CN112321568B - 4-methylpyrrole substituted indolone derivative, preparation method and medical application thereof - Google Patents

Abstract

The invention discloses a 4-methylpyrrole substituted indolone derivative, a preparation method and medical application thereof. Relates to the field of pharmaceutical chemistry; compared with the compound 10d discovered in the early stage, the 4-methylpyrrole substituted indolone derivative has better chemical stability, higher selectivity on FLT3-ITD, more ideal pharmacokinetic property and stronger in-vivo anti-tumor activity, particularly has lower onset dosage for treating acute myeloid leukemia, has a tumor inhibition rate obviously superior to that of the compound 10d and sunitinib under the same dosage, and is difficult to generate drug resistance.

Description

4-methylpyrrole substituted indolone derivative, preparation method and medical application thereof

Technical Field

The invention relates to the field of medicinal chemistry, in particular to 4-methylpyrrole substituted indolone derivatives or pharmaceutically acceptable salts thereof, a medicinal composition containing the compounds, a preparation method of the compounds and application of the compounds in the aspect of tumor resistance.

Background

Malignant tumors (cancers) are serious chronic diseases threatening human health, and have become one of the most serious public health problems in China or even in the world in the 21 st century. The anticancer drugs used in clinic at present mainly comprise cytotoxic drugs, but the traditional chemotherapy drugs have the defects of easy drug resistance, strong toxic and side effects, more adverse reactions, poor curative effect and the like. With the development of molecular biology and tumor pharmacology, the mechanism of tumorigenesis is further elucidated, and the research on antitumor drugs is progressing from traditional cytotoxic drugs to novel molecular-targeted antitumor drugs.

Tyrosine kinases exist in most pathogenic genes in a plurality of tumor diseases, so that research and development of medicaments taking tyrosine kinases as targets (such as EGFR, FGFR, PDGFR, VEGFR, IGFR, CSF1R, c-Kit, FLT3, RET and the like) are widely concerned, and Tyrosine Kinase Inhibitors (TKI) become hot spots in the field of research of antitumor medicaments. Moreover, the tyrosine kinase inhibitor has the advantages of high selectivity, low toxicity, less side effect and the like, so that the tyrosine kinase inhibitor is superior to the traditional antitumor drugs. According to statistics, more than 80% of currently marketed kinase drugs are tyrosine kinase inhibitors, and even part of the currently marketed kinase drugs become the first choice drugs for treating tumors.

FMS-like receptor tyrosine kinase 3 (Fms-like tyrosine kinase 3, FLT3) is an important receptor tyrosine kinase in cell signaling, and abnormal activation thereof is closely related to the occurrence and development of various tumors, particularly Acute Myeloid Leukemia (AML). Studies have shown that FLT3 is highly expressed in more than 70% of AML patients and Acute Lymphoblastic Leukemia (ALL) patients, with more than 1/3 of AML patients being accompanied by FLT3 gene mutations. Therefore, targeted therapy against FLT3 has become one of the important means of treating AML, and large amounts of FLT3 inhibitors have entered clinical studies or are undergoing preclinical studies.

Sunitinib (Sunitinib) is a multi-target tyrosine kinase inhibitor developed by the company pfeiri, is approved by the FDA to be on the market in 2006, is mainly used for the treatment of gastrointestinal stromal tumors and renal cell carcinoma, and is the first anticancer drug approved for two types of indications at the same time. It can effectively inhibit receptor tyrosine kinases such as VEGFR-1, VEGFR-2, VEGFR-3, PDGFR-beta, c-Kit, FLT3 and the like. By inhibiting the expression of various angiogenesis promoting factors in cancer cells, the aim of inhibiting the angiogenesis and 'starvation' of cancer cells is fulfilled. Although the Sunitinib has obvious antitumor effect, the Sunitinib still shows the side effects of hypodynamia, bone marrow suppression, fever and the like in clinic, has strong tissue accumulation and can not be taken continuously, and the scheme of stopping taking for 2 weeks after continuously taking for 4 weeks is adopted in clinic. It is worth noting that Sunitinib has a direct inhibitory effect on tumor cells with FLT3 mutation, but shows strong toxic and side effects in clinical trials for treating AML, and has no therapeutic window. In earlier research, the structure of Sunitinib is modified by adopting a biological electronic isostere strategy, and a derivative 10d [ Eur.J.Med.chem.2017,127,72-86 ] with a larger treatment window than Sunitinib is found; CN 104829596B. However, in the subsequent studies, it was found that compound 10d has low chemical stability and generates impurities after a long time at room temperature, resulting in a decrease in purity; the purity of compound 10d was maintained at low temperature (-20 ℃) and vacuum drying, while the purity decreased to 80.8% after 150 days at room temperature. In addition, the in vivo efficacy of compound 10d in a nude mouse transplantation tumor model with acute myelogenous leukemia (MV 4-11) is still a certain gap compared with Sunitinib. Therefore, we need to further elucidate the reason why the compound 10d is chemically unstable, modify the structure of the unstable part, improve the chemical stability, and find out a candidate compound with better drug property and stronger activity.

The LC-MS detection shows that the new impurities are mainly derivatives with the molecular weight reduced by 2 (12.42%) and the molecular weight reduced by 4 (6.00%). Through the separation and identification of impurities and the guess of reaction mechanism, we guess the mechanism of impurity generation and generally guess ¹ H-NMR、 ¹³ And performing structural analysis on C-NMR, HMQC, HMBC and ROSEY to determine that the two main impurities are 10d-X and 10d-Y respectively. The above results provide the basis for further improvement of chemical stability of the compounds. Since the methyl group at the 2-position of the pyrrole ring in the structure of the compound 10d is easily oxidized, the 2-position of the pyrrole ring needs to be structurally modified, and the methyl group is removed or replaced by a functional group which is not easily oxidized, such as Cl, br, H and the like, and finally, the chemical stability can be improved and the activity can be maintained after the methyl group is removed. The invention is characterized by a series of structural optimization and activityScreening and finding out 4-methylpyrrole substituted indolone anti-tumor candidate compounds with high chemical stability and stronger activity.

Disclosure of Invention

In order to solve the problems, the invention provides a 4-methylpyrrole substituted indolone derivative shown in a general formula (I), and a tautomer, an enantiomer, a diastereomer, a racemate or pharmaceutically acceptable salt thereof.

The technical scheme of the invention is as follows: a4-methylpyrrole substituted indolone derivative or a pharmaceutically acceptable salt, isomer, prodrug, polymorph or solvate thereof, wherein the chemical structural formula of the derivative is shown as formula (I):

wherein:

R ¹ selected from hydrogen atom, halogen, alkyl, alkoxy, halogenated alkyl, amino and NHCOR ³ Or NHSO ₂ R ³ One or more of the above;

R ² selected from alkyl, heterocycloalkyl, alkyl-substituted heterocycloalkyl, alkenyl, aryl, heteroaryl, CH ₂ R ⁴ 、(CH ₂ ) _n NR ⁵ R ⁶ And NHR ⁷ R ⁸ One or more of the above;

R ³ selected from alkyl or alkenyl;

R ⁴ selected from heterocycloalkyl or alkyl-substituted heterocycloalkyl;

R ⁵ and R ⁶ Are respectively selected from one or more of hydrogen, alkyl, cycloalkyl and heterocycloalkyl;

at the same time, R ⁵ And R ⁶ Forming a 4-7 membered heterocycloalkyl group, wherein the 4-7 membered heterocycle contains one or more N, O, S atoms and the 4-7 membered heterocycle is substituted with one or more alkyl groups;

R ⁷ and R ⁸ Are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroarylOne or more of the groups;

at the same time, R ⁷ And R ⁸ Forming a 4-7 membered heterocycloalkyl group, wherein the 4-7 membered heterocycle contains one or more N, O, S atoms and the 4-7 membered heterocycle is substituted with one or more alkyl groups;

n is 1 to 6.

Typical compounds preferred in the present invention are as follows, but are not limited to:

another object of the present invention is to provide a method for preparing a 4-methylpyrrole-substituted indolone derivative, comprising the steps of:

the first step is as follows: compound II in POCl ₃ And DMF, performing Vilsmeier reaction to generate a compound III; in particular, under low temperature (0 ℃ C.), POCl ₃ Dropwise adding the mixture into anhydrous DMF, stirring at room temperature for 0.5 hour, adding compound II dissolved in the anhydrous DMF, and heating to 80 ℃ for reaction for 0.5 hour; after the reaction is finished, quenching the reaction by using an ice-water bath, adjusting the pH value to about 7, extracting by using dichloromethane, washing by using brine, and drying to obtain a compound III;

the second step is that: carrying out aldol condensation reaction on the compounds III and IV under the action of alkali to generate a compound IV; specifically, dissolving the compounds III and IV in ethanol, adding pyrrolidine, and heating to 50 ℃ for reaction for 2 hours; after the reaction is finished, performing suction filtration, washing a filter cake by ethanol, and drying to obtain a compound V;

the third step: performing nitro reduction reaction on the compound V under the action of zinc powder to obtain a compound VI; specifically, dissolving a compound in a mixed solvent of tetrahydrofuran and methanol, adding a saturated solution of ammonium chloride, heating to 50 ℃, and then adding zinc powder to react for 0.5 hour; after the reaction is finished, distilling under reduced pressure to remove the solvent, dissolving the residue with ethyl acetate, washing with saturated sodium carbonate solution and saline solution in sequence, and drying; distilling under reduced pressure to remove the solvent, pulping with methanol to obtain purified compound VI;

the fourth step: reacting the compound IV with a corresponding carboxylic acid compound under the action of a condensing agent; or firstly reacting with acyl chloride and then reacting with the corresponding amine compound to obtain a target compound I; specifically, dissolving the compound V and a corresponding carboxylic acid compound in DMF, adding condensing agents PyBOP and DIPEA, and reacting at room temperature for 12 hours; after the reaction is finished, adding the reaction solution into water, extracting by ethyl acetate, washing by using salt water, drying, and purifying by column chromatography; finally, removing the Boc protecting group through HCl ethyl acetate solution treatment to obtain a target compound I;

or dissolving the compound IV in tetrahydrofuran, adding DIPEA, cooling to 0 ℃, adding corresponding acyl chloride, reacting at room temperature for 0.5 hour, then distilling under reduced pressure to remove the solvent, dissolving the residue with a mixed solvent of ethyl acetate and methanol, pulping and purifying to obtain an acylation product; further dissolving the product in anhydrous DMF, adding corresponding amine compounds, heating to 50 ℃ and reacting for 12 hours; after the reaction is finished, pouring the reaction liquid into water, extracting by ethyl acetate, washing by using brine, drying, distilling under reduced pressure to remove the solvent, and pulping and purifying the residue by using ethyl acetate to obtain the target compound I.

Further, the isomers are selected from one or more of enantiomers, diastereomers, geometric isomers or stereoisomers.

