Indole acrylic acid amide derivative as well as preparation method and application thereof

Abstract

The invention belongs to the field of medicinal chemistry, and relates to an indole acrylic acid amide derivative as well as a preparation method and application thereof. Functional activity screening of G protein dependent signaling pathways is carried out on all synthesized compounds, and it is found that the new derivatives can be used as allosteric antagonism regulators of beta2-adrenergic receptors. Trans-indole acrylic acid with different substituent groups is used as a raw material, amide coupling is performed to obtain the novel indole acrylic acid amide derivative, and R1 is a hydrogen atom, a halogen atom, a methyl group or a methoxyl group; r2 is any one of phenyl, m-bromophenyl, m-chlorphenyl, m-fluorophenyl, m-cyanophenyl, m-methoxyphenyl, cyclohexyl and piperidyl. A biological activity test result shows that the indole acrylic acid amide derivative disclosed by the invention has relatively good antagonistic activity on beta2AR and can be used for carrying out negative allosteric regulation on the functional activity of isoproterenol.

Description

Technical Field

[0001] The invention belongs to the field of medicinal chemistry, and particularly relates to an indole acrylic acid amide compound and a synthesis method thereof, and an application of the indole acrylic acid amide compound as a β2-adrenaline receptor allosteric antagonist modulator. Background Art

[0002] G protein-coupled receptors (GPCRs) are the largest family of cell signal transduction receptors in the human body and the most important drug target. They are involved in almost all life activities and are responsible for regulating the response of cells to light, odor, hormones, neurotransmitters, chemokines, etc. They are of great significance to maintaining life and health and are widely used in biological cell signal transduction research and drug screening and development. At present, there are about 500 drugs targeting GPCRs, accounting for 34% of FDA-approved drugs. GPCR research has won 10 Nobel Prizes, which also indirectly illustrates its importance. GPCRs have conservative structural features, namely seven transmembrane helices (7TMs). The transmembrane domain composed of these seven helices divides the receptor into an extracellular N-terminus, an intracellular C-terminus, three extracellular loops, and three intracellular loops. The extracellular region of GPCR undergoes conformational changes after binding to agonist signal molecules (such as odors, hormones, etc.), which in turn triggers the movement of transmembrane helices. The activation mechanism of GPCR is the only way to understand its physiological functions and the causes of related diseases. It is also the basis for precise drug development and has always been the core content and frontier hotspot of research in this field. The activation of GPCR refers to the recruitment of downstream effector proteins triggered by agonist binding. The essence of GPCR activation is the process of mutual coupling and allosteric communication between extracellular agonist binding and intracellular downstream effector protein recruitment, that is, the conformational transition of the receptor from the inactive state to the activated state.

[0003] β2-adrenergic receptor (β2AR) is a typical GPCR and has long been used as a model system for studying GPCR regulatory mechanisms. β2-receptor blockers, or orthosteric antagonists of β2AR, are mainstay cardiovascular drugs used to treat a variety of diseases. Adrenergic receptors are a type A subfamily of GPCRs that play an important role in the sympathetic nervous system.

[0004] In the human body, there are nine subtypes of adrenergic receptors, and their sequences and structures are relatively conserved. Traditionally, drug development for GPCRs has mainly focused on the orthosteric binding site (i.e., the location where the endogenous ligand of the receptor binds). The high conservation of this site between different subtypes has brought great challenges to the development of selective drugs. Summary of the invention

[0005] The purpose of the present invention is to prepare new indole acrylic acid amide derivatives to develop target compounds with high biological activity, stable chemical structure, good water solubility and novel skeleton as new β2AR allosteric antagonists.

[0006] To achieve the above purpose, the technical solution adopted by the present invention is as follows:

[0007] The present invention provides an indole acrylic acid amide derivative, the structure of the indole acrylic acid amide derivative is shown in Formula 1:

[0008]

[0009] R1=H,F,Cl,Br,CN,Me,OCH3;

[0010]

[0011] The present invention also provides a preparation method of indole acrylic acid amide derivatives, which comprises: dissolving trans-indole acrylic acid with different substituents in N,N-dimethylformamide (DMF), adding 1-hydroxy-7-azabenzotriazole (HOAt), stirring at room temperature for 10 minutes, adding N-methylmorpholine and aniline, aliphatic amine or heterocyclic amine with different substituents at 0°C, and then adding 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI), and then reacting at room temperature for several hours to obtain the indole acrylic acid amide derivatives.

[0012] Furthermore, the trans-indole acrylic acid with different substituents includes (E3)-3-(4-bromo-1H-indol-3-yl)acrylic acid, (E3)-3-(5-bromo-1H-indol-3-yl)acrylic acid, and (E3)-3-(6-bromo-1H-indol-3-yl)acrylic acid.

[0013] Furthermore, the anilines with different substituents include m-bromoaniline, m-chloroaniline, and m-fluoroaniline; the aliphatic amines with different substituents include cyclohexylamine; and the heterocyclic amines with different substituents include piperidine and piperazine.

[0014] Furthermore, the preparation method specifically comprises: dissolving 1 mmol of trans-indolepropene with different substituents in N,N-dimethylformamide, adding 1.20 mmol of N-hydroxy-7-azabenzotriazole, 1.20 mmol of aniline, aliphatic amine or heterocyclic amine with different substituents and 0.70 mmol of N-methylmorpholine respectively under ice bath conditions, reacting for 10 minutes, then adding 1.20 mmol of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, and then reacting at room temperature for 4 hours; after the reaction is completed, the reaction solution is diluted with ethyl acetate, washed with water, and then dried with anhydrous sodium sulfate; after rotary evaporation to remove the solvent, the obtained solid crude product is purified by silica gel column chromatography, eluted with an eluent, and the product after column chromatography is concentrated, and then filtered with dichloromethane to obtain the new indoleacrylic acid amide derivative.

[0015] Furthermore, the eluent is prepared by petroleum ether and ethyl acetate in a volume ratio of 1:1.

[0016] The present invention also provides the use of the above-mentioned indole acrylic acid amide derivatives as β2 adrenergic receptor (β2AR) allosteric antagonists.

[0017] Furthermore, the indole acrylic acid amide derivatives are used to exert an allosteric antagonistic effect on the G-protein signaling pathway of β2AR, and negatively regulate the agonistic effect of the endogenous agonist isoproterenol (ISO) on β2AR.

