of the downstream target genes BCL-XL, C-myc and Mcl-1 of STAT3 can be inhibited, and the concentration dependence is realized.

2. The compound I-1 inhibits the nuclear translocation of cancer cell p-STAT3, and the experimental method is as follows: (1) plate paving: collecting MDA-MB-231 and MGC-803 cells in logarithmic growth phase, discarding culture medium, washing with PBS once, digesting with pancreatin, terminating digestion of culture medium, centrifuging, re-suspending with culture medium to obtain single cell suspension, suspending appropriate amount of cells, dripping into confocal dish, standing for 30min, transferring to 37 deg.C, and adding 5% CO₂And (5) culturing for 24 hours in an incubator. (2) Adding medicine: after 24h, different concentrations of W1078 were added to the confocal dish, treated for 12-18h, and stimulated for 30min with 100ng/mL IL-6. (3) Fixing: after 6h, the medium was discarded, washed once with PBS, and fixed for 15min with 4% paraformaldehyde. (4) Membrane permeation: discarding paraformaldehyde, washing with PBS (shaking table) for 3 times (5 min/time), adding 0.3% Triton-X100, and allowing to permeate membrane for 10min, and washing with PBS (shaking table) for 3 times (5 min/time). (5) And (3) sealing: adding goat serum, placing in a shaking table, and sealing at room temperature for 1 h. (6) Incubating primary antibody: discarding goat serum, adding pY705-STAT3 primary antibody diluted with goat serum, placing in a wet box, and standing overnight at 4 deg.C. (7) Hatching a secondary antibody: the primary antibody was recovered and washed 3 times with PBS on a shaker for 5 min/time. Then, a fluorescent secondary antibody diluted by goat serum is added, and the mixture is incubated for 1h at room temperature in a dark place. (8) Dyeing the core: discard the secondary antibody, wash 3 times for 5 min/time in PBS shaker. DAPI was added and incubated for 10min in the dark. DAPI was then discarded and washed 3 times 5 min/time in PBS shaker. (9) And (3) photographing: the photographs were taken with a laser scanning ultra high resolution microscope (FV 3000).

As shown in FIG. 7, it is clear from FIG. 7 that Compound I-1 significantly inhibited the nuclear translocation of pY705-STAT3 in MDA-MB-231 and A549 cells, and was concentration-dependent.

3. Inhibition of STAT3 dimerization by Compound I-1

The inhibition activity of the compound I-1 on STAT3 dimerization is measured by adopting a fluorescence confocal method, and the experimental method is as follows: (1) plate paving: taking a logarithmic growth phase (HEK-293T cell, abandoning a culture medium, washing with PBS once, trypsinizing, stopping digestion of the culture medium, suspending the cell into a single cell suspension by using the culture medium after centrifugation, suspending a proper amount of cells into a confocal dish uniformly, standing for 30min, smoothly transferring the cell into a 5% CO2 incubator at 37 ℃, culturing for 24h, (2) transfecting for 24h, transfecting HA-STAT3 and Flag-STAT3 plasmids (3) into the HEK-293T cell, adding the compounds I-1 with different concentrations into the HEK-293T cell after 24h, treating for 24h, adding 100 ng/mL/IL-6, treating for 1h, (4) abandoning the culture medium, washing with PBS once, adding 4% paraformaldehyde, fixing for 15min, (5) removing paraformaldehyde, placing the HEK-293T into a shaking table, washing for 3 times, 5 min/time, adding 0.3% Triton-X100 for 10min, then, the mixture was washed 3 times for 5 min/time in PBS shaker. (6) And (3) sealing: adding goat serum, placing in a shaking table, and sealing at room temperature for 1 h. (7) Incubating primary antibody: discarding goat serum, adding HA-tag and Flag-tag primary antibody diluted with goat serum, placing in a wet box, and standing overnight at 4 deg.C. (8) Hatching a secondary antibody: the primary antibody was recovered and washed 3 times with PBS on a shaker for 5 min/time. Then, a fluorescent secondary antibody diluted by goat serum is added, and the mixture is incubated for 1h at room temperature in a dark place. (9) Dyeing the core: discard the secondary antibody, wash 3 times for 5 min/time in PBS shaker. DAPI was added and incubated for 10min in the dark. DAPI was then discarded and washed 3 times 5 min/time in PBS shaker. (10) And (3) photographing: the photographs were taken with a laser scanning ultra high resolution microscope (FV 3000).