Further, the 4-methyl pyrrole substituted indolone derivative is selected from the following group:

(S, Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-2-carboxamide hydrochloride;

(R, Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-2-carboxamide hydrochloride;

(R, Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-3-carboxamide hydrochloride;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) piperidine-4-carboxamide hydrochloride;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) piperidine-2-carboxamide hydrochloride;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (pyrrolidin-1-yl) acetamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (pyrrolidin-2-yl) acetamide hydrochloride;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (piperidin-1-yl) acetamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (4-methylpiperazin-1-yl) acetamide;

(Z) -2- (diethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) acetamide;

(Z) -2- (cyclopentylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) acetamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propionamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (piperidin-1-yl) propionamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (4-methylpiperazin-1-yl) propanamide;

(Z) -3- (diethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide;

(Z) -3- (dimethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide;

(Z) -3- (ethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propionamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (propylamino) propionamide;

(Z) -3- (tert-butylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propionamide;

(Z) -3- (cyclopentylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propionamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3-morpholinepropionamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -6- (trifluoromethyl) nicotinamide;

(Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -4-methylpiperazine-1-carboxamide;

(Z) -N- (5- ((5-chloro-2-oxindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propanamide

(Z) -N- (5- ((5-bromo-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propionamide;

(Z) -N- (5- ((5-methoxy-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propanamide.

The invention aims to provide a pharmaceutical composition, which comprises the 4-methylpyrrole substituted indolone derivative, pharmaceutically acceptable salt, isomer, prodrug, polymorph or solvate thereof, and at least one pharmaceutically acceptable carrier, additive, auxiliary agent or excipient.

The invention also aims to provide the application of the 4-methylpyrrole substituted indolone derivative or the pharmaceutically acceptable salt and the pharmaceutical composition thereof in preparing tyrosine kinase inhibitors. The compound or the pharmaceutically acceptable salt thereof and the pharmaceutical composition thereof have obvious inhibitory activity on receptor tyrosine kinases such as VEGFR-1, VEGFR-2, PDGFR alpha, PDGFR-beta, FLT3, CSF-1R and the like, and particularly have strong inhibitory activity on FLT3, so the compound or the pharmaceutically acceptable salt thereof can be used for preparing medicines for treating related tumor diseases caused by abnormal expression of the tyrosine kinases.

The invention also aims to provide the 4-methylpyrrole substituted indolone derivative or pharmaceutically acceptable salt and application of the pharmaceutical composition thereof in preparing antitumor drugs. In vivo and in vitro antitumor tests show that the compound can obviously inhibit the growth of malignant tumors such as liver cancer, breast cancer, colon cancer and the like. Therefore, the compound or the pharmaceutically acceptable salt thereof and the pharmaceutical composition thereof can be used as a single therapeutic agent or used in combination with other antitumor drugs for treating various malignant tumors caused by abnormal expression of tyrosine kinase.

The invention also aims to provide the application of the 4-methylpyrrole substituted indolone derivative or the pharmaceutically acceptable salt and the pharmaceutical composition thereof in preparing the drugs for treating acute myelogenous leukemia. In vitro anti-tumor activity experiments show that the compound can obviously inhibit the proliferation of acute myelogenous leukemia cells. The whole animal experiment shows that the compound has good curative effect on acute myelogenous leukemia, and the effective dose is lower than that of Sunitinib and the compound 10d. Therefore, the compound or the pharmaceutically acceptable salt thereof and the pharmaceutical composition thereof can be used as a single therapeutic agent or combined with other anti-tumor medicaments for treating acute myelogenous leukemia.

The beneficial effects of the invention are: compared with the compound 10d discovered in the early stage, the 4-methylpyrrole substituted indolone derivative disclosed by the invention has better chemical stability, higher selectivity on FLT3-ITD, more ideal pharmacokinetic characteristics and stronger in-vivo anti-tumor activity, particularly lower onset dosage for treating acute myelogenous leukemia, obviously better tumor inhibition rate than the compound 10d and sunitinib under the same dosage, difficulty in generating drug resistance and good industrial prospect.

Detailed Description

In order to more clearly illustrate the technical solution of the present invention, the following further detailed description is made on the technical solution of the present invention:

example 1

Synthesis of (S, Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-2-carboxamide hydrochloride (A01)

And (3) synthesis of an intermediate III: at 0 deg.C, adding POCl ₃ (1 mL) was slowly added dropwise to anhydrous DMF (1.1 mL) and after stirring at room temperature for 0.5h, a solution of Compound II (0.80g, 6.35mmol) in DMF (2 mL) was added to the reaction mixture, which was then heated to 80 ℃ for 0.5h. After the reaction was monitored by LC-MS, the heating was stopped, cooled to room temperature, cooled in an ice bath, and saturated Na was added ₂ CO ₃ The reaction solution was adjusted to pH7, then extracted with methylene chloride (20 mL. Times.2), and several layers were combined, washed successively with water and a saturated sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction, concentrated, and purified by column chromatography (petroleum ether: ethyl acetate =20:1 to 4). ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm): 13.03(1H,brs),9.79(1H,s),8.17(1H,s),2.59(3H,s).

Synthesis of intermediate V-1: compound III (1.2g, 7.79mmol) and IV-1 (1.3g, 8.56mmol) were dissolved in ethanol (10 mL), and pyrrolidine (0.83mL, 10.13mmol) was added and heated to 50 ℃ for 2 hours. After the LC-MS monitoring reaction is finished, stopping heating, cooling to room temperature, performing suction filtration, washing a filter cake with a small amount of ethanol, and performing vacuum drying to obtain orange yellow solid 2.09g with the yield of 85%. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):14.26(1H,brs),11.17(1H,s),8.29 (1H,d,J＝3.6Hz),7.90(1H,dd,J ₁ ＝9.2Hz,J ₂ ＝2.5Hz),7.85(1H,s),7.03(1H,td,J ₁ ＝9.5Hz, J ₂ ＝2.5Hz),6.88(1H,dd,J ₁ ＝8.5Hz,J ₂ ＝4.4Hz),2.61(3H,s).

Synthesis of intermediate VI-1: compound V-1 (0.40g, 1.39mmol) was dissolved in a mixed solvent of tetrahydrofuran and methanol (27 mL/15 mL, V/V), a saturated solution (1 mL) of ammonium chloride (0.73g, 13.9mmol) was added, the mixture was heated to 50 ℃ and zinc powder (0.45g, 6.96mmol) was added in portions, and the reaction was continued for 0.5 hour. After the completion of the reaction, LC-MS monitored the reaction, heating was stopped, cooling to room temperature, concentration under reduced pressure was carried out, the residue was dissolved in ethyl acetate (100 mL) and a saturated sodium carbonate solution (100 mL), the organic layer was separated, washed with water and a saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. And (4) carrying out suction filtration, concentrating the filtrate, pulping and purifying methanol to obtain a crude product VI-1, and directly using the crude product VI-1 for the next reaction.

Synthesis of intermediate VII-1: mixing compound VI-1 (0.40g, 1.55mmol) and Boc-L-prolineThe amino acid (0.37g, 1.71 mmol) was dissolved in DMF (50 mL), and the condensing agents PyBOP (1.05g, 2.02mmol) and DIPEA (0.77mL, 4.66 mmol) were added and reacted at room temperature for 12 hours. After completion of the reaction was monitored by TLC, the reaction mixture was added to water (50 mL), extracted with ethyl acetate (30 mL. Times.2), and the organic layers were combined, washed once with water and sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction, concentrated, and purified by column chromatography (dichloromethane: methanol = 50). ¹ H NMR(500 MHz,DMSO-d ₆ )δ(ppm):13.28(1H,s),9.47(1H,s),8.03(1H,s),7.80(1H,s),7.35(1H,s),7.19 (1H,dd,J ₁ ＝8.7Hz,J ₂ ＝2.0Hz),6.86(1H,td,J ₁ ＝8.7Hz,J ₂ ＝2.1Hz),6.81(1H,dd,J ₁ ＝8.4Hz, J ₂ ＝4.3Hz),4.51-4.58(1H,m),3.37-3.57(2H,m),2.59-2.65(1H,m),2.28(3H,s),1.90-2.07(3H, m).

Synthesis of compound a 01: compound VII-1 (100mg, 0.22mmol) was dissolved in ethanol (5 mL), and a saturated HCl solution in ethanol (5 mL) was added to stir the reaction at room temperature. After the completion of the TLC monitoring reaction, the filtrate was concentrated under reduced pressure, and the residue was slurried with ethyl acetate (5 mL) for purification, filtered under suction, and dried in vacuo to give 72mg of a yellow solid in 84% yield. mp:239-241 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.50(1H,s),10.96(1H,s),10.26(1H,s),10.11(1H,brs), 8.68(1H,brs),7.77(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.0Hz),7.74(1H,s),7.68(1H,d,J＝2.8Hz),6.94 (1H,td,J ₁ ＝9.6Hz,J ₂ ＝2.3Hz),6.84-6.87(1H,m),4.46-4.55(1H,m),3.26-3.28(2H,m), 2.39-2.46(1H,m),2.34(3H,s),1.92-1.99(3H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm): 169.9,166.8,159.6,157.7,135.3,127.4(d,J＝9.5Hz),125.7(d,J＝9.2Hz),123.8,121.2,119.5, 116.1(d,J＝2.9Hz),113.0(d,J＝24.0Hz),110.5(d,J＝8.8Hz),106.5(d,J＝25.4Hz),59.5, 46.1,30.5,24.1,9.4.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₁₉ H ₂₀ FN ₄ O ₂ ⁺ ,355.1565；found, 355.1572.

Example 2

Synthesis of (R, Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-2-carboxamide hydrochloride (A02)

Synthesis of intermediate VII-2: compound VI-1 (0.40g, 1.55mmol) and Boc-D-proline (0.37g, 1.71 mmol) were dissolved in DMF (50 mL), and the condensing agents PyBOP (1.05g, 2.02mmol) and DIPEA (0.77mL, 4.66 mmol) were added and reacted at room temperature for 12 hours. After completion of the reaction was monitored by TLC, the reaction mixture was added to water (50 mL), extracted with ethyl acetate (30 mL. Times.2), and the organic layers were combined, washed once with water and sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction, concentrated, and purified by column chromatography (dichloromethane: methanol =50:1 to 20) to obtain 0.53g of a yellow solid in a yield of 75% which was directly used in the next reaction.