[0018] Compared with the prior art, the present invention has at least the following beneficial effects:

[0019] The present invention provides a new derivative of indole acrylic acid amide and a preparation method thereof. By screening the functional activity of all synthesized compounds for G protein-dependent signaling pathways, the new derivative can be used as an allosteric antagonist modulator (NAM) of β2-adrenergic receptors. The present invention uses trans-indole acrylic acid with different substituents as raw materials, and amide coupling to obtain the following new indole acrylic acid amide derivatives, which have the characteristics of high biological activity, stable chemical structure, good water solubility, and novel skeleton, wherein R1 is a hydrogen atom, a halogen atom, a methyl or a methoxyl group; R2 is any one of phenyl, m-bromophenyl, m-chlorophenyl, m-fluorophenyl, m-cyanophenyl, m-methoxyphenyl, cyclohexyl, and piperidinyl. The results of biological activity tests show that the indole acrylic acid amide derivatives of the present invention have good antagonistic activity to β2AR, and can negatively allosterically regulate the functional activity of isoproterenol (ISO). The indole acrylic acid amide derivatives can be used as allosteric antagonists of β2AR, providing a useful reference for the structure-activity relationship and drugability research of such compounds. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1For the design of early pyrazole compounds;

[0021] Figure 2 Design of indole acrylic acid amide target compounds;

[0022] Figure 3 For the synthesis of indole acrylic acid amide compounds L1-L27;

[0023] Figure 4 A method for synthesizing a type M compound;

[0024] Figure 5 Figure 2 is a graph of the ISO dose-response curve mediated by . DETAILED DESCRIPTION

[0025] In 1961, Jacques Monod of the Pasteur Institute in France, Jacob and Jean-Pierre Changeux first proposed the concept of allostery. Allosteric sites (other regions that are different from orthosteric binding sites in space and topology) have relatively large degrees of freedom, low evolutionary pressure, and relatively high mutation frequency. Compared with orthosteric drugs targeting active sites, allosteric modulators are more selective and less toxic. Allosteric modulators bind outside the orthosteric ligand pocket of the receptor. Due to the relatively low conservation of their binding position, they may have better subtype selectivity. Its discovery provides a new idea for obtaining highly selective drugs. In addition, orthosteric modulators and allosteric modulators exert their efficacy based on different mechanisms of action, which makes it possible for allosteric modulators to overcome the acquired resistance of orthosteric drugs during treatment and have better selectivity and safety.

[0026] In 2017, the applicant collaborated with scientists from Duke University in the United States to report the first small molecule negative allosteric modulator compound 15 (Cmpd-15, as shown in Formula 2) of β2AR (Ahn S, et al. Proc. Natl. Acad. Sci. USA, 2017, 114: 1708-1713; Liu X, et al. Nature, 2017, 548: 480-484). However, since Cmpd-15 is a dipeptide compound with poor water solubility and low biological activity, its structure is also relatively unstable, which may affect its drugability. Therefore, this project uses Cmpd-15 as the lead compound and uses the skeleton transition strategy, structural simplification and bioisostere concept for drug design. The synthesized new compound was detected by GloSensor TMcAMPAccumulation assay is used to screen the biological activity of the classical signaling pathway (G-protein signaling) (Binkowski BF et al. ACS Chem Biol. 2011; 6(11): 1193-1197). It is expected to obtain a series of new indoleamide derivatives with stable and simplified structures, novel skeletons, enhanced allosteric activity, improved water solubility and metabolic stability as allosteric antagonists of β2AR.

[0027]

[0028] Heterocycles such as pyrazoles and indoles are common drug skeletons. Therefore, the inventors replaced the peptide structure of Cmpd-15 with a pyrazole skeleton in the early stage, such as Figure 1 As shown, keeping the (S)-2-amino-3-(3-bromophenyl)-N-methylpropionamide on the right unchanged, a series of pyrazole derivatives were designed and synthesized (China Invention Patent Publication No.: CN115745891A).

[0029] According to the inventor's previous research, the results of cAMP accumulation experiments show that most pyrazole amide derivatives have significantly better allosteric antagonism to β2AR than their lead compound Cmpd-15, and the water solubility of the new derivative compounds is significantly improved compared with Cmpd-15. However, the phenylalanine part on the right side of Cmpd-15 has the disadvantage of large steric hindrance, which may affect the drugability of the compound (China Invention Patent Publication No.: CN115894373A). For this reason, it is necessary to optimize its structure. Indole is an aromatic heterocyclic organic compound containing a six-membered benzene ring and a five-membered nitrogen-containing pyrrole ring, so it is also called benzopyrrole, which is an extremely important part in drug design. It is widely present in nature and exhibits a broad spectrum of pharmacological activities, such as antidepressant, anticancer, and anti-inflammatory effects. The present invention uses the principles of skeleton transition, structural simplification and bioisostere to replace the pyrazole skeleton with the indole acrylic acid skeleton, and replaces the phenylalanine part on the right side of Cmpd-15 with substituted aniline, aliphatic amine and heterocyclic amine to design and synthesize a series of indole acrylic acid amide derivatives, such as Figure 2 As shown, the biological activity of the new derivatives on β2AR was studied.

[0030] In view of this, the present invention uses trans-indole acrylic acid 1 with different substituents as raw materials, and carries out amine acyl coupling with substituted aniline, aliphatic amine and heterocycle 2 respectively to obtain a series of new indole acrylic acid amide derivatives as follows: Figure 3 shown.

[0031] The specific synthesis method steps of the indoleamide derivatives provided by the present invention are as follows:

[0032] Compound 1 (1 mmol) was dissolved in N, N-dimethylformamide (DMF), and N-hydroxy-7-azabenzotriazole (HOAT) (1.20 mmol), compound 2 (1.20 mmol) and N-methylmorpholine (NMM) (0.70 mmol) were added under ice bath conditions. After reacting for 10 min, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI) (1.20 mmol) was added, and then reacted at room temperature for 4 h. After the reaction was completed by TLC monitoring, the reaction solution was diluted with ethyl acetate (50 mL), washed with water (50 mL × 3), and then dried over anhydrous sodium sulfate. After the solvent was removed by rotary evaporation, the obtained solid crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 1:1], the product after column chromatography was concentrated, and then filtered with an appropriate amount of dichloromethane to obtain a new derivative of indole acrylic acid amide (hereinafter referred to as target compound L), as shown in Table 1.

[0033] Among them, compound 1 is trans-indolepropene with different substituents;

[0034] Compound 2 is aniline, aliphatic amine or heterocyclic amine with different substituents.

[0035] Table 1 Structures of new derivatives of indole acrylic acid amide

[0036]

[0037]

[0038]

[0039]

[0040] In order to study the effect of double bonds in indoleacrylic acid amide on the biological activity of derivatives, the inventors synthesized compounds M1 and M2 without double bonds, such as Figure 4 As shown, the specific synthesis method is the same as that of L series derivatives.

[0041] The present invention is described in detail below in conjunction with specific implementation modes.

[0042] The preparation method of the indole acrylic acid amide derivatives provided by the present invention is specifically as follows.

[0043] Embodiment 1:

[0044] Preparation of (E3)-(1H-indol-3-yl)-N-phenylacrylamide (L1)

[0045] Trans-1H-3-indole-acrylic acid (250 mg, 1.34 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAt (218 mg, 1.60 mmol) was added and reacted for 10 min, and then aniline (146 μL, 1.20 mmol) and NMM (104 μL, 0.94 mmol) were added under ice bath conditions and reacted for 10 min, and then EDCI (307 mg, 1.60 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, it was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 15:1], and the crude product of the concentrated column chromatography was filtered with dichloromethane to obtain the target compound L1 as a pale yellow solid powder with a yield of 49%. 1 HNMR (400MHz, DMSO-d6): δ11.69(s,1H),10.04(s,1H),7.99(d,J=7.1Hz,1H),7.88–7.76(m,2H),7.73(d,J=8.0Hz, 2H),7.49(d,J=7.2Hz,1H),7.33(t,J=7.3Hz,2H),7.26–7.15(m,2H),7.03(t,J=7.4Hz,1H),6.85(d,J=15.7Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ165.1,139.9,137.6,134.9,131.2,128.9,124.9,122. 8,122.5,120.6,120.1,118.9,116.0,112.5,112.3.HRMS(ESI,m / z):Calcd.for C 17 H 14 N2O[M+Na] + 285.0998; found:285.1003.