The results are shown in FIG. 8, where the yellow moiety is significantly reduced, i.e., co-localization of HA-STAT3 with Flag-STAT3 is reduced, indicating that compound I-1 inhibits STAT3 dimer formation, i.e., compound I-1 inhibits STAT3 dimerization.

4. Compound I-1 inhibits cancer cell STAT3 binding to DNA

The inhibition activity of compound I-1 on the binding of STAT3 to DNA in breast cancer cells was determined by the gel migration (EMSA) method as follows: (1) extracting nucleoprotein: the nucleoprotein is extracted by adopting a Biyunshi nuclear protein and cytoplasm protein extraction kit (P0027). Utensil for cleaning buttockThe body is as follows: MDA-MB-231 cells that had been treated for 3h with different concentrations of Compound I-1 were taken, placed on ice, medium was discarded, washed once with pre-cooled PBS, PBS was discarded, cells were scraped with a cell scraper, and cells were pipetted off. Cells were collected by centrifugation and the supernatant was aspirated with maximum effort, leaving a pellet of cells. 200. mu.L of the cell plasma protein extraction reagent A containing 1% PMSF was added to each 20. mu.L of the cell pellet. The Vortex was vigorous at maximum speed for 5 seconds to completely suspend and disperse the cell pellet. Ice-bath for 10-15 min. 10 μ L of cytoplasmic protein extraction reagent B was added. Vortex 5 seconds at maximum speed and ice bath 1 minute. The highest speed was Vortex 5 seconds vigorously, centrifuged at 12,000 and 16,000g for 5min at 4 ℃. The supernatant was completely aspirated, and 50. mu.L of a nuclear protein extraction reagent containing 1% PMSF was added. The highest speed, vigorous Vortex 15-30 seconds, completely suspended and dispersed the cell pellet. Then put back into ice bath, and then put into high speed violent Vortex 15-30 seconds every 1-2min for 30 min. Centrifugation at 12,000-16,000g for 10min at 4 ℃. Immediately sucking the supernatant into a precooled plastic tube to obtain the extracted cell nucleus protein. BCA quantification was performed and buffered at-80 ℃. (2) EMSA: prepared with EMSA glue (TBE buffer (5x) 1mL, ddH2O,5mL, acylamide/bisacrylamide (30%, w/v) 1mL, glycerin 250 μ L, 10% ammonium persulfate, 100 μ L, TEMED,10 μ L); EMSA binding reaction (negative control: nuclear-Free Water + EMSA/Gel-Shift binding buffer (5X) + STAT3 or STAT5 probe; sample reaction: nuclear-Free Water + EMSA/Gel-Shift binding buffer (5X) +8 μ g nuclear protein + STAT3 or STAT5 probe; adding various reagents in this order, mixing well before adding STAT3 or STAT5 probe, standing at room temperature for 10min, adding probe, mixing well, standing at room temperature for 20 min.); electrophoretic analysis (0.5 XTBE as electrophoretic fluid, 100V on ice, pre-electrophoresis for 30 min. then sample of mixed loading buffer is added into loading well, 10. mu.L diluted 1 Xloading buffer (blue) is added into redundant loading well for observing electrophoresis progress, 100V on ice, 60-70 min. shearing nylon membrane with size consistent with glue size, soaking with 0.5XTBE for 10min, on ice, 380mA, 70 min. taking out membrane, UV crosslinking for 15min, then soaking membrane in confining liquid (dissolving at 37 ℃), sealing at room temperature for 1h, taking 50. mu.L Streptavidin-HRPConjute, adding into 15ml confining liquid (1:300 diluted solution)Release), mixing well for standby. After the blocking was completed, the blocking solution used for nylon membrane blocking was removed, and 15ml of the blocking solution containing Streptavidin-HRP Conjugate prepared in the previous step was added. Slowly shake on a horizontal shaker for 15 minutes. The blocking solution was then discarded and the eluent (37 ℃ solution, ddH) was added₂O is diluted to 1 x), and washed for 4 times and 10 min/time. Preparing chemiluminescence working solution (A: B ═ 1:1) on site, adding the working solution on the surface of the nylon membrane to enable the working solution to completely cover the nylon membrane, carrying out development at room temperature for 2min, and carrying out development by using a chemiluminescence developing instrument.