Synthesis of compound a 02: compound VII-2 (100mg, 0.22mmol) was dissolved in ethanol (5 mL), and a saturated HCl solution in ethanol (5 mL) was added to stir the reaction at room temperature. After the completion of the TLC monitoring reaction, the filtrate was concentrated under reduced pressure, and the residue was slurried with ethyl acetate (5 mL) and purified, filtered under suction, and dried under vacuum to give 75mg of a yellow solid in 87% yield. mp 247-249 deg.C. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.50(1H,s),10.97(1H,s),10.25(1H,s),10.03(1H,brs), 8.68(1H,brs),7.77(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.0Hz),7.74(1H,s),7.68(1H,d,J＝2.9Hz),6.94 (1H,td,J ₁ ＝9.6Hz,J ₂ ＝2.3Hz),6.84-6.87(1H,m),4.45-4.53(1H,m),3.25-3.29(2H,m), 2.39-2.42(1H,m),2.34(3H,s),1.91-1.99(3H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm): 169.9,166.8,159.6,157.7,135.3,127.4(d,J＝9.4Hz),125.7(d,J＝8.8Hz),123.8,121.2,119.5, 116.1(d,J＝3.0Hz),113.0(d,J＝23.9Hz),110.5(d,J＝8.5Hz),106.5(d,J＝25.4Hz),59.5, 46.1,30.5,24.1,9.4.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₁₉ H ₂₀ FN ₄ O ₂ ⁺ ,355.1565；found, 355.1556.

Example 3

Synthesis of (R, Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-3-carboxamide hydrochloride (A03)

Synthesis of intermediate VII-3: compound VI-1 (0.40g, 1.55mmol) and (R) -1-Boc-3-carboxypyrrolidine (0.37 g, 1.71mmol) were dissolved in DMF (50 mL) and reacted at room temperature for 12 hours with the addition of the condensing agent PyBOP (1.05g, 2.02mmol) and DIPEA (0.77 mL,4.66 mmol). After completion of the TLC monitoring reaction, the reaction mixture was added to water (50 mL), extracted with ethyl acetate (30 mL. Times.2), and the organic layers were combined, washed successively with water and sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction and concentrated, and purified by column chromatography (dichloromethane: methanol =50:1 to 20) to obtain 0.54g of a yellow solid, yield 77%, which was used directly in the next reaction.

Synthesis of compound a 03: compound VII-3 (100mg, 0.22mmol) was dissolved in ethanol (5 mL), and a saturated HCl solution in ethanol (5 mL) was added to stir the reaction at room temperature. After the completion of the TLC monitoring reaction, the filtrate was concentrated under reduced pressure, and the residue was slurried with ethyl acetate (5 mL) and purified, filtered under suction, and dried under vacuum to give 70mg of a yellow solid in 82% yield. mp 259-261 deg.C. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.45(1H,s),10.92(1H,s),9.77(1H,s),9.40(1H,brs), 9.13(1H,brs),7.76(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝1.7Hz),7.72(1H,s),7.67(1H,d,J＝2.4Hz),6.94 (1H,td,J ₁ ＝9.1Hz,J ₂ ＝2.1Hz),6.83-6.86(1H,m),3.28-3.34(2H,m),3.12-3.26(3H,m),2.31 (3H,s),2.23-2.27(1H,m),2.01-2.06(1H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):170.1, 169.9,159.6,157.7,135.2,127.6(d,J＝9.5Hz),125.6(d,J＝9.1Hz),124.5,121.1,119.6,115.7 (d,J＝3.1Hz),113.9(d,J＝24.3Hz),110.5(d,J＝8.7Hz),106.3(d,J＝25.8Hz),47.4,45.2, 42.5,29.6,9.3.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₁₉ H ₂₀ FN ₄ O ₂ ⁺ ,355.1565；found,355.1563.

Example 4

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) piperidine-4-carboxamide hydrochloride (A04)

Synthesis of intermediate VII-4: compound VI-1 (0.40g, 1.55mmol) and Boc-4-piperidinecarboxylic acid (0.39g, 1.71 mmol) were dissolved in DMF (50 mL), and the condensing agents PyBOP (1.05g, 2.02mmol) and DIPEA (0.77mL, 4.66 mmol) were added and reacted at room temperature for 12 hours. After completion of the reaction was monitored by TLC, the reaction mixture was added to water (50 mL), extracted with ethyl acetate (30 mL. Times.2), and the organic layers were combined, washed successively with water and sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction, concentrated, and purified by column chromatography (dichloromethane: methanol =50:1 to 20) to obtain 0.60g of a yellow solid in a yield of 83%, which was directly used in the next reaction step.

Synthesis of compound a 04: compound VII-4 (100mg, 0.22mmol) was dissolved in ethanol (5 mL), and a saturated HCl solution in ethanol (5 mL) was added to stir the reaction at room temperature. After the completion of the TLC monitoring reaction, the filtrate was concentrated under reduced pressure, and the residue was slurried with ethyl acetate (5 mL) and purified, filtered under suction, and dried under vacuum to give 68mg of a yellow solid in 77% yield. mp is 244-246 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):12.60(1H,s),9.67(1H,s),9.18(1H,s),9.02(1H,brs), 8.90(1H,brs),8.09-8.10(1H,m),7.92(1H,d,J＝7.3Hz),7.84(1H,s),7.75-7.79(1H,m), 7.00-7.08(1H,m),3.95-4.03(1H,m),2.98-3.05(2H,m),2.77-2.90(2H,m),2.34(3H,s), 2.06-2.17(2H,m),1.78-1.88(2H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.9,167.9, 159.6,157.7,135.2,127.5(d,J＝9.4Hz),125.6(d,J＝11.6Hz),124.6,120.8,119.4,115.7(d,J ＝3.1Hz),112.9(d,J＝23.7Hz),110.4(d,J＝8.3Hz),106.3(d,J＝25.3Hz),56.5,45.0,37.5, 30.3,23.4,9.4.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₀ H ₂₂ FN ₄ O ₂ ⁺ ,369.1721；found,369.1741.

Example 5

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) piperidine-2-carboxamide hydrochloride (A05)

Synthesis of intermediate VII-5: compound VI-1 (0.40g, 1.55mmol) and N-Boc-2-piperidinecarboxylic acid (0.39g, 1.71 mmol) were dissolved in DMF (50 mL), and the condensing agents PyBOP (1.05g, 2.02mmol) and DIPEA (0.77mL, 4.66 mmol) were added and reacted at room temperature for 12 hours. After completion of the TLC monitoring reaction, the reaction mixture was added to water (50 mL), extracted with ethyl acetate (30 mL. Times.2), and the organic layers were combined, washed successively with water and sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction, concentrated, and purified by column chromatography (dichloromethane: methanol =50:1 to 20) to obtain 0.57g of a yellow solid in 78% yield, which was used directly in the next reaction step.

Synthesis of compound a 05: compound VII-5 (100mg, 0.22mmol) was dissolved in ethanol (5 mL), and a saturated HCl solution in ethanol (5 mL) was added to stir the reaction at room temperature. After the completion of the TLC monitoring reaction, the filtrate was concentrated under reduced pressure, and the residue was slurried with ethyl acetate (5 mL) and purified, filtered under suction, and dried under vacuum to give 72mg of a yellow solid in 81% yield. mp is 263-265 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.50(1H,s),10.95(1H,s),10.13(1H,s),9.26-9.28(1H, m),8.76-8.82(1H,m),7.77(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.1Hz),7.74(1H,s),7.68(1H,d,J＝2.7 Hz),6.94(1H,td,J ₁ ＝9.6Hz,J ₂ ＝2.3Hz),6.84-6.87(1H,m),4.00-4.06(1H,m),3.25-3.28(1H, m),2.92-2.99(1H,m),2.32(3H,s),2.22-2.25(1H,m),1.82-1.85(1H,m),1.52-1.76(4H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.9,167.2,159.6,157.7,135.3,127.4(d,J＝9.5Hz), 125.7(d,J＝10.4Hz),123.7,121.3,119.5,116.1(d,J＝2.9Hz),113.0(d,J＝24.1Hz),110.5(d, J＝8.6Hz),106.4(d,J＝25.4Hz),57.6,43.8,27.9,22.1,21.7,9.4.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₀ H ₂₂ FN ₄ O ₂ ⁺ ,369.1721；found,369.1716.

Example 6

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (pyrrolidin-1-yl) acetamide (A06)

Synthesis of intermediate VIII-1: compound VI-1 (0.30g, 1.15mmol) was dissolved in THF (20 mL), DIPEA (0.57mL, 3.45mmol) was added, the temperature was lowered to 0 ℃ and chloroacetyl chloride (0.46mL, 5.75mmol) was added dropwise and after completion of addition, the reaction was carried out at room temperature for 0.5h. After the completion of the TLC monitoring reaction, concentration was performed under reduced pressure, and the residue was slurried with ethyl acetate (10 mL) and methanol (0.5 mL). Suction filtration, the filter cake washed with ethyl acetate (1 mL) and dried in vacuo to give 0.30g of a yellow solid in 76% yield. ¹ H NMR(500 MHz,DMSO-d ₆ )δ(ppm):13.46(1H,s),10.90(1H,s),9.75(1H,s),7.77(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.2Hz),7.73(1H,s),7.68(1H,d,J＝2.9Hz),6.94(1H,td,J ₁ ＝8.4Hz,J ₂ ＝2.4Hz),6.83-6.85 (1H,m),4.29(2H,s),2.30(3H,s).

Synthesis of compound a 06: compound VIII-1 (100mg, 0.3mmol) was dissolved in DMF (5 mL) and addedPyrrolidine (123. Mu.L, 1.50 mmol) was reacted at 50 ℃ for 12 hours. After TLC monitoring of the completion of the reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed successively with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. Suction filtration, filtrate concentration, residue adding ethyl acetate (5 mL), pulping and purification, suction filtration, vacuum drying to obtain yellow solid 88mg, yield 80%. mp is 250-252 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.44(1H,s), 10.89(1H,s),9.17(1H,s),7.76(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝1.9Hz),7.73(1H,s),7.62(1H,d,J＝ 2.5Hz),6.94(1H,td,J ₁ ＝10.0Hz,J ₂ ＝2.0Hz),6.83-6.85(1H,m),3.26(2H,s),2.57-2.66(4H, m),2.27(3H,s),1.71-1.81(4H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.9,168.2,159.6, 157.7,135.2,127.5(d,J＝9.3Hz),125.7(d,J＝6.3Hz),124.3,121.4,119.6,115.6(d,J＝3.0Hz), 112.8(d,J＝23.8Hz),110.4(d,J＝8.6Hz),106.3(d,J＝25.4Hz),59.0,54.3(2C),31.1,23.9, 9.1.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₀ H ₂₂ FN ₄ O ₂ ⁺ ,369.1721；found,369.1718.

Example 7

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (pyrrolidin-2-yl) acetamide hydrochloride (A07)

Synthesis of intermediate VII-7: compound VI-1 (0.40g, 1.55mmol) and 2- (1- (tert-butoxycarbonyl) pyrrolidin-2-yl) acetic acid (0.39g, 1.71mmol) were dissolved in DMF (50 mL), and the condensing agents PyBOP (1.05g, 2.02mmol) and DIPEA (0.77mL, 4.66mmol) were added and reacted at room temperature for 12 hours. After completion of the reaction was monitored by TLC, the reaction mixture was added to water (50 mL), extracted with ethyl acetate (30 mL. Times.2), and the organic layers were combined, washed successively with water and sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction, concentrated, and purified by column chromatography (dichloromethane: methanol =50:1 to 20) to obtain 0.58g of a yellow solid in 78% yield, which was used directly in the next reaction step.