[0046] Embodiment 2:

[0047] Preparation of (E3)-N-(3-bromophenyl)-3-(1H-indol-3-yl)acrylamide (L2)

[0048] Trans-1H-3-indole-acrylic acid (300 mg, 1.60 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAt (261 mg, 1.92 mmol) was added and reacted for 10 min, then m-bromoaniline (0.35 ml, 3.20 mmol) and NMM (0.12 ml, 1.12 mmol) were added under ice bath conditions and reacted for 10 min, and then EDCI (368 mg, 1.20 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, it was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation, and the crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 1:1], and the crude product of the concentrated column chromatography was filtered with dichloromethane to obtain the target compound L2 as a yellow solid with a yield of 89%. 1 H NMR (400MHz, DMSO-d6): δ11.73(s,1H),10.20(s,1H),8.16(s,1H),8.00–7.91(m,1H),7.88(s,1H),7.81(d,J=15.6 Hz,1H),7.55(d,J=8.2Hz,1H),7.51-7.44(m,1H),7.29(t,J=8.0Hz,1H),7.25-7.18(m,3H),6.79(d,J=15.7Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ165.4,141.5,137.6,135.6,131.7,130.9,125.4,124.9,122.5,121.8,121.2,120.7,120.1,117.6,115.4,112.6,112.2.

[0049] Embodiment 3:

[0050] Preparation of (E3)-N-(3-chlorophenyl)-3-(1H-indol-3-yl)acrylamide (L3)

[0051] Trans-1H-3-indole-acrylic acid (250 mg, 1.34 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAt (218 mg, 1.60 mmol) was added, and the mixture was reacted for 10 min. Then, m-chloroaniline (0.17 ml, 1.60 mmol) and NMM (104 μL, 0.94 mmol) were added under ice bath conditions and reacted for 10 min. Then, EDCI (307 mg, 1.60 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 15:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L3 as a yellow solid with a yield of 47%. 1 H NMR (300MHz, DMSO-d6): δ11.70(s,1H),10.21(s,1H),7.97(d,J=10.6Hz,2H),7.83(t,J=10.8Hz,2H),7.50 (t,J=8.3Hz,2H),7.36(t,J=8.0Hz,1H),7.29–7.18(m,2H),7.09(d,J=8.0Hz,1H),6.80(d,J=15.7Hz,1H). 13 C NMR (75MHz, DMSO-d6): δ165.4,141.4,137.6,135.6,133.2,131.6,130.5,124.9,122.5,122.4,120.7,120.1,118.4,117.3,115.5,112.5,112.2.

[0052] Embodiment 4:

[0053] Preparation of (E3)-N-(3-fluorophenyl)-3-(1H-indol-3-yl)acrylamide (L4)

[0054] Trans-1H-3-indole-acrylic acid (250 mg, 1.34 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAt (218 mg, 1.60 mmol) was added, and the mixture was reacted for 10 min. 3-fluoroaniline (0.15 ml, 1.60 mmol) and NMM (100 μL, 0.94 mmol) were added under ice bath conditions and reacted for 10 min. EDCI (307 mg, 1.60 mmol) was then added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 15:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L4 as a yellow solid with a yield of 73%. 1 HNMR (400MHz, DMSO-d6): δ11.75(s,1H),10.31(s,1H),7.99(d,J=7.1Hz,1H),7.88(s,1H),7 .85-7.64(m,2H),7.50(d,J=7.3Hz,1H),7.38(s,2H),7.29–7.18(m,2H),6.93–6.79(m,2H). 13 C NMR (100MHz, DMSO-d6): δ165.4,141.7,141.6,137.6,135.5,131.6,130.5,130.4,124.9,122.5,1 20.7,120.1,115.5,114.7,112.5,112.2,109.3,109.1,105.8,105.6.HRMS(ESI,m / z):Calcd.for C 17 H 13 FN2O[M+Na] + 303.0904; found:303.0908.

[0055] Embodiment 5:

[0056] Preparation of (E3)-3-(1H-indol-3-yl)-N-(m-tolyl) acrylamide (L5)

[0057] Trans-1H-3-indole-acrylic acid (250 mg, 1.34 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAt (218 mg, 1.60 mmol) was added and reacted for 10 min, then m-methylaniline (172 μL, 1.60 mmol) and NMM (103.90 μL, 0.94 mmol) were added under ice bath conditions and reacted for 10 min, and then EDCI (307 mg, 1.60 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, it was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 1:1], and the crude product of the concentrated column chromatography was filtered with dichloromethane to obtain the target compound L5 as a light yellow solid with a yield of 96%. 1 H NMR (400MHz, DMSO-d6): δ11.68(s,1H),9.96(s,1H),7.99(d,J=7.4Hz,1H),7.85(s,1H),7.80(d,J=15.6Hz ,1H),7.58(s,1H),7.56–7.46(m,2H),7.28–7.16(m,3H),6.87(d,J=5.6Hz,1H),6.84(s,1H),2.30(s,3H). 13 C NMR (100MHz, DMSO-d6): δ165.2,165.1,139.8,139.7,138.1,137.6,137.4,134.8,131.2,131.1,128.8,1 25.0,124.9,123.7,122.5,120.7,120.2,119.5,119.4,116.2,116.1,112.5,112.5,112.3,112.3,21.4.

[0058] Embodiment 6:

[0059] Preparation of (E3)-N-(3-cyanophenyl)-3-(1H-indol-3-yl)acrylamide (L6)

[0060] Trans-1H-3-indole-acrylic acid (250 mg, 1.34 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAt (218 mg, 1.60 mmol) was added, and the mixture was reacted for 10 min. 3-aminobenzonitrile (0.29 ml, 2.67 mmol) and NMM (100 μL, 0.94 mmol) were added under ice bath conditions and reacted for 10 min. EDCI (307 mg, 1.60 mmol) was then added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 15:1], and the crude product of the concentrated column chromatography was filtered with dichloromethane to obtain the target compound L6 as a yellow solid with a yield of 37%. 1 HNMR (400MHz, DMSO-d6): δ11.72(s,1H),10.35(s,1H),8.27(s,1H),7.98–7.92(m,1H),7.89(d,J=2.8Hz,1 H),7.88–7.80(m,2H),7.55(t,J=7.8Hz,1H),7.52–7.46(m,2H),7.29-7.18(m,2H),6.79(d,J=15.6Hz,1H). 13 C NMR(100MHz,DMSO-d6)δ165.6,140.6,137.6,135.9,131.8,130.3,126.3,124.9,123.4,1 22.5,121.5,120.7,120.0,118.9,115.1,112.5,112.1,111.6.HRMS(ESI,m / z):Calcd.for C 18 H 13 N3O[M+Na] + 310.0951 found:310.0955.