The result is shown in fig. 9, the result of the gel migration (EMSA) experiment of selective inhibition of binding of compound I-1 adenocarcinoma STAT3 and DNA shows that the drug inhibits the STAT3DNA binding activity, and meanwhile, the STAT5DNA binding activity is not greatly affected by the drug through STAT5DNA binding observation, so that certain selectivity of the STAT5DNA binding activity is proved.

Therefore, compound I-1 can inhibit STAT3 binding to DNA in MDA-MB-231 cells, but does not affect STAT5 binding to DNA, i.e., compound I-1 has a specific inhibitory effect on STAT 3.

5. Inhibition of STAT3 transcriptional activity by Compound I-1

The effect of compound I-1 on the transcriptional activity of STAT3 was determined using a dual luciferase reporter as follows: (1) plate paving: taking HEK-293T cells in logarithmic growth phase, removing the culture medium, washing with PBS, digesting with pancreatin, stopping digestion of the culture medium, suspending into single cell suspension with the culture medium after centrifugation, and inoculating into a 96-well plate at 20000/well. (2) Transfection: after 24h, 50ng pGL3-STAT3+50ng STAT3C +40ng Renilla luciferase reporter plasmid TK-RL were transfected per well with lipo 2000. (3) Adding medicine: after 24h of transfection, different concentrations of Compound I-1 were added and treated for 24 h. (4) And (3) detection: the detection is carried out by using a Biyunshi dual-luciferase reporter gene detection kit (RG 028). Discard the culture medium, add 50. mu.L of reporter gene cell lysate to each well, shake for 5 min. And (3) melting the firefly luciferase detection reagent and the renilla luciferase detection buffer solution, and reaching the room temperature. Renilla luciferase assay substrate (100X) was placed on an ice bath or ice box for use. According to the amount of 100 mu L of each sample, a proper amount of renilla luciferase detection buffer solution is taken, and a renilla luciferase detection substrate (100X) is added according to the ratio of 1:100 to prepare renilla luciferase detection working solution. After completion of shaking, 50. mu.L of firefly luciferase assay reagent was added to each well, and RLU1(relative light unit) was measured after homogenizing with a gun. Reporter cell lysates were used as blank controls. After completion of the above procedure for measuring firefly luciferase, 100. mu.L of Renilla luciferase assay working solution was added and homogenized with a pipette, and then RLU2(relative light unit) was measured. The ratio of RLU1/RLU2 was used to compare the differences in STAT3 transcriptional activity between different samples.

As a result, as shown in FIG. 10, Compound I-1 inhibited the transcriptional activity of STAT3 and was concentration-dependent.

6. Compound I-1 inhibits growth and proliferation of tumor in animal model

a. Drug effect, pharmacology and toxicology research in tumor-bearing mouse model

Establishing a tumor-bearing mouse model: the log phase cancer cells were centrifuged, washed 3 times with sterile PBS and counted to adjust the cell concentration to about 2 x10 ^7 cells/ml, and then 100. mu.L of cell suspension was injected subcutaneously on the ventral side of the mice. Grouping experiments: after the tumor-bearing mouse model was established (about 1-3 weeks), the mice were randomly divided into control groups and administration groups, each of which had about 6-10 mice. And (3) medicine intervention: after the model was established, drug intervention was initiated. The control group was intraperitoneally injected with 15% castor oil-containing PBS (drug solvent group), and the administration group was intraperitoneally injected with drugs in an administration volume of 100. mu.L/mouse. Mice body weight and tumor volume were measured daily for 3-4 weeks, and mice were monitored for behavioral status. Collecting samples: mice were sacrificed by cervical dislocation 3-4 weeks after administration, tumor bodies were removed, weighed and volume measured, and blood, heart, spleen, liver and other organs and tissues were taken for further pharmacological and toxicological studies.

b. Study of drug efficacy and pharmacology in a human tumor xenograft (PDX) mouse model.