Synthesis of compound a 07: compound VII-7 (100mg, 0.22mmol) was dissolved in ethanol (5 mL), and a saturated HCl solution in ethanol (5 mL) was added to stir the reaction at room temperature. After the completion of the TLC monitoring reaction, the filtrate was concentrated under reduced pressure, and the residue was slurried with ethyl acetate (5 mL) and purified, filtered under suction, and dried under vacuum to give 70mg of a yellow solid in 81% yield. mp is 234-236 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.45(1H,s),10.93(1H,s),9.76(1H,s),9.25(1H,brs), 9.05(1H,brs),7.76(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.4Hz),7.72(1H,s),7.69(1H,d,J＝2.9Hz),6.93 (1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.5Hz),6.83-6.86(1H,m),3.73-3.79(1H,m),3.13-3.18(2H,m),2.91 (2H,d,J＝6.9Hz),2.32(3H,s),2.10-2.18(1H,m),1.93-1.96(2H,m),1.81-1.91(1H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.8,168.0,159.6,157.7,135.2,127.5(d,J＝9.4Hz), 125.7(d,J＝12.1Hz),124.6,120.8,119.4,115.7(d,J＝2.9Hz),112.9(d,J＝24.2Hz),110.4(d, J＝8.5Hz),106.3(d,J＝25.5Hz),56.5,44.9,37.5,30.3,23.4,9.4.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₀ H ₂₂ FN ₄ O ₂ ⁺ ,369.1721；found,369.1717.

Example 8

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (piperidin-1-yl) acetamide (A08)

Synthesis of compound a 08: compound VIII-1 (100mg, 0.3 mmol) was dissolved in DMF (5 mL), and piperidine (149. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After TLC monitoring of the completion of the reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), the organic layers were combined, washed successively with water and saturated sodium chloride solution, and then dried with anhydrous sulfuric acidAnd (4) drying sodium. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 89mg of a yellow solid in 78% yield. mp 275-277 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.44(1H,s), 10.89(1H,s),9.17(1H,s),7.77(1H,dd,J ₁ ＝9.1Hz,J ₂ ＝1.4Hz),7.74(1H,s),7.65(1H,d,J＝ 2.2Hz),6.94(1H,td,J ₁ ＝10.0Hz,J ₂ ＝2.0Hz),6.83-6.85(1H,m),3.08(2H,s),2.42-2.50(4H, m),2.28(3H,s),1.57-1.60(4H,m),1.37-1.47(2H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm): 169.9,167.9,159.6,157.7,135.2,127.5(d,J＝9.5Hz),125.6(d,J＝7.2Hz),124.3,120.5,119.0, 115.7(d,J＝2.8Hz),112.9(d,J＝24.1Hz),110.4(d,J＝8.5Hz),106.4(d,J＝25.4Hz),62.3, 54.6(2C),26.2(2C),23.9,8.9.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₄ FN ₄ O ₂ ⁺ ,383.1878； found,383.1872.

Example 9

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -2- (4-methylpiperazin-1-yl) acetamide (A09)

Synthesis of compound a 09: compound VIII-1 (100mg, 0.3 mmol) was dissolved in DMF (5 mL), and N-methylpiperazine (166. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After the completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed successively with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 101mg of yellow solid in 85% yield. mp 265-267 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm): 13.44(1H,s),10.89(1H,s),9.18(1H,s),7.76(1H,dd,J ₁ ＝9.4Hz,J ₂ ＝2.4Hz),7.73(1H,s), 7.65(1H,d,J＝2.9Hz),6.94(1H,td,J ₁ ＝9.5Hz,J ₂ ＝2.5Hz),6.83-6.85(1H,m),3.12(2H,s), 2.55-2.64(4H,m),2.33-2.45(4H,m),2.28(3H,s),2.19(3H,s). ¹³ C NMR(126MHz,DMSO-d ₆ ) δ(ppm):169.9,167.7,159.6,157.7,135.2,127.5(d,J＝9.5Hz),125.6(d,J＝7.0Hz),124.3, 120.7,119.2,115.7(d,J＝3.0Hz),112.9(d,J＝24.1Hz),110.4(d,J＝8.6Hz),106.4(d,J＝25.2 Hz),61.4,55.2(2C),53.2(2C),46.2,9.0.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₅ FN ₅ O ₂ ⁺ , 398.1987；found,398.1984.

Example 10

Synthesis of (Z) -2- (diethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) acetamide (A10)

Synthesis of compound a 10: compound VIII-1 (100mg, 0.3mmol) was dissolved in DMF (5 mL), to which was added diethylamine (155. Mu.L, 1.5 mmol), followed by stirring the reaction at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 80mg of a yellow solid in 72% yield. mp 265-267 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.44(1H, s),10.89(1H,s),9.21(1H,s),7.77(1H,d,J＝9.2Hz),7.74(1H,s),7.65-7.70(1H,m),6.94(1H, td,J ₁ ＝9.0Hz,J ₂ ＝1.8Hz),6.83-6.85(1H,m),3.15(2H,s),2.60(4H,q,J＝6.8Hz),2.28(3H,s), 1.05(6H,t,J＝7.0Hz). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.9,169.1,159.6,157.7, 135.2,127.5(d,J＝9.4Hz),125.6(d,J＝10.3Hz),124.3,120.3,118.9,115.8(d,J＝2.9Hz), 112.9(d,J＝21.4Hz),110.4(d,J＝8.6Hz),106.4(d,J＝13.9Hz),57.5,48.4(2C),12.7(2C), 8.9.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₀ H ₂₄ FN ₄ O ₂ ⁺ ,371.1878；found,371.1876.

Example 11

Synthesis of (Z) -2- (dimethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) acetamide (A11)

Synthesis of compound a 11: compound VIII-1 (100mg, 0.3 mmol) was dissolved in DMF (5 mL), and dimethylamine (76. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to yield 76mg of yellow solid in 74% yield. mp is 271-273 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.44(1H, s),10.89(1H,s),9.16(1H,s),7.76(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.0Hz),7.73(1H,s),7.61(1H,d,J ＝2.7Hz),6.93(1H,td,J ₁ ＝9.8Hz,J ₂ ＝2.2Hz),6.83-6.85(1H,m),3.09(2H,s),2.31(6H,s), 2.27(3H,s). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.9,168.2,159.6,157.7,135.2,127.5(d, J＝9.5Hz),125.7(d,J＝8.2Hz),124.3,121.4,119.7,115.6(d,J＝2.9Hz),112.9(d,J＝24.0 Hz),110.4(d,J＝8.5Hz),106.4(d,J＝25.6Hz),63.0,45.9(2C),9.1.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₁₈ H ₂₀ FN ₄ O ₂ ⁺ ,343.1565；found,343.1560.

Example 12

Synthesis of (Z) -2- (cyclopentylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) acetamide (A12)

Synthesis of compound a 12: compound VIII-1 (100mg, 0.3 mmol) was dissolved in DMF (5 mL), cyclopentylamine (148. Mu.L, 1.5 mmol) was added thereto, followed by stirring at 50 ℃And (4) stirring and reacting. After TLC monitoring of the completion of the reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed successively with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 95mg of yellow solid, 83% yield. mp:283-285 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.43(1H, s),10.88(1H,s),9.37(1H,s),7.76(1H,dd,J ₁ ＝9.4Hz,J ₂ ＝1.7Hz),7.73(1H,s),7.70(1H,d,J ＝2.5Hz),6.93(1H,td,J ₁ ＝9.7Hz,J ₂ ＝1.8Hz),6.83-6.85(1H,m),3.27(2H,s),3.02-3.07(1H, m),2.28(3H,s),1.62-1.75(4H,m),1.47-1.54(2H,m),1.34-1.39(2H,m). ¹³ C NMR(126MHz, DMSO-d ₆ )δ(ppm):169.9,169.8,159.6,157.7,135.2,127.5(d,J＝9.4Hz),125.6(d,J＝10.7 Hz),124.5,120.1,118.8,115.7(d,J＝3.0Hz),112.9(d,J＝24.1Hz),110.4(d,J＝8.4Hz),106.4 (d,J＝25.6Hz),59.7,51.3,32.8(2C),23.9(2C),9.0.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₄ FN ₄ O ₂ ⁺ ,383.1878；found,383.1873.

Example 13

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propionamide (A13)

Synthesis of intermediate IX-1: compound VI-1 (0.30g, 1.15mmol) was dissolved in THF (20 mL), DIPEA (0.57mL, 3.45mmol) was added, the temperature was lowered to 0 deg.C, 3-chloropropionyl chloride (0.55mL, 5.75mmol) was added dropwise, and after completion of the addition, the reaction was carried out at room temperature for 0.5h. After the completion of the TLC monitoring reaction, concentration was performed under reduced pressure, and the residue was slurried with ethyl acetate (10 mL) and methanol (0.5 mL). Suction filtration was carried out, and the filter cake was washed with ethyl acetate (1 mL) and dried in vacuo to give 0.32g of a yellow solid in 80% yield. ¹ H NMR(500 MHz,DMSO-d ₆ )δ(ppm):13.44(1H,s),10.88(1H,s),9.50(1H,s),7.75(1H,dd,J ₁ ＝9.4Hz,J ₂ ＝2.5Hz),7.72(1H,s),7.68(1H,d,J＝3.1Hz),6.93(1H,td,J ₁ ＝9.6Hz,J ₂ ＝2.5Hz),6.83-6.85 (1H,m),3.88(2H,t,J＝6.4Hz),2.85(2H,t,J＝6.3Hz),2.30(3H,s).

Synthesis of compound a 13: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), and pyrrolidine (123. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After TLC monitoring of the completion of the reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed successively with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 90mg of a yellow solid, yield 79%. mp at 274-276 deg.C. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.40(1H, s),10.88(1H,s),9.99(1H,s),7.74(1H,dd,J ₁ ＝9.2Hz,J ₂ ＝1.0Hz),7.72(1H,s),7.71(1H,d,J ＝3.2Hz),6.93(1H,td,J ₁ ＝8.9Hz,J ₂ ＝2.4Hz),6.82-6.85(1H,m),2.72(2H,t,J＝6.9Hz), 2.51-2.56(4H,m),2.47-2.49(2H,m),2.27(3H,s),1.68-1.76(4H,m). ¹³ C NMR(126MHz, DMSO-d ₆ )δ(ppm):169.8,169.7,159.6,157.7,135.2,127.6(d,J＝9.3Hz),125.6(d,J＝23.3 Hz),125.3,119.8,118.9,115.5(d,J＝2.8Hz),112.8(d,J＝23.8Hz),110.3(d,J＝8.5Hz),106.4 (d,J＝25.4Hz),53.7(2C),52.0,35.1,23.6(2C),9.0.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₄ FN ₄ O ₂ ⁺ ,383.1878；found,383.1866.