[0061] Embodiment 7:

[0062] Preparation of (E3)-3-(1H-indol-3-yl)-N-(3-methoxyphenyl)acrylamide (L7)

[0063] Trans-1H-3-indole-acrylic acid (250 mg, 1.34 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAT (218 mg, 1.60 mmol) was added, and the mixture was reacted for 10 min. 3-methoxyaniline (0.18 ml, 1.60 mmol) and NMM (100 μL, 0.94 mmol) were added under ice bath conditions and reacted for 10 min. EDCI (307 mg, 1.60 mmol) was then added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 10:1], and the crude product of the concentrated column chromatography was filtered with dichloromethane to obtain the target compound L7 as a pale yellow solid with a yield of 72%. 1H NMR (400MHz, DMSO-d6): δ11.70(s,1H),10.06(s,1H),7.99(d,J=7.5Hz,1H),7.90–7.77(m,2H) ,7.55–7.43(m,2H),7.32–7.16(m,4H),6.85(d,J=15.7Hz,1H),6.66–6.58(m,1H),3.75(s,3H). 13 C NMR (100MHz, DMSO-d6): δ165.1,159.6,141.0,137.5,134.9,131.3,129.6,124.9,122.4, 120.6,120.1,116.0,112.5,112.2,111.3,108.2,104.7,79.2.HRMS(ESI,m / z):Calcd.for C 18 H 16 N2O2[M+Na] + 315.1104; found:315.1112.

[0064] Embodiment 8:

[0065] Preparation of (E3)-3-(4-bromo-1H-indol-3-yl)-N-phenylacrylamide (L8)

[0066] 4-Bromo-1H-indole-3-carboxaldehyde (770 mg, 3.44 mmol) was dissolved in pyridine in a 50 ml round-bottom flask, malonic acid (358 mg, 3.44 mmol) and piperidine were added as catalysts, and the mixture was reacted at room temperature for 10 h under nitrogen protection. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with the eluent [ V (petroleum ether): V (ethyl acetate) = 10: 1], concentrated column chromatography to obtain a crude product of 280.20 mg, then take the acid 100 mg and dissolve it in DMF in a 25mL round-bottom flask, add HOAt (61 mg, 0.45 mmol) and react for 10 min, then add aniline (34 μL, 0.38 mmol) and NMM (30 μL, 0.26 mmol) under ice bath conditions to react for 10 min, then add EDCI (86 mg, 0.45 mmol) and react at room temperature for 4 h. After the reaction is completed as monitored by TLC, it is extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase is collected, dried over anhydrous sodium sulfate, and then the solvent is removed by rotary evaporation, and the crude product is purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 8: 1], the crude product of the concentrated column chromatography is then filtered with dichloromethane to obtain the target compound L8 as a yellow solid 92.60 mg, with a yield of 93%. 1 HNMR (400MHz, DMSO-d6): δ11.95(s,1H),10.07(s,1H),7.93(s,1H),7.87(d,J=15.2Hz,1H),7.74(d,J=8.0Hz,2H),7.33( t,J=8.1Hz,3H),7.18(td,J=8.0,5.2Hz,1H),7.05(t,J=7.4Hz,1H),6.94(dd,J=11.8,7.8Hz,1H),6.73(d,J=15.6Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.5,139.8,138.0,133.6,128.8,126.7,124.7,123. 9,123.0,123.0,119.1,117.2,112.9,112.5,112.1.HRMS(ESI,m / z):Calcd.for C 17 H 13 BrN2O[M+Na] + 363.0103; found:363.0109.

[0067] Embodiment 9:

[0068] Preparation of (E3)-3-(4-bromo-1H-indol-3-yl)-N-(3-fluorophenyl)acrylamide (L9)

[0069] 4-Bromo-1H-indole-3-carboxaldehyde (770 mg, 3.44 mmol) was dissolved in a 50 ml round-bottom flask with pyridine, and malonic acid (358 mg, 3.44 mmol) and piperidine were added as catalysts. The mixture was reacted at room temperature for 10 h under nitrogen protection. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V(petroleum ether):V(ethyl acetate)] =10:1], concentrated by column chromatography to obtain a crude product (E3)-3-(4-bromo-1H-indol-3-yl)acrylic acid 280.20 mg, and then 91 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask, HOAt (56 mg, 0.41 mmol) was added, and the mixture was reacted for 10 min. Then 3-fluoroaniline (33 μL, 0.34 mmol) and NMM (26 μL, 0.24 mmol) were added in an ice bath and the mixture was reacted for 10 min. Then EDCI (79 mg, 0.41 mmol) was added and the mixture was reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V (petroleum ether): V (ethyl acetate) = 8:1]. The crude product from the concentrated column chromatography was filtered with dichloromethane to obtain 88 mg of the target compound L9 as a yellow solid with a yield of 97%. 1 H NMR (400MHz, Methanol-d4): δ8.82(d,J=15.6Hz,1H),7.88(s,1H),7.66(dt,J=11.4,2.2Hz,1H),7.42(d,J=4.4 Hz,1H),7.36–7.31(m,2H),7.31–7.27(m,1H),7.04(t,J=7.9Hz,1H),6.85-6.76(m,1H),6.54(d,J=15.5Hz,1H). 13C NMR(100MHz,Methanol-d4)δ167.9,165.5,163.1,142.2,142.1,139.6,137.3,131.2,131.1,127.2,12 6.1,125.8,124.0,116.4,114.7,114.6,112.5,111.2,111.0,108.1,107.8.HRMS(ESI,m / z):Calcd.for C 17 H 12 BrFN2O[M+Na] + 381.0009; found:381.0014.

[0070] Embodiment 10:

[0071] (E3)-3-(4-bromo-1H-indol-3-yl)-N-cyclohexyl acrylamide (L 10 )

[0072] Same as Example 9, (E3)-3-(4-bromo-1H-indol-3-yl)acrylic acid was first generated, and 280 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask, HOAt (172 mg, 1.26 mmol) was added, and the reaction was continued for 10 min. Cyclohexylamine (120 μL, 1.05 mmol) and NMM (82 μL, 0.74 mmol) were added under ice bath conditions and the reaction was continued for 10 min. EDCI (242 mg, 1.26 mmol) was then added and the reaction was continued at room temperature for 4 h. After the reaction was completed as monitored by TLC, DMF was removed by extraction with ethyl acetate (50 mL) and water (50 mL×3) for 3 times. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The obtained crude product was purified by silica gel column chromatography, and the eluent was [V(dichloromethane V(methanol)=30:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 10 199 mg of yellow solid powder, yield 75%. 1 HNMR (400MHz, DMSO-d6): δ11.90(s,1H),8.36(s,1H),7.84(d,J=27.8Hz,2H),7.38(d, J=73.4Hz,2H),7.04(s,1H),6.39(s,1H),3.66(s,1H),2.14–1.45(m,5H),1.20(s,6H). 13C NMR (100MHz, DMSO-d6): δ164.7,137.9,131.6,125.9,124.4,123.9,122.8,118.0,112.9,112.7,112.0,47.5,32.7,25.3,24.7.HRMS(ESI,m / z):Calcd.for C 17 H 19 BrN2O[M+Na] + 369.0573; found:369.0570.