Collecting tumor specimens of patients: the specimen can be derived from tissue biopsy and tumor radical operation specimen, collected after tumor is isolated, fresh tumor tissue is completely soaked in 0 ℃ serum-free culture medium without double antibody. Tumor tissue was cut into 2 x 2mm tissue pieces with sterile tissue scissors and washed three times with culture medium. The skin on both sides of the abdomen and the back of the mouse in an anesthesia state is respectively provided with a small opening with the diameter of about 3mm, a small pocket-shaped space is separated, a tumor tissue block is planted under the skin, and the wound is sutured. 100 Xdouble-resistant solution is dropped on the wound to prevent infection. Each tumor was implanted in 5 mice (F1), and the implanted tumors were observed at least once a week. Observations included the presence or absence of tumor growth and measurement of tumor volume. After about 12-16 weeks, the transplanted tumor began to grow to a size of 1-2cm 3. F1 mice were removed from their ventral tumor mass, and the tumor mass was cut into 2 x 2mm tissue pieces using sterile tissue scissors, and washed by soaking in serum-free and double-resistant RPMI1640 medium. The tissue blocks were implanted subcutaneously on both ventral and dorsal sides of mice, and 5 mice were implanted per tumor (F2). After the tumor was loaded, the transplanted tumor grew to 1-2cm3 size, F2 mice were removed from the ventral tumor mass, cut into 2X 2mm tissue pieces, and the tissue pieces were then implanted subcutaneously on both ventral and dorsal sides of the mice, and several mice were implanted per tumor (F3). When the transplanted tumor grows to about 100mm3 size, F3 was randomly divided into a control group and an administration group, after the division, the control group was intraperitoneally injected with PBS (drug solvent) containing 15% castor oil, and the administration group was intraperitoneally injected with a drug in a volume of about 100. mu.L/mouse, during which the tumor volume and body weight were measured daily. After three to four weeks of continuous administration, mice were sacrificed by dislocation of cervical vertebrae, tumor bodies were taken out, weighed and weighed, and blood, heart, spleen, liver and other organs and tissues were taken for further pharmacological and toxicological studies.

The results are shown in FIGS. 11-14, and it can be seen from FIG. 11 that Compound I-1 can significantly reduce the volume and weight of tumors (breast cancer) in mice, indicating that Compound I-1 can inhibit the growth and proliferation of tumors in mice; FIG. 12 shows that compound I-1 has substantially no effect on the mouse major organ morphology and other indicators; FIG. 13 shows that I-1 significantly inhibits the growth and proliferation of breast cancer in a human tumor xenograft (PDX) mouse model (and the inhibition effect is more significant at higher concentrations), and FIG. 14 shows that I-1 has little effect on the indicators such as the morphology of the major organs of animals.

It should be finally noted that the above examples are only intended to illustrate the technical solutions of the present invention, and not to limit the scope of the present invention, and that other variations and modifications based on the above description and thought may be made by those skilled in the art, and that all embodiments need not be exhaustive. Any modification, equivalent replacement, and improvement made within the spirit and principle of the present invention should be included in the protection scope of the claims of the present invention.

Claims (5)

1. An imidazo [1,2-a ] pyridine compound, characterized in that the compound has the following structural formula:

2. a STAT3 specific inhibitor, wherein the STAT3 specific inhibitor is an imidazo [1,2-a ] pyridine compound according to claim 1, or a pharmaceutically acceptable salt thereof.

3. The use of a compound according to claim 1 for the preparation of a medicament for the prophylactic and/or therapeutic inhibition of tumors, wherein said tumors are one or more of lung, gastric or breast cancer tumors.

4. The use according to claim 3, wherein the medicament is in the form of an injection, tablet, pill, capsule, suspension or emulsion.

5. A pharmaceutical composition comprising a compound of any one of claim 1 and an EGFR inhibitor.

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