Example 14

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (piperidin-1-yl) propanamide (A14)

Synthesis of compound a 14: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), to whichPiperidine (149. Mu.L, 1.5 mmol) was added, and the reaction was stirred at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 87mg of a yellow solid in 73% yield. mp is 270-272 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.40(1H,s), 10.88(1H,s),9.83(1H,s),7.74(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.4Hz),7.72(1H,s),7.70(1H,d,J＝ 3.0Hz),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.5Hz),6.82-6.85(1H,m),2.58(2H,t,J＝6.6Hz), 2.37-2.48(6H,m),2.31(3H,s),1.52-1.56(4H,m),1.38-1.45(2H,m). ¹³ C NMR(126MHz, DMSO-d ₆ )δ(ppm):169.9,169.8,159.6,157.8,135.1,127.6(d,J＝9.2Hz),125.5(d,J＝25.1 Hz),125.2,120.0,119.1,115.5(d,J＝3.0Hz),112.8(d,J＝24.6Hz),110.4(d,J＝8.6Hz),106.3 (d,J＝25.7Hz),55.1,54.1(2C),33.2,26.0(2C),24.5,9.3.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₂ H ₂₆ FN ₄ O ₂ ⁺ ,397.2034；found,397.2022.

Example 15

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (4-methylpiperazin-1-yl) propanamide (A15)

Synthesis of compound a 15: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), and N-methylpiperazine (166. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After the completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed successively with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 103mg of a yellow solid in 84% yield. mp is 285-287 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm): 13.40(1H,s),10.88(1H,s),9.74(1H,s),7.74(1H,d,J＝9.3Hz),7.72(1H,s),7.66-7.69(1H,m), 6.93(1H,t,J ₁ ＝7.8Hz),6.82-6.85(1H,m),2.61(2H,t,J＝6.0Hz),2.35-2.48(10H,m),2.31 (3H,s),2.16(3H,s). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.8,169.7,159.6,157.7,135.1, 127.6(d,J＝9.4Hz),125.5(d,J＝25.0Hz),125.2,120.1,119.2,115.5(d,J＝3.0Hz),112.8(d,J ＝23.9Hz),110.4(d,J＝8.4Hz),106.3(d,J＝25.5Hz),55.2(2C),54.4,52.8(2C),46.2,33.2, 9.6.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₂ H ₂₇ FN ₅ O ₂ ⁺ ,412.2143；found,412.2137.

Example 16

Synthesis of (Z) -3- (diethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide (A16)

Synthesis of compound a 16: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), and diethylamine (155. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 84mg of a yellow solid in 73% yield. mp:251-253 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.41(1H, s),10.87(1H,s),9.86(1H,s),7.74(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝1.6Hz),7.71(1H,s),7.69(1H,d,J ＝2.5Hz),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.3Hz),6.83-6.85(1H,m),2.72(2H,t,J＝6.8Hz), 2.53-2.56(4H,m),2.44(2H,t,J＝6.7Hz),2.29(3H,s),1.00(6H,t,J＝7.1Hz). ¹³ C NMR(126 MHz,DMSO-d ₆ )δ(ppm):170.0,169.8,159.6,157.7,135.1,127.5(d,J＝9.4Hz),125.5(d,J＝ 21.5Hz),125.3,120.0,119.1,115.4(d,J＝3.0Hz),112.8(d,J＝24.0Hz),110.4(d,J＝8.6Hz), 106.4(d,J＝25.6Hz),49.0,46.2(2C),33.4,12.0(2C),9.2.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₆ FN ₄ O ₂ ⁺ ,385.2034；found,385.2030.

Example 17

Synthesis of (Z) -3- (dimethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide (A17)

Synthesis of compound a 17: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), and dimethylamine (76. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After TLC monitoring of the completion of the reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed successively with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to yield 69mg of yellow solid, 65% yield. mp is 270-272 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.41 (1H,s),10.88(1H,s),9.84(1H,s),7.75(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.2Hz),7.72(1H,s),7.69(1H, d,J＝2.9Hz),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.3Hz),6.83-6.85(1H,m),2.57(2H,t,J＝6.9Hz), 2.46(2H,t,J＝6.7Hz),2.28(3H,s),2.22(6H,s). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm): 169.8,169.7,159.6,157.7,135.1,127.6(d,J＝9.4Hz),125.5(d,J＝23.4Hz),125.2,120.1, 119.1,115.5(d,J＝3.0Hz),112.8(d,J＝23.9Hz),110.4(d,J＝8.6Hz),106.4(d,J＝25.5Hz), 55.6,45.2(2C),33.9,9.1.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₁₉ H ₂₂ FN ₄ O ₂ ⁺ ,357.1721；found, 357.1707.

Example 18

Synthesis of (Z) -3- (ethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide (A18)

Synthesis of compound a 18: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), to which was added ethylamine (83. Mu.L, 1.5 mmol), and the reaction was stirred at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 76mg of a yellow solid, yield 71%. mp is 252-254 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.41(1H,s), 10.88(1H,s),9.84(1H,s),7.75(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝2.2Hz),7.72(1H,s),7.70(1H,d,J＝ 2.9Hz),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.3Hz),6.83-6.85(1H,m),2.81(2H,t,J＝6.3Hz), 2.58-2.63(2H,m),2.45(2H,t,J＝6.2Hz),2.29(3H,s),1.06(3H,t,J＝7.1Hz). ¹³ C NMR(126 MHz,DMSO-d ₆ )δ(ppm):170.0,169.8,159.6,157.7,135.1,127.6(d,J＝9.2Hz),125.5(d,J＝ 24.1Hz),125.3,120.0,119.1,115.4(d,J＝2.9Hz),112.8(d,J＝23.9Hz),110.3(d,J＝8.2Hz), 106.4(d,J＝25.3Hz),45.7,43.6,35.8,15.4,9.2.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₁₉ H ₂₂ FN ₄ O ₂ ⁺ ,357.1721；found,357.1715.

Example 19

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (propylamino) propionamide (A19)

Synthesis of compound a 19: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), to which was added n-propylamine (123. Mu.L, 1.5 mmol), and the reaction was stirred at 50 ℃. After TLC monitoring of the completion of the reaction, the reaction mixture was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), the organic layers were combined, and dissolved in water and saturated sodium chloride in that orderThe solution was washed and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 83mg of a yellow solid in 75% yield. mp 243-245 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.40(1H, s),10.88(1H,s),10.00(1H,s),7.74(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝1.5Hz),7.71(1H,s),7.69(1H,d,J ＝2.0Hz),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.3Hz),6.83-6.85(1H,m),2.79(2H,t,J＝6.3Hz), 2.53-2.58(2H,m),2.44(2H,t,J＝6.4Hz),2.29(3H,s),1.43-1.48(2H,m),0.89(3H,t,J＝7.3 Hz). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):170.0,169.8,159.6,157.7,135.1,127.6(d,J＝9.5 Hz),125.5(d,J＝20.9Hz),125.3,120.1,119.1,115.4(d,J＝2.9Hz),112.8(d,J＝24.1Hz), 110.3(d,J＝8.6Hz),106.4(d,J＝25.4Hz),51.3,45.9,35.9,23.0,12.3,9.2.HRMS(ESI):m/z [M+H] ⁺ calcd for C ₂₀ H ₂₄ FN ₄ O ₂ ⁺ ,371.1878；found,371.1873.

Example 20

Synthesis of (Z) -3- (tert-butylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide (A20)

Synthesis of compound a 20: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), to which was added tert-butylamine (158. Mu.L, 1.5 mmol), and the reaction was stirred at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 94mg of a yellow solid in 82% yield. mp is 237-239 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.40(1H, s),10.87(1H,s),10.11(1H,s),7.74(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝1.5Hz),7.71(1H,s),7.69-7.70 (1H,m),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.3Hz),6.82-6.85(1H,m),2.78(2H,t,J＝6.1Hz),2.42 (2H,t,J＝6.1Hz),2.30(3H,s),1.09(9H,s). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):170.0, 169.8,159.6,157.7,135.1,127.6(d,J＝9.6Hz),125.5(d,J＝21.4Hz),125.3,119.8,119.0, 115.4(d,J＝2.9Hz),112.8(d,J＝23.9Hz),110.4(d,J＝8.5Hz),106.2(d,J＝25.4Hz),50.6, 38.8,36.8,29.0(3C),9.4.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₆ FN ₄ O ₂ ⁺ ,385.2034；found, 385.2029.

Example 21

Synthesis of (Z) -3- (cyclopentylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide (A21)

Synthesis of compound a 21: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), and cyclopentylamine (148. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After TLC monitoring of the completion of the reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed successively with water and saturated sodium chloride solution, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 90mg of a yellow solid in 76% yield. mp is 258-260 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.40(1H, s),10.88(1H,s),10.06(1H,s),7.74(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝1.5Hz),7.71(1H,s),7.69(1H,d,J ＝2.5Hz),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.3Hz),6.82-6.85(1H,m),3.02-3.07(1H,m),2.78(2H, t,J＝6.3Hz),2.43(2H,t,J＝6.2Hz),2.29(3H,s),1.74-1.81(2H,m),1.59-1.66(2H,m), 1.46-1.53(2H,m),1.29-1.36(2H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):170.2,169.8, 159.6,157.7,135.1,127.6(d,J＝9.4Hz),125.5(d,J＝20.5Hz),125.3,120.0,119.1,115.4(d,J ＝2.9Hz),112.8(d,J＝23.9Hz),110.4(d,J＝8.5Hz),106.2(d,J＝25.5Hz),59.5,44.6,36.2, 32.9(2C),24.1(2C),9.2.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₂ H ₂₆ FN ₄ O ₂ ⁺ ,397.2034；found, 397.2030.

Example 22

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3-morpholinepropionamide (A22)

Synthesis of compound a 22: compound IX-1 (104mg, 0.3 mmol) was dissolved in DMF (5 mL), to which morpholine (131. Mu.L, 1.5 mmol) was added, and then the reaction was stirred at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 92mg of a yellow solid in 77% yield. mp:257-259 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.40(1H,s), 10.88(1H,s),9.61(1H,s),7.74(1H,dd,J ₁ ＝9.3Hz,J ₂ ＝1.5Hz),7.71(1H,s),7.68(1H,d,J＝ 2.8Hz),6.93(1H,td,J ₁ ＝9.4Hz,J ₂ ＝2.3Hz),6.82-6.85(1H,m),3.57-3.64(4H,m),2.62(2H,t, J＝6.9Hz),2.37-2.50(6H,m),2.30(3H,s). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.9, 169.7,159.6,157.7,135.1,127.6(d,J＝9.5Hz),125.5(d,J＝19.6Hz),125.1,120.4,119.3, 115.4(d,J＝3.1Hz),112.8(d,J＝24.0Hz),110.4(d,J＝8.5Hz),106.3(d,J＝25.5Hz),66.6 (2C),54.8,53.5(2C),33.1,9.3.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₄ FN ₄ O ₃ ⁺ ,399.1827； found,399.1826.