[0073] Embodiment 11:

[0074] (E3)-3-(4-bromo-1H-indol-3-yl)-1-(piperidin-1-yl)prop-2-en-1-one (L 11 )

[0075] As in Example 10, (E3)-3-(4-bromo-1H-indol-3-yl)acrylic acid was first generated, and 300 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask. HOAt (155 mg, 1.36 mmol) was added and reacted for 10 min. Cyclohexylamine (112 μL, 1.13 mmol) and NMM (88 μL, 0.79 mmol) were added under ice bath conditions and reacted for 10 min. EDCI (260 mg, 1.36 mmol) was then added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, it was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, and the eluent was [V (dichloromethane V (methanol) = 40:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 11 145 mg of yellow solid powder, yield 55%. 1 HNMR (400MHz, DMSO-d6): δ12.00(s,1H),8.49(d,J=15.2Hz,1H),8.24(s,1H),7.46(d,J=8.1Hz,1H),7 .29(d,J=7.6Hz,1H),7.10–7.03(m,1H),7.01(s,1H),3.58(s,4H),1.60(d,J=5.8Hz,2H),1.49(s,4H). 13C NMR(100MHz, DMSO-d6): δ164.9,137.8,134.4,127.2,124.5,123.8,122.7,112.9,112.8,112.7,111.9,24.3.HRMS(ESI,m / z):Calcd.for C 16 H 17 BrN2O[M+Na] + 355.0416; found:355.0414.

[0076] Embodiment 12:

[0077] (E3)-3-(4-chloro-1H-indol-3-yl)-N-phenylacrylamide (L 12 )

[0078] 4-Chloro-1H-indole-3-carboxaldehyde (970 mg, 5.40 mmol) was dissolved in a 50 ml round-bottom flask with pyridine, and malonic acid (562 mg, 5.40 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, it was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methanol)] )=45:1], concentrated column chromatography to obtain a crude product (E3)-3-(4-chloro-1H-indol-3-yl) acrylic acid 530mg, then take the acid 147mg and dissolve it in DMF in a 25mL round-bottom flask, add HOAt (108mg, 0.79mmol) and react for 10min, then add aniline (60μL, 0.66mmol) and NMM (50μL, 0.46mmol) in an ice bath and react for 10min, then add EDCI (151mg, 0.79mmol) and react at room temperature for 4h. After the reaction is completed as monitored by TLC, it is extracted 3 times with ethyl acetate (50mL) and water (50mL×3) to remove DMF, the organic phase is collected, dried over anhydrous sodium sulfate and then rotary evaporated to remove the solvent, the obtained crude product is purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate)=6:1], the crude product of the concentrated column chromatography is then filtered with dichloromethane to obtain the target compound L 12 135.60 mg of yellow solid, yield 92%. 1H NMR (400MHz, DMSO-d6): δ12.02(s,1H),10.07(s,1H),8.44(d,J=15.5Hz,1H),8.01(s,1H),7.73(d,J=7.9Hz,2 H),7.48-7.41(m,1H),7.32(t,J=7.8Hz,2H),7.19–7.09(m,2H),7.03(t,J=7.4Hz,1H),6.63(d,J=15.5Hz,1H). 13 CNMR (100MHz, DMSO-d6): δ164.5,139.8,138.2,133.8,128.8,126.6,124.8,123 .0,122.7,122.6,121.3,119.2,117.4,112.0,111.7.HRMS(ESI,m / z):Calcd.for C 17 H 13 ClN2O[M+Na] + 319.0608; found:319.0610.

[0079] Embodiment 13:

[0080] (E3)-N-(3-bromophenyl)-3-(4-chloro-1H-indol-3-yl)acrylamide (L 13 )

[0081] Same as Example 12 above, (E3)-3-(4-chloro-1H-indol-3-yl)acrylic acid was first generated, and 200 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask, HOAt (147 mg, 1.08 mmol) was added, and the mixture was reacted for 10 min. Then, m-bromoaniline (98 μL, 0.90 mmol) and NMM (70 μL, 0.63 mmol) were added under ice bath conditions and reacted for 10 min. Then, EDCI (207 mg, 1.08 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, and the eluent was [V(petroleum ether: V(ethyl acetate)=8:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 13 Yellow solid powder 75.30 mg, yield 22%. 1HNMR (400MHz, DMSO-d6): δ12.03(s,1H),10.22(s,1H),8.44(d,J=15.5Hz,1H),8.12(s,1H),8.03(s,1H),7.57(d,J=8.4Hz,1H ),7.45(dd,J=6.6,2.5Hz,1H),7.28(t,J=8.0Hz,1H),7.22(dt,J=8.2,1.5Hz,1H),7.19–7.11(m,2H),6.56(d,J=15.6Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.7,141.3,138.1,134.5,130.7,126.9,125.5,124.7,122.7,122.5,121.7,121.4,121.4,117.8,116.7,111.9,111.7. Embodiment 14:

[0082] (E3)-3-(4-chloro-1H-indol-3-yl)-N-(3-fluorophenyl)acrylamide (L 14 )

[0083] As in Example 12 above, (E3)-3-(4-chloro-1H-indol-3-yl)acrylic acid was first generated, and 131 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask. HOAt (97 mg, 0.71 mmol) was added and reacted for 10 min. 3-fluoroaniline (57 μL, 0.59 mmol) and NMM (46 μL, 0.41 mmol) were added under ice bath conditions and reacted for 10 min. EDCI (136 mg, 0.71 mmol) was then added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, it was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 8:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 14 Yellow solid powder: 67.30 mg, yield: 53%. 1H NMR (400MHz, DMSO-d6): δ12.04(s,1H),10.27(s,1H),8.45(d,J=15.5Hz,1H),8.04(d,J=2.8Hz,1H),7.77(d,J=12 .2Hz,1H),7.51-7.41(m,1H),7.41-7.28(m,2H),7.20-7.10(m,2H),6.86(t,J=7.4Hz,1H),6.59(d,J=15.5Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.7,163.4,141.6,141.4,138.1,134.4,130.4,130.3,126.9,124.7,122. 8,122.6,121.4,116.8,114.9,111.9,111.7,109.5,109.3,106.0,105.8.HRMS(ESI,m / z):Calcd.for C 17 H 12 ClFN2O[M+Na] + 337.0514; found:337.0513.

[0084] Embodiment 15:

[0085] (E3)-3-(4-Fluoro-1H-indol-3-yl)-N-phenylacrylamide (L 15 )

[0086] 4-Fluoro-1H-indole-3-carboxaldehyde (960 mg, 5.88 mmol) was dissolved in a 50 ml round-bottom flask with pyridine, and malonic acid (612 mg, 5.88 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methanol=40:1] and concentrated by column chromatography to obtain 650 mg of crude product (E3)-3-(4-fluoro-1H-indol-3-yl)acrylic acid. 50 mg of the acid was dissolved in DMF in 2 In a 5mL round-bottom flask, HOAt (40mg, 0.29mmol) was added to react for 10min, and then aniline (22μL, 0.24mmol) and NMM (18μL, 0.17mmol) were added under ice bath conditions to react for 10min, and then EDCI (56mg, 0.29mmol) was added to react at room temperature for 4h. After the reaction was completed by TLC monitoring, ethyl acetate (50mL) and water (50mL×3) were extracted 3 times to remove DMF, and the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 8:1], and the crude product of the concentrated column chromatography was filtered with dichloromethane to obtain the target compound L 15 42.5 mg of yellow solid powder, with a yield of 85%. 1 H NMR (400MHz, DMSO-d6): δ11.93(s,1H),10.06(s,1H),7.87(t,J=16.3Hz,2H),7.72(d,J=8.0Hz,2H),7.31(t,J=8.2 Hz,3H),7.17(td,J=8.0,5.02Hz,1H),7.03(t,J=7.4Hz,1H),6.92(dd,J=11.8,7.8Hz,1H),6.71(d,J=15.6Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.8,157.4,154.9,139.9,139.9,133.9,129.2,128. 8,123.1,123.0,122.9,119.2,117.7,117.6,113.7,111.0,108.9,105.8,105.6.