Example 23

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -6- (trifluoromethyl) nicotinamide (A23)

Synthesis of compound a 23: compound VI-1 (0.40g, 1.55mmol) and 4-trifluoromethylbenzoic acid (0.59g, 3.10 mmol) were dissolved in DMF (50 mL) and the condensing agents PyBOP (1.05g, 2.02mmol) and DIPEA (0.77mL, 4.66 mmol) were added and reacted at room temperature for 12 hours. After completion of the reaction was monitored by TLC, the reaction mixture was added to water (50 mL), extracted with ethyl acetate (30 mL. Times.2), and the organic layers were combined, washed successively with water and sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered with suction and concentrated, and purified by column chromatography (dichloromethane: methanol = 50. mp:>300℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.57(1H,s),10.94(1H,s),10.21(1H,s),9.26(1H,s), 8.57(1H,d,J＝6.8Hz),8.11(1H,d,J＝8.2Hz),7.79(1H,d,J＝2.4Hz),7.78(1H,s),7.74(1H, d,J＝3.0Hz),6.95(1H,td,J ₁ ＝9.5Hz,J ₂ ＝2.5Hz),6.84-6.87(1H,m),2.37(3H,s). ¹³ C NMR (126MHz,DMSO-d ₆ )δ(ppm):169.9,162.9,157.7,149.8,138.3,135.2,133.8,127.5(d,J＝9.4 Hz),125.8(d,J＝23.0Hz),125.1,124.0,123.0,121.1,121.0,120.8,120.7,116.0(d,J＝3.0Hz), 113.0(d,J＝24.5Hz),110.5(d,J＝8.7Hz),106.4(d,J＝24.9Hz),9.5.HRMS(ESI):m/z [M+H] ⁺ calcd for C ₂₁ H ₁₅ F ₄ N ₄ O ₂ ⁺ ,431.1131；found,431.1114.

example 24

Synthesis of (Z) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -4-methylpiperazine-1-carboxamide (A24)

Synthesis of compound a 24: compound VI-1 (0.40g, 1.55mmol) was dissolved in THF (20 mL), DIPEA (0.51mL, 3.1 mmol) and p-nitrophenyl chloroformate (469mg, 2.32mmol) were added. After 20min at room temperature, N-methylpiperazine (687. Mu.L, 6.2 mmol) was added to the reaction mixture, and the reaction was continued for 0.5h. After the completion of the reaction was detected by LC-MS, the reaction solution was concentrated under reduced pressure and purified by column chromatography (dichloromethane: methanol =1000.41g, yield 70%. mp:257-259 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.43(1H,s),10.84(1H,s),7.91(1H,s), 7.73(1H,dd,J ₁ ＝9.4Hz,J ₂ ＝2.2Hz),7.70(1H,s),7.37(1H,d,J＝2.8Hz),6.91(1H,td,J ₁ ＝ 9.0Hz,J ₂ ＝2.4Hz),6.82-6.85(1H,m),3.39-3.51(4H,m),2.35-2.50(4H,m),2.28(3H,s),2.23 (3H,s). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.9,159.5,157.7,156.1,135.0,127.6(d,J＝9.4Hz),125.9(d,J＝19.8Hz),124.4,121.6 118.1,114.7(d,J＝2.9Hz),112.8(d,J＝24.4Hz), 110.3(d,J＝8.7Hz),106.2(d,J＝25.2Hz),66.6(2C),54.9(2C),46.1,44.1(2C),9.3.HRMS (ESI):m/z[M+H] ⁺ calcd for C ₂₀ H ₂₃ FN ₅ O ₂ ⁺ ,384.1836；found,384.1826.

Example 25

Synthesis of (Z) -N- (5- ((5-chloro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propionamide (A25)

Synthesis of intermediate V-2: compound III (1.2g, 7.79mmol) and IV-2 (1.4g, 8.56mmol) were dissolved in ethanol (10 mL), and then pyrrolidine (0.83mL, 10.13mmol) was added and heated to 50 ℃ for 2 hours. After the LC-MS monitoring reaction is finished, stopping heating, cooling to room temperature, performing suction filtration, washing a filter cake with a small amount of ethanol, and performing vacuum drying to obtain orange yellow solid 2.3g with the yield of 85%. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):14.19(1H,brs),11.25(1H,s),8.28 (1H,dd,J1＝3.6Hz,J ₂ ＝1.4Hz),8.11(1H,s),7.88(1H,s),7.22(1H,dd,J ₁ ＝8.3Hz,J ₂ ＝1.9 Hz),6.88(1H,dd,J ₁ ＝8.3Hz,J ₂ ＝1.4Hz),2.60(3H,s).

Synthesis of intermediate VI-1: compound V-2 (0.42g, 1.39mmol) was dissolved in a mixed solvent of tetrahydrofuran and methanol (27 mL/15 mL, V/V), a saturated solution (1 mL) of ammonium chloride (0.73g, 13.9 mmol) was added, heated to 50 deg.C, zinc powder (0.45g, 6.96mmol) was added in portions, and the reaction was continued for 0.5 hour. After the completion of the reaction, LC-MS monitored the reaction, heating was stopped, cooling to room temperature, concentration under reduced pressure was carried out, the residue was dissolved in ethyl acetate (100 mL) and a saturated sodium carbonate solution (100 mL), the organic layer was separated, washed with water and a saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. And (3) carrying out suction filtration, concentrating the filtrate, pulping and purifying methanol to obtain a crude product VI-1, and directly using the crude product VI-1 in the next reaction.

Synthesis of intermediate IX-2: compound VI-2 (0.31g, 1.15mmol) was dissolved in THF (20 mL), DIPEA (0.57mL, 3.45mmol) was added, the temperature was lowered to 0 deg.C, 3-chloropropionyl chloride (0.55mL, 5.75mmol) was added dropwise, and after completion of addition, reaction was carried out at room temperature for 0.5h. After the completion of the TLC monitoring reaction, concentration was performed under reduced pressure, and the residue was slurried with ethyl acetate (10 mL) and methanol (0.5 mL). Suction filtration, filter cake washing with ethyl acetate (1 mL), vacuum drying to obtain yellow solid 0.31g, yield 75%. ¹ H NMR(500 MHz,DMSO-d ₆ )δ(ppm):13.40(1H,s),11.00(1H,s),9.52(1H,s),7.98((1H,d,J＝1.7Hz), 7.78(1H,s),7.69(1H,d,J＝3.0Hz),7.13(1H,dd,J ₁ ＝8.3Hz,J ₂ ＝2.0Hz),6.86(1H,d,J＝8.2 Hz),3.88(2H,t,J＝6.3Hz),2.85(2H,t,J＝6.3Hz),2.30(3H,s).

Synthesis of compound a 25: compound IX-2 (109mg, 0.3 mmol) was dissolved in DMF (5 mL), and pyrrolidine (123. Mu.L, 1.5 mmol) was added thereto, followed by stirring at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Filtering, concentrating the filtrate, adding ethyl acetate (5 mL) into the residue, pulping, purifying, filtering, and vacuum filteringDrying gave 96mg of a yellow solid in 80% yield. mp:251-253 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.35(1H, s),10.99(1H,s),9.99(1H,s),7.97(1H,d,J ₁ ＝1.9Hz),7.77(1H,s),7.71(1H,d,J＝3.0Hz), 7.13(1H,dd,J ₁ ＝8.2Hz,J ₂ ＝2.0Hz),6.86(1H,d,J＝8.2Hz),2.72-2.74(2H,m),2.51-2.57(4H, m),2.46-2.50(2H,m),2.28(3H,s),1.71-1.76(4H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm): 169.6,137.5,128.0,126.0,125.9,125.8,125.6,125.3,123.1,120.1,119.2,118.9,114.7,111.1, 53.7(2C),51.8,35.1,23.6(2C),9.0.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₄ ClN ₄ O ₂ ⁺ , 399.1582；found,399.1579.

Example 26

Synthesis of (Z) -N- (5- ((5-bromo-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propionamide (A26)

Synthesis of intermediate V-3: compound III (1.2g, 7.79mmol) and IV-3 (1.8g, 8.56mmol) were dissolved in ethanol (10 mL), and pyrrolidine (0.83mL, 10.13mmol) was added and heated to 50 ℃ for 2 hours. After the LC-MS monitoring reaction is finished, stopping heating, cooling to room temperature, performing suction filtration, washing a filter cake with a small amount of ethanol, and performing vacuum drying to obtain 1.84g of orange yellow solid with the yield of 80%. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):14.19(1H,brs),11.26(1H,s),8.28 (1H,d,J＝3.5Hz),8.24(1H,s),7.90(1H,s),7.36(1H,d,J＝8.3Hz),6.85(1H,d,J＝8.3Hz), 2.61(3H,s).

Synthesis of intermediate VI-3: compound V-3 (0.48g, 1.39mmol) was dissolved in a mixed solvent of tetrahydrofuran and methanol (27 mL/15 mL, V/V), a saturated solution (1 mL) of ammonium chloride (0.73g, 13.9mmol) was added, the mixture was heated to 50 ℃ and zinc powder (0.45g, 6.96mmol) was added in portions, and the reaction was continued for 0.5 hour. After the completion of the reaction, LC-MS monitored the reaction, heating was stopped, cooling to room temperature, concentration under reduced pressure was carried out, the residue was dissolved in ethyl acetate (100 mL) and a saturated sodium carbonate solution (100 mL), the organic layer was separated, washed with water and a saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. And (4) carrying out suction filtration, concentrating the filtrate, pulping and purifying methanol to obtain a crude product VI-3, and directly using the crude product VI-3 in the next reaction.

Synthesis of intermediate IX-3: compound VI-3 (0.36g, 1.15mmol) was dissolved in THF (20 mL), DIPEA (0.57mL, 3.45mmol) was added, the temperature was lowered to 0 deg.C, 3-chloropropionyl chloride (0.55mL, 5.75mmol) was added dropwise, and after the addition was completed, the reaction was carried out at room temperature for 0.5h. After the completion of the TLC monitoring reaction, concentration was performed under reduced pressure, and the residue was slurried with ethyl acetate (10 mL) and methanol (0.5 mL). Suction filtration, filter cake washing with ethyl acetate (1 mL), vacuum drying to obtain yellow solid 0.30g, yield 65%. ¹ H NMR(500 MHz,DMSO-d ₆ )δ(ppm):13.40(1H,s),11.01(1H,s),9.52(1H,s),8.10(1H,s),7.78(1H,s), 7.69(1H,d,J＝2.9Hz),7.27(1H,dd,J ₁ ＝8.2Hz,J ₂ ＝0.7Hz),6.82(1H,d,J＝8.2Hz),3.88(2H, t,J＝6.3Hz),2.85(2H,t,J＝6.3Hz),2.31(3H,s).