[0087] Embodiment 16:

[0088] (E3)-3-(4-methyl-1H-indol-3-yl)-N-phenylacrylamide (L16 )

[0089] 4-Methyl-1H-indole-3-carboxaldehyde (1000 mg, 6.28 mmol) was dissolved in a 100 mL round-bottom flask with pyridine, and malonic acid (1307 mg, 12.56 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methanol)] =50:1], concentrated by column chromatography to obtain a crude product (E3)-3-(4-methyl-1H-indol-3-yl)acrylic acid 736.10 mg, and then 150.00 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask, HOAt (121 mg, 0.89 mmol) was added, and the mixture was reacted for 10 min. Aniline (68 μL, 0.75 mmol) and NMM (58 μL, 0.53 mmol) were added in an ice bath and reacted for 10 min. EDCI (171 mg, 0.89 mmol) was added and the mixture was reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The obtained crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 6:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 16 Yellow solid 76.60 mg, yield 51%. 1H NMR (400MHz, DMSO-d6): δ11.47 (s, 1H), 9.85 (s, 1H), 7.95 (d, J = 15.4 Hz, 1H), 7.67 (s, 1H), 7.55 (d, J = 8.0 Hz, 2H), 7.17-7.06 (m, 3H), 6.84 (t, J = 7.5 Hz, 2H), 6.67 (d, J = 7.2 Hz, 1H), 6.39 (d, J = 15.4 Hz, 1H). 13 CNMR (100MHz, DMSO-d6): δ164.7,139.9,136.8,135.2,129.8,128.8,125.0,124.8, 122.9,122.1,121.9,119.1,116.8,113.2,110.3,21.1.HRMS(ESI,m / z):Calcd.for C 18 H 16 N2O[M+Na] + 299.1155; found:299.1156.

[0090] Embodiment 17:

[0091] (E3)-N-(3-bromophenyl)-3-(4-methyl-1H-indol-3-yl)acrylamide (L 17 )

[0092] Same as Example 16 above, first synthesize (E3)-3-(4-methyl-1H-indol-3-yl)acrylic acid, then take 200 mg of the acid and dissolve it in DMF in a 25 mL round-bottom flask, add HOAt (162 mg, 1.19 mmol) and react for 10 min, then add m-bromoaniline (108 μL, 0.99 mmol) and NMM (78 μL, 0.69 mmol) in an ice bath and react for 10 min, then add EDCI (228 mg, 1.19 mmol) and react at room temperature for 4 h. After the reaction is completed as monitored by TLC, extract 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, collect the organic phase, dry it over anhydrous sodium sulfate, and then remove the solvent by rotary evaporation. The obtained crude product is purified by silica gel column chromatography, eluent [V (dichloromethane: V (ethyl acetate) = 60: 1], concentrate the crude product of column chromatography and then filter it with dichloromethane to obtain the target compound L 17 90 mg of yellow solid powder, with a yield of 26%. 1 H NMR (400MHz, DMSO-d6): δ11.69(s,1H),10.19(s,1H),8.14(d,J=15.6Hz,2H),7.89(d,J=2.7Hz,1H),7.57(d,J=8.0Hz,1H), 7.32–7.18(m,2H),7.22(d,J=8.0Hz,1H),7.03(t,J=7.5Hz,1H),6.87(d,J=7.1Hz,1H),6.52(d,J=15.4Hz,1H),2.68(s,3H). 13 C NMR (100MHz, DMSO-d6): δ164.9,141.4,136.8,135.9,130.7,129.8,125.4,1 25.3,124.8,122.1,122.0,121.7,121.4,117.8,116.0,113.0,110.3,20.9.

[0093] Embodiment 18:

[0094] (E3)-3-(5-Bromo-1H-indol-3-yl)-N-phenylacrylamide (L 18 )

[0095] 5-Bromo-1H-indole-3-carboxaldehyde (2000 mg, 8.93 mmol) was dissolved in a 100 ml round-bottom flask with pyridine, and malonic acid (1394 mg, 13.40 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V (methanol) = 40:1], concentrated column chromatography to obtain a crude product (E3)-3-(5-bromo-1H-indol-3-yl) acrylic acid 771mg, then take the acid 133mg and dissolve it in DMF in a 25mL round-bottom flask, add HOAt (82mg, 0.60mmol) and react for 10min, then add aniline (46μL, 0.50mmol) and NMM (39μL, 0.35mmol) under ice bath conditions to react for 10min, then add EDCI (115mg, 0.60mmol) and react at room temperature for 4h. After the reaction is completed as monitored by TLC, it is extracted 3 times with ethyl acetate (50mL) and water (50mL×3) to remove DMF, the organic phase is collected, dried over anhydrous sodium sulfate and then rotary evaporated to remove the solvent, the obtained crude product is purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 6:1], the crude product of the concentrated column chromatography is then filtered with dichloromethane to obtain the target compound L 18 98 mg of light yellow solid powder, with a yield of 74%. 1 H NMR (400MHz, DMSO-d6): δ11.87(s,1H),10.04(s,1H),8.11(d,J=1.8Hz,1H),7.91(s,1H),7.78–7.71(m,2 H),7.70(s,1H),7.46(d,J=8.6Hz,1H),7.38–7.30(m,3H),7.04(t,J=7.3Hz,1H),6.77(d,J=15.8Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.9,139.8,136.3,134.2,132.7,128.9,126.5,125.0,122.9,122.3,119.0,116.5,114.5,113.5,111.9.

[0096] Embodiment 19:

[0097] (E3)-3-(5-bromo-1H-indol-3-yl)-N-(3-fluorophenyl)acrylamide (L 19 )

[0098] Same as Example 18 above, (E3)-3-(5-bromo-1H-indol-3-yl)acrylic acid was first generated, and 133 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask, HOAt (74 mg, 0.54 mmol) was added, and the reaction was continued for 10 min. 3-fluoroaniline (43 μL, 0.45 mmol) and NMM (35 μL, 0.32 mmol) were added under ice bath conditions and the reaction was continued for 10 min. EDCI (104 mg, 0.54 mmol) was added and the reaction was continued at room temperature for 4 h. After the reaction was completed as monitored by TLC, DMF was removed by extraction with ethyl acetate (50 mL) and water (50 mL×3) for 3 times, and the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, and the eluent was [V (petroleum ether): V (ethyl acetate) = 6:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 19 White solid powder 81.20 mg, yield 50%. 1 H NMR (400MHz, DMSO-d6): δ12.04(s,1H),10.44(s,1H),8.11(d,J=2.0Hz,1H),7.91(s,1H),7.84-7.70(m,2H ),7.46(d,J=8.6Hz,1H),7.42-7.35(m,2H),7.35-7.31(m,1H),6.91-6.82(m,1H),6.80(d,J=15.6Hz,1H).

[0099] Embodiment 20:

[0100] (E3)-N-(3-bromophenyl)-3-(5-fluoro-1H-indol-3-yl)acrylamide (L 20 )

[0101] 5-Fluoro-1H-indole-3-carboxaldehyde (973 mg, 5.96 mmol) was dissolved in a 100 mL round-bottom flask with pyridine, and malonic acid (930 mg, 1.50 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography, and the eluent was [V(dichloromethane):V(methanol)= 60:1], concentrated by column chromatography to obtain 856.20 mg of a crude product (E3)-3-(5-fluoro-1H-indol-3-yl)acrylic acid, 150 mg of the acid was dissolved in a 25 mL round-bottom flask with DMF, HOAt (120 mg, 0.88 mmol) was added, the mixture was reacted for 10 min, m-bromoaniline (79 μL, 0.73 mmol) and NMM (57 μL, 0.51 mmol) were added in an ice bath and the mixture was reacted for 10 min, and EDCI (169 mg, 0.88 mmol) was added and the mixture was reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The obtained crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 6:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 20 125 mg of yellow solid powder, with a yield of 83%. 1 H NMR (400MHz, DMSO-d6): δ11.81(s,1H),10.14(s,1H),8.11(s,1H),7.94(s,1H),7.77(d,J=15.8Hz,1H),7.69(d,J=10. 2Hz,1H),7.58–7.41(m,2H),7.30(t,J=7.9Hz,1H),7.35-7.18(m,1H),7.09(t,J=9.2,Hz,1H),6.73(d,J=15.8Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ165.2,141.4,135.0,134.1,133.2,130.8,125.4,125. 1,125.0,121.7,121.2,117.6,115.7,113.5,112.3,110.6,110.4,105.2,105.0.

[0102] Embodiment 21:

[0103] (E3)-N-(3-bromophenyl)-3-(5-methyl-1H-indol-3-yl)acrylamide (L 21 )

[0104] 5-Methyl-1H-indole-3-carboxaldehyde (977 mg, 6.14 mmol) was dissolved in a 100 mL round-bottom flask with pyridine, and malonic acid (958 mg, 9.21 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography, and the eluent was [V(dichloromethane):V(methanol)= 60:1], concentration column chromatography was performed to obtain 666 mg of a crude product (E3)-3-(5-methyl-1H-indol-3-yl)acrylic acid, and 153 mg of the acid was dissolved in a 25 mL round-bottom flask with DMF, HOAt (124 mg, 0.91 mmol) was added, and the mixture was reacted for 10 min. Then, m-bromoaniline (83 μL, 0.76 mmol) and NMM (59 μL, 0.53 mmol) were added in an ice bath and the mixture was reacted for 10 min. Then, EDCI (174.45 mg, 0.91 mmol) was added and the mixture was reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The obtained crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 6:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L 21 192.60 mg of yellow solid powder, with a yield of 37%. 1 H NMR (400MHz, DMSO-d6): δ11.63(s,1H),10.21(s,1H),8.17(s,1H),7.88–7.79(m,2H),7.76(s,1H),7.60(d,J=8.1Hz,1 H),7.39(d,J=8.3Hz,1H),7.31(t,J=8.0Hz,1H),7.23(d,J=8.0Hz,1H),7.08(d,J=8.3Hz,1H),6.76(d,J=15.6Hz,1H). 13C NMR (100MHz, DMSO-d6): δ165.4,141.5,135.9,135.9,131.9,130.8,129.5,125.3,125.1, 124.0,121.7,121.2,119.8,117.6,114.9,112.1,111.8,21.5.HRMS(ESI,m / z):Calcd.for C 18 H 15 BrN2O[M+Na] + 377.0260; found:377.0259.

[0105] Embodiment 22:

[0106] (E3)-3-(6-Bromo-1H-indol-3-yl)-N-phenylacrylamide (L 22 )

[0107] 6-Bromo-1H-indole-3-carboxaldehyde (972 mg, 3.98 mmol) was dissolved in a 100 mL round-bottom flask with pyridine, and malonic acid (622 mg, 5.97 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methane)] The crude product (E3)-3-(6-bromo-1H-indol-3-yl)acrylic acid ( ... 22 211 mg of white solid, yield 91%. 1H NMR (400MHz, DMSO-d6): δ11.78(s,1H),10.03(s,1H),7.90(d,J=8.6Hz,1H),7.88(s,1H),7.76(d,J=9.7Hz,1H ),7.72–7.68(m,2H),7.67(d,J=1.8Hz,1H),7.40-7.27(m,3H),7.04(t,J=7.3Hz,1H),6.81(d,J=15.8Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.8,139.8,138.4,134.1,131.9,128.8,123.9,123.3,122.9,121.6,118.9,116.9,115.1,115.1,112.4.

[0108] Embodiment 23:

[0109] (E3)-3-(6-Bromo-1H-indol-3-yl)-N-(3-bromophenyl)acrylamide (L 23 )

[0110] 6-Bromo-1H-indole-3-carboxaldehyde (972 mg, 3.98 mmol) was dissolved in a 100 ml round-bottom flask with pyridine, and malonic acid (622 mg, 5.97 mmol) and piperidine were added as catalysts. The mixture was reacted at room temperature for 10 h under nitrogen protection. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methane)] The crude product (E3)-3-(6-bromo-1H-indol-3-yl)acrylic acid ( ... 23160 mg of yellow solid powder, yield 54%. 1 H NMR (400MHz, DMSO-d6): δ11.92(s,1H),10.21(s,1H),8.11(s,1H),7.95(s,2H),7.79(d,J=15.1 Hz,1H),7.54(d,J=8.0Hz,1H),7.47(d,J=7.1Hz,1H),7.35–7.11(m,3H),6.81(d,J=15.6Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ165.1,141.4,138.4,134.9,132.3,130.9,125.5,123.9,123. 4,121.7,121.6,121.2,117.6,116.2,115.1,115.1,112.3.HRMS(ESI,m / z):Calcd.for C 17 H 12 Br2N2O[M+Na] + 440.9208; found:440.9203.

[0111] Embodiment 24:

[0112] (E3)-3-(6-chloro-1H-indol-3-yl)-N-phenylacrylamide (L 24 )

[0113] 6-Chloro-1H-indole-3-carboxaldehyde (990 mg, 5.51 mmol) was dissolved in a 100 ml round-bottom flask with pyridine, and malonic acid (1032 mg, 9.90 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, it was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methanol)] )=60:1], concentrated column chromatography to obtain a crude product (E3)-3-(6-chloro-1H-indol-3-yl) acrylic acid 1136mg, then take 250mg of the acid and dissolve it in DMF in a 25mL round-bottom flask, add HOAt (185mg, 1.36mmol) and react for 10min, then add aniline (103μL, 1.13mmol) and NMM (88μL, 0.79mmol) in an ice bath and react for 10min, then add EDCI (260mg, 1.36mmol) and react at room temperature for 4h. After the reaction is completed as monitored by TLC, it is extracted 3 times with ethyl acetate (50mL) and water (50mL×3) to remove DMF, the organic phase is collected, dried over anhydrous sodium sulfate and then rotary evaporated to remove the solvent, the obtained crude product is purified by silica gel column chromatography, the eluent [V(petroleum ether:V(ethyl acetate)=6:1], the crude product of the concentrated column chromatography is then filtered with dichloromethane to obtain the target compound L 24 374.80 mg of yellow solid powder, with a yield of 88%. 1 HNMR (400MHz, DMSO-d6): δ11.77(s,1H),10.03(s,1H),7.95(d,J=8.6Hz,1H),7.89(s,1H),7.81–7.64(m,3H ),7.54(s,1H),7.33(t,J=7.7Hz,2H),7.25(d,J=8.7Hz,1H),7.04(t,J=7.2Hz,1H),6.82(d,J=15.4Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.9,139.8,138.0,134.2,132.0,128.9,127.1,123.7,123.0,121.3,120.8,119.0,116.9,112.4,112.2.