Synthesis of compound a 26: compound IX-3 (122mg, 0.3 mmol) was dissolved in DMF (5 mL), and pyrrolidine (123. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 97mg of yellow solid, yield 73%. mp:251-253 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.35(1H, s),10.99(1H,s),10.00(1H,s),8.10(1H,d,J ₁ ＝1.8Hz),7.78(1H,s),7.71(1H,d,J＝2.9Hz), 7.26(1H,dd,J ₁ ＝8.2Hz,J ₂ ＝1.8Hz),6.82(1H,d,J＝2.9Hz),2.72(2H,t,J＝6.6Hz), 2.52-2.57(4H,m),2.46-2.50(2H,m),2.28(3H,s),1.70-1.76(4H,m). ¹³ C NMR(126MHz, DMSO-d ₆ )δ(ppm):169.6,169.4,137.8,128.8,128.4,125.9,125.6,125.4,121.7,120.2,119.2, 114.5,113.7,111.6,53.7(2C),51.8,35.1,23.6(2C),9.0.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₁ H ₂₄ BrN ₄ O ₂ ⁺ ,443.1077；found,443.1075.

Example 27

Synthesis of (Z) -N- (5- ((5-methoxy-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propionamide (A27)

Synthesis of intermediate V-4: compound III (1.2g, 7.79mmol) and IV-4 (1.4g, 8.56mmol) were dissolved in ethanol (10 mL), and pyrrolidine (0.83mL, 10.13mmol) was added and heated to 50 ℃ for 2 hours. After the LC-MS monitoring reaction is finished, stopping heating, cooling to room temperature, performing suction filtration, washing a filter cake with a small amount of ethanol, and performing vacuum drying to obtain 1.91g of orange yellow solid with the yield of 82%. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):14.38(1H,brs),10.97(1H,s),8.26 (1H,d,J＝3.7Hz),7.79(1H,s),7.61(1H,s),6.78-6.82(2H,m),3.79(3H,s),2.62(3H,s).

Synthesis of intermediate VI-4: compound V-4 (0.42g, 1.39mmol) was dissolved in a mixed solvent of tetrahydrofuran and methanol (27 mL/15 mL, V/V), a saturated solution (1 mL) of ammonium chloride (0.73g, 13.9mmol) was added, the mixture was heated to 50 ℃ and zinc powder (0.45g, 6.96mmol) was added in portions, and the reaction was continued for 0.5 hour. After the completion of the reaction, LC-MS monitored the reaction, heating was stopped, cooling to room temperature, concentration under reduced pressure was carried out, the residue was dissolved in ethyl acetate (100 mL) and a saturated sodium carbonate solution (100 mL), the organic layer was separated, washed with water and a saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. And (4) carrying out suction filtration, concentrating the filtrate, pulping and purifying methanol to obtain a crude product VI-4, and directly using the crude product VI-4 in the next reaction.

Synthesis of intermediate IX-4: compound VI-4 (0.31g, 1.15mmol) was dissolved in THF (20 mL), DIPEA (0.57mL, 3.45mmol) was added, the temperature was decreased to 0 deg.C, 3-chloropropionyl chloride (0.55mL, 5.75mmol) was added dropwise, and after completion of addition, reaction was carried out at room temperature for 0.5h. After the completion of the TLC monitoring reaction, concentration was performed under reduced pressure, and the residue was slurried with ethyl acetate (10 mL) and methanol (0.5 mL). Suction filtration was performed, and the filter cake was washed with ethyl acetate (1 mL) and dried in vacuo to give 0.29g of a yellow solid in 70% yield. ¹ H NMR(500 MHz,DMSO-d ₆ )δ(ppm):13.39(1H,s),10.70(1H,s),9.49(1H,s),7.66(1H,s),7.62(1H,d,J＝2.8Hz),7.46(1H,d,J＝2.0Hz),6.76(1H,d,J＝8.4Hz),6.71(1H,dd,J ₁ ＝8.4Hz,J ₂ ＝1.9Hz), 3.88(2H,t,J＝6.3Hz),3.78(3H,s),2.85(2H,t,J＝6.3Hz),2.30(3H,s).

Synthesis of compound a 27: compound IX-4 (107mg, 0.3 mmol) was dissolved in DMF (5 mL), and pyrrolidine (123. Mu.L, 1.5 mmol) was added thereto, followed by stirring the reaction at 50 ℃. After completion of the TLC monitoring reaction, the reaction solution was poured into water, extracted with ethyl acetate solution (30 mL. Times.2), and the organic layers were combined, washed with water and saturated sodium chloride solution in this order, and dried over anhydrous sodium sulfate. Suction filtration, concentration of the filtrate, addition of ethyl acetate (5 mL) to the residue, pulping for purification, suction filtration, vacuum drying to give 102mg of a yellow solid in 86% yield. mp is 254-256 ℃. ¹ H NMR(500MHz,DMSO-d ₆ )δ(ppm):13.43 (1H,s),10.68(1H,s),9.98(1H,s),7.63-7.68(2H,m),7.47(1H,s),6.74-6.80(1H,m),6.70(1H, dd,J ₁ ＝8.4Hz,J ₂ ＝1.8Hz),3.78(3H,s),2.72(2H,t,J＝6.7Hz),2.52-2.56(4H,m),2.45-2.49 (2H,m),2.28(3H,s),1.69-1.78(4H,m). ¹³ C NMR(126MHz,DMSO-d ₆ )δ(ppm):169.8,169.6, 155.3,132.9,126.9,125.4,125.1,124.5,119.0,118.0,116.6,113.0,110.3,104.5,56.1,53.7(2C), 52.0,35.1,23.6(2C),9.0.HRMS(ESI):m/z[M+H] ⁺ calcd for C ₂₂ H ₂₇ N ₄ O ₃ ⁺ ,395.2078；found, 395.2073.

Example 28

Long term stability test of Compounds

The experimental method comprises the following steps: the compounds were left open at room temperature for more than 90 days and the purity of the compounds was determined by HPLC normalization (table 1).

TABLE 1 Long-term stability test results for the example compounds

The results show that: the series of compounds have good chemical stability, and can be prepared by standing at room temperature for 90 days in the open air, and subjecting to HPLC-UV ¹ And through H-NMR spectrum analysis, the purity of each compound is not obviously reduced, and the chemical stability of each compound is obviously superior to that of the compound 10d.

Example 29

Stability test of Compound A13 in pure Water and acidic aqueous solution

The experimental method comprises the following steps: compound a13 was dissolved in pure water at pH =7 and an acidic aqueous solution at pH =2, samples were taken at different time points, and the purity of the compound was determined by HPLC normalization (table 2).

TABLE 2 stability test results of Compound A13 in pure Water and acidic aqueous solution

The results show that: the purity of compound a13 did not change significantly in aqueous solutions at pH7 and pH2 for one week, indicating that compound a13 has good stability in aqueous and strongly acidic solutions.

Example 30

Proliferation inhibitory Activity of Compound on human AML cell line

The experimental method comprises the following steps: human acute myeloid leukemia MV-4-11 and MOLM-13 are FLT3 mutant cell lines. The CCK-8 method was used to determine the in vitro antiproliferative activity of the compounds on MV-4-11: the test compounds were diluted to twice the final concentration with a 2-fold gradient of the culture medium and taken 200 μ L to 2mL EP tubes for use. An appropriate amount of cells in the logarithmic growth phase is taken and resuspended in the culture medium, the cells are added into the culture medium containing the tested compound in equal volume, the cells are evenly mixed by being inverted for 10 times, and the cells are sequentially added into a 96-well plate, wherein each well is 100 mu L. At 37 ℃ C, 5% CO ₂ After incubation for 70h, 10ul CCK-8 was added to each well and incubation continued for 2h. The microplate reader reads the OD450 absorbance value, and the experiment is repeated for three times. The data were analyzed and processed using Graphpad Prism 5 software to determine IC ₅₀ 。

The results show that: most of the compounds have good proliferation inhibition activity on human MV-4-11 and MOLM-13 cells, and maintain strong FLT3 inhibition activity (Table 3). Wherein the proliferation inhibition activity of the compounds A13, A18, A25 and A27 on two AML cells, namely MV-4-11 and MOLM-13, is superior to sunitinib.

TABLE 3 proliferation inhibitory Activity of the example Compounds on human AML cells and in vitro inhibitory Activity of FLT3

Example 31

In vitro proliferation inhibitory Activity of Compound A13 on solid tumor cells

The experimental method comprises the following steps: HCT-116 is a colon cancer cell line; MDA-MB-231 is a breast cancer cell line; HT-29Is a colon cancer cell line; hela is a cervical cancer cell line; hepG2 is a liver cancer cell line. The in vitro antiproliferative activity of compounds on solid tumor cells was determined using the MTT method: trypsinizing the cells in logarithmic growth phase, counting, resuspending the appropriate amount of cells in culture medium, 100. Mu.L per well in 96-well plates, overnight culturing, 100. Mu.L per well of 2-fold gradient diluted test compound or control medium, 5% CO at 37 ℃ in ₂ After culturing in an incubator for 44h, 20 μ L of MTT is added into each well, incubation is continued for 4h at 37 ℃, an enzyme-linked immunosorbent assay (OD 490) absorbance value is read by an enzyme-linked immunosorbent assay (ELISA) instrument, and the experiment is repeated for three times. Analyzing the processed data by adopting Graphpad Prism 5 software to obtain IC ₅₀ The value is obtained.

TABLE 4 in vitro proliferation inhibitory Activity of Compound A13 on solid tumor cells

The results show that: the compound A13 has better proliferation inhibition activity on tumor cells HCT-116, MDA-MB-231, HT-29, hela and HepG2 (table 4), and the proliferation inhibition activity is better than that of Sunitinib.

Example 32

In vitro inhibitory Activity of Compound A13 on partial tyrosine kinases

The experimental method comprises the following steps: the in vitro inhibitory activity of compound a13 on partial tyrosine kinases was determined using the method of Mobility Shift Assay: compounds were diluted 3-fold from 1. Mu.M starting concentration to 9 concentrations, then transferred 250nL to 384-well plates using Echo550, and 250nL of 100% DMSO was added to each of negative and positive control wells. 10 μ L of 2.5 fold final concentration kinase solution was added to each of the compound wells and the positive control wells; mu.L of kinase buffer was added to the negative control wells. After centrifugation at 1000rpm for 30 seconds, shaking and mixing, incubation at room temperature for 10 minutes, 15. Mu.L of a mixed solution of ATP and kinase substrate at 25/15 fold final concentration was added. Centrifuging 384-well plate at 1000rpm for 30 s, shaking for 30 s, incubating at room temperature for 30 min, adding 30 μ L of termination detection solution, stopping kinase reaction, centrifuging at 1000rpm for 30 s, shaking for mixing, reading with Caliper EZ Reader II, and transferringThe conversion rate. The data were analyzed using Graphpad Prism 5 software to determine IC ₅₀ The value is obtained.

The results show that: the compound A13 shows certain inhibitory activity on various tyrosine kinases such as FLT3, CSF-1R, PDGFR alpha, PDGFR beta, VEGFR1, VEGFR2 and the like, and especially shows strong inhibitory activity on FLT3, FLT3-ITD and FLT3-D835Y mutants (Table 5). The compound A13 has strong and selective inhibitory activity on FLT3-ITD, has over 50 times of selectivity on other receptor tyrosine kinases and over 1000 times of selectivity on c-Kit, and can reduce myelosuppressive toxicity caused by synchronous inhibition of FLT3 and c-Kit of other FLT3 inhibitors such as Midostaurin, AC220 and the like.

TABLE 5 in vitro inhibitory Activity of Compound A13 on partial tyrosine kinases

Example 33

Effect of compound A13 in vivo on inhibiting growth of MV-4-11 nude mouse transplanted tumor

The experimental method comprises the following steps: MV-4-11 cells are expanded in vitro culture, and a proper amount of cells in the logarithmic growth phase are taken and suspended in a serum-free IMDM medium and Matrigel (1) suspension, and are prepared into 5X 10 cells under the aseptic condition ⁶ mu.L of the cell suspension, 100. Mu.L of the cell suspension was inoculated subcutaneously into the anterior left axillary fossa of male Balb/c nude mice by means of a syringe. When the tumor volume grows to 100-200mm ³ In the course of treatment, animals with moderate tumor size were randomly grouped into 5 animals per group. Separately administering blank Medium (ddH) ₂ O), sunitinib (5 mg/kg/d), A13 low dose (2.5 mg/kg/d), A13 medium dose (5 mg/kg/d) and A13 high dose (10 mg/kg/d), were gavaged once a day for 3 weeks. During the administration period, body weight and tumor size of nude mice were measured daily. After the experiment, the cervical vertebra is dislocated and killed, and the tumor is taken and weighed.

The formula for Tumor Volume (TV) is: TV =1/2 × a × b ² And a represents the tumor major axis; b represents the tumor minor axis.

The results show that: compound A13 can inhibit tumor growth dose-dependently on MV-4-11 nude mouse transplantation tumor model by continuous gavage for 3 weeks, and has no effect on mouse body weight. Wherein the tumor inhibition rate under the dosage of 2.5mg/kg/d reaches 55.68 percent, is obviously superior to the tumor inhibition rate of sunitinib under the dosage of 5mg/kg/d (29.92 percent), and is superior to the tumor inhibition rate of 10d under the dosage of 10mg/kg/d (52.43 percent), the in vivo pharmacodynamic data of 10d are derived from different batches of tests, which are introduced from Eur.J.Med. Chem.2017,127 and 72-86 degrees, the compound A13 can enable the tumor to be nearly completely regressed under the dosage of 10mg/kg/d, and the tumor inhibition rate reaches 93.44 percent (Table 6).

TABLE 6 antitumor Effect of Compounds A13 and 10d on MV4-11 acute myeloid leukemia model

* P <0.05; * P <0.01; * P <0.001 (compared to solvent control).

Example 34

Effect of compound A24 in vivo inhibition of MV-4-11 nude mouse transplanted tumor growth

Experimental methods MV-4-11 cells were cultured in vitro and the transplanted tumors were inoculated in the same manner as in example 33, and nude mice with moderate tumor sizes were randomly selected and divided into 5 groups of 5 mice each. Separately administering blank Medium (ddH) ₂ O), sunitinib (5 mg/kg/d), A24 low dose (2.5 mg/kg/d), A24 medium dose (5 mg/kg/d) and A24 high dose (10 mg/kg/d), and the administration was performed by intragastric administration once a day for 2 weeks. During the administration period, body weight and tumor size of nude mice were measured. After the experiment, the cervical vertebrae were dislocated and sacrificed, and the tumor was weighed.

TABLE 7 antitumor Effect of Compound A24 on MV4-11 acute myeloid leukemia model

* P <0.05; * P <0.01; * P <0.001 (compared to solvent control).

The results show that: compound A24 was able to dose-dependently inhibit tumor growth on MV-4-11 nude mouse transplantable tumor model with no effect on mouse body weight following 2 weeks of intragastric administration. Wherein the tumor inhibition rate under the dosage of 5mg/kg/d reaches 58.42 percent, the dosage of 10mg/kg/d can completely eliminate the transplanted tumor, and the tumor inhibition rate reaches 92.16 percent (Table 7).

Example 35

Function of compound A13 in vivo for inhibiting growth of MOLM-13 nude mouse transplanted tumor

The experimental method comprises the following steps of MOLM-13 cell in-vitro culture and amplification, taking a proper amount of cells in a logarithmic growth phase, suspending the cells in a serum-free 1640 culture medium and Matrigel (1) ⁶ mu.L of the cell suspension, 100. Mu.L of the cell suspension was inoculated subcutaneously into the axilla of the anterior left limb of male Balb/c nude mice by a syringe. When the tumor volume grows to 100-200mm ³ Animals with moderate tumor size were selected and randomly divided into 6 groups. Blank medium (ddH 2O), positive drug sunitinib (20 mg/kg/d), sunitinib (40 mg/kg/d), A13 low dose (10 mg/kg/d), A13 medium dose (20 mg/kg/d) and A13 high dose (40 mg/kg/d) are respectively given, and the mixture is subjected to intragastric administration once a day for 16 days in an administration period. During the administration period, body weight and tumor size of nude mice were measured daily. After the experiment, the cervical vertebrae were dislocated and sacrificed, and the tumor was weighed.

TABLE 8 antitumor Effect of Compound A13 on MOLM-13 acute myeloid leukemia model

* P <0.05; * P <0.01; * P <0.001 (compared to solvent control).

The results show that: after continuous gavage for 16 days in a MOLM-13 nude mouse transplantation tumor model, the compound A13 can inhibit the growth of tumors dose-dependently and has no influence on the body weight of mice. Wherein the tumor inhibition rate under the dosage of 10mg/kg/d reaches 76.24 percent, the dosage of 20mg/kg/d can ensure that the transplanted tumor is nearly completely regressed, and the tumor inhibition rate reaches 97.95 percent, which is obviously superior to 80.27 percent of the tumor inhibition rate of sunitinib under the dosage of 20mg/kg/d (Table 8).

Example 36

Effect of compound A13 in vivo inhibition of HepG2 nude mouse transplanted tumor growth

The experimental method comprises the following steps: hepG2 cells were expanded by in vitro culture, and cells at the log phase of growth were counted, and an appropriate amount of cells were resuspended in a serum-free DMEM medium and Matrigel (1) suspension, and prepared to 1X 10 under sterile conditions ⁶ mu.L of the cell suspension, 100. Mu.L of the cell suspension was inoculated subcutaneously into the anterior left axillary fossa of male Balb/c nude mice by means of a syringe. When the tumor volume grows to 100-200mm ³ Animals with moderate tumor size were selected and randomly divided into 5 groups. Separately administering blank Medium (ddH) ₂ O), sunitinib 40mg/kg, A13 low dose (20 mg/kg/d), A13 medium dose (40 mg/kg/d) and A13 high dose (80 mg/kg/d), and the administration period is 2 weeks after the administration of the composition by intragastric administration once a day. During the administration period, the body weight and tumor size of nude mice were measured daily. After the experiment, the cervical vertebrae were dislocated and sacrificed, and the tumor was weighed.

The results show that: compound a13 was able to dose-dependently inhibit tumor growth with no effect on mouse body weight by continuous gavage administration for 2 weeks on HepG2 nude mouse graft tumor model. Wherein the tumor inhibition rate under the dosage of 20mg/kg/d reaches 54.94 percent, which is equivalent to the tumor inhibition rate (55.11 percent) of sunitinib under the dosage of 40mg/kg/d, and the tumor inhibition rate under the dosage of 80mg/kg/d reaches 87.70 percent.

TABLE 9 tumor-inhibiting effect of Compound A13 on HepG2 liver cancer model

* P <0.05; * P <0.01; * P <0.001 (compared to solvent control).

Example 37

In vivo efficacy of Compound A13 on HT-29 Colon cancer nude mouse transplantable tumors

The experimental method comprises the following steps: HT-29 cells are cultured and amplified in vitro, the cells in the logarithmic phase of growth are taken, counted, and an appropriate amount of cells are taken to be re-suspended in a serum-free 1640 medium and Matrigel (1) suspension to be prepared into 1 × 10 ⁶ mu.L of the cell suspension, 100. Mu.L of the cell suspension was inoculated subcutaneously into the axilla of the anterior left limb of male Balb/c nude mice by using a syringe, respectively. Periodically observing the animal and movingGrowing the implanted tumor; regularly observing the growth of animals and transplanted tumors; when the tumor volume grows to 100-200mm ³ At this time, animals with moderately large tumors were selected and randomized into 5 groups. The animals were dissected for grouping, dose, mode of administration and end of experiment as in example 36.

TABLE 10 antitumor Effect of Compound A13 on HT-29 Colon cancer model

* P <0.05; * P <0.01; * P <0.001 (compared to solvent control).

The results show that: compound A13 was able to dose-dependently inhibit tumor growth on HT-29 nude mouse transplantable tumor model by continuous gavage for 2 weeks, with no effect on mouse body weight. Wherein the tumor inhibition rate under the dosage of 80mg/kg/d reaches 78.22 percent.

Claims (4)

1. A 4-methylpyrrole substituted indolone derivative or a pharmaceutically acceptable salt thereof, characterized by being selected from the group consisting of:

   (S,Z)-N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-2-carboxamide hydrochloride,

   (R,Z)-N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) pyrrolidine-3-carboxamide hydrochloride,

   (Z)-N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propanamide,

   Z)-N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (4-methylpiperazin-1-yl) propanamide,

   (Z) -3- (diethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide,

   (Z) -3- (dimethylamino) substituted benzeneN- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide,

   (Z) -3- (ethylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide,

   (Z)-N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (propylamino) propanamide,

   (Z) -3- (cyclopentylamino) -N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) propanamide,

   (Z)-N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -6- (trifluoromethyl) nicotinamide,

   (Z)-N- (5- ((5-fluoro-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -4-methylpiperazine-1-carboxamide,

   (Z)-N- (5- ((5-bromo-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propanamide,

   (Z)-N- (5- ((5-methoxy-2-oxoindol-3-ylidene) methyl) -4-methyl-1H-pyrrol-3-yl) -3- (pyrrolidin-1-yl) propanamide.
2. 2. Use of a 4-methylpyrrole-substituted indolone derivative or a pharmaceutically acceptable salt thereof as claimed in claim 1 for the preparation of FLT3 inhibitors.
3. 3. Use of a 4-methylpyrrole-substituted indolone derivative or a pharmaceutically acceptable salt thereof as described in claim 1 for the preparation of a medicament for the treatment and/or prevention of diseases associated with abnormal FLT3 expression.
4. 4. Use of a 4-methylpyrrole-substituted indolone derivative or a pharmaceutically acceptable salt thereof as claimed in claim 1 for the preparation of a medicament for the treatment of acute myeloid leukemia.