[0114] Embodiment 25:

[0115] (E3)-N-(3-bromophenyl)-3-(6-chloro-1H-indol-3-yl)acrylamide (L 25 )

[0116] As in Example 24, (E3)-3-(6-chloro-1H-indol-3-yl)acrylic acid was first generated, and 198 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask. HOAt (146 mg, 1.07 mmol) was added and reacted for 10 min. Then, m-bromoaniline (97 μL, 0.89 mmol) and NMM (69 μL, 0.62 mmol) were added under ice bath conditions and reacted for 10 min. Then, EDCI (205 mg, 1.07 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, DMF was removed by extraction with ethyl acetate (50 mL) and water (50 mL×3) for 3 times. The organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, and the eluent was [V (petroleum ether: V (ethyl acetate) = 8:1]. The crude product of the concentrated column chromatography was then filtered with dichloromethane to obtain the target compound L. 25 Yellow solid powder 158.30 mg, yield 71%. 1 H NMR (400MHz, DMSO-d6): δ11.91(s,1H),10.45(s,1H),8.14(s,1H),7.98(d,J=8.6Hz,1H),7.90(s,1H),7.77(d,J= 15.7Hz,1H),7.60(d,J=8.2Hz,1H),7.54(s,1H),7.28(t,J=8.0Hz,1H),7.24-7.17(m,2H),6.87(d,J=15.7Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ165.2,141.5,138.0,134.7,132.3,130.8,127.0,125.4,123. 6,121.7,121.4,121.2,120.8,117.7,116.4,112.3,112.1.HRMS(ESI,m / z):Calcd.forC 17 H 12 BrClN2O[M+Na] + 396.9714; found:396.9709.

[0117] Embodiment 26:

[0118] (E3)-3-(7-Bromo-1H-indol-3-yl)-N-phenylacrylamide (L 26 )

[0119] 7-Bromo-1H-indole-3-carboxaldehyde (972 mg, 3.98 mmol) was dissolved in a 100 ml round-bottom flask with pyridine, malonic acid (622 mg, 5.97 mmol) and piperidine were added as catalysts, and the mixture was reacted at room temperature for 10 h under nitrogen protection. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methane)] The crude product (E3)-3-(7-bromo-1H-indol-3-yl)acrylic acid (460 mg) was obtained by concentration column chromatography. 230 mg of the acid was dissolved in DMF in a 25 mL round-bottom flask. HOAt (140 mg, 1.03 mmol) was added and reacted for 10 min. Aniline (78 μL, 0.86 mmol) and NMM (67 μL, 0.60 mmol) were added in an ice bath and reacted for 10 min. EDCI (198 mg, 1.03 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The obtained crude product was purified by silica gel column chromatography, eluent [V(dichloromethane:V(methanol)=60:1], and the crude product of the column chromatography was concentrated and filtered with dichloromethane to obtain the target compound L 26 229 mg of white solid powder, yield 78%. 1 H NMR (400MHz, DMSO-d6): δ11.89(s,1H),10.04(s,1H),7.98(d,J=8.0Hz,1H),7.92(s,1H),7.77(d,J=15.8Hz,1H),7.71(d,J=8 .0Hz,2H),7.47(d,J=7.6Hz,1H),7.33(t,J=7.8Hz,2H),7.17(t,J=7.8Hz,1H),7.04(t,J=7.4Hz,1H),6.86(d,J=15.8Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ164.8,139.8,135.8,134.2,131.9,128.9,126.6,125.1,123.0,121.9,119.5,119.0,117.3,113.4,105.2.

[0120] Embodiment 27:

[0121] (E3)-3-(7-Bromo-1H-indol-3-yl)-N-(3-bromophenyl)acrylamide (L27 )

[0122] 7-Bromo-1H-indole-3-carboxaldehyde (972 mg, 3.98 mmol) was dissolved in a 100 ml round-bottom flask with pyridine, and malonic acid (622 mg, 5.97 mmol) and piperidine were added as catalysts. The mixture was protected by nitrogen and reacted at room temperature for 10 h. After the reaction was completed as monitored by TLC, it was extracted three times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF. The organic phase was collected, dried over anhydrous sodium sulfate, and then rotary evaporated to remove the solvent. The crude product was purified by silica gel column chromatography with an eluent of [V(dichloromethane):V(methanol)] )=60:1], concentrated column chromatography to obtain a crude product (E3)-3-(7-bromo-1H-indol-3-yl) acrylic acid 460mg, then take the acid 230mg and dissolve it in DMF in a 25mL round-bottom flask, add HOAt (140mg, 1.03mmol) and react for 10min, then add m-bromoaniline (94μL, 0.86mmol) and NMM (67μL, 0.60mmol) in an ice bath and react for 10min, then add EDCI (198mg, 1.03mmol) and react at room temperature for 4h. After the reaction is completed as monitored by TLC, it is extracted 3 times with ethyl acetate (50mL) and water (50mL×3) to remove DMF, the organic phase is collected, dried over anhydrous sodium sulfate and then rotary evaporated to remove the solvent, the obtained crude product is purified by silica gel column chromatography, the eluent [V(petroleum ether:V(ethyl acetate)=8:1], the crude product of the concentrated column chromatography is then filtered with dichloromethane to obtain the target compound L 27 332.90 mg of white solid powder, with a yield of 92%. 1 H NMR (400MHz, DMSO-d6): δ11.92(s,1H),10.21(s,1H),8.11(s,1H),7.96(d,J=7.8Hz,2H),7.79(d,J=15.8Hz,1H),7.54(d,J=8 .2Hz,1H),7.47(d,J=7.7Hz,1H),7.30(t,J=8.0Hz,1H),7.23(d,J=8.1Hz,1H),7.17(t,J=7.7Hz,1H),6.81(d,J=15.8Hz,1H). 13 C NMR (100MHz, DMSO-d6): δ165.0,141.3,135.7,134.8,132.2,130.8,126.5,125.4,125.1,122.0,121.7,121.2,119.4,117.6,116.6,113.2,105.1.

[0123] Embodiment 28:

[0124] Preparation of 3-(1H-indol-3-yl)-N-phenylpropionamide (M1)

[0125] 3-(1H-indol-3-yl)propionic acid (500 mg, 2.64 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAT (432 mg, 1.20 mmol) was added, and the mixture was reacted for 10 min. Aniline (0.29 ml, 3.17 mmol) and NMM (206 μL, 1.85 mmol) were added under ice bath conditions and reacted for 10 min. EDCI (608 mg, 3.17 mmol) was then added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, the mixture was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation. The crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 10:1], and the crude product was obtained by column chromatography, and the target compound M1 was obtained as a white solid with a yield of 72%. 1H NMR (400MHz, DMSO-d6): δ10.78(s,1H),9.93(s,1H),7.61(s,1H)7.59–7.55(m,2H),7.33(d,J=8.1Hz,1H),7.28(t,J=8. 0Hz, 2H), 7.13 (d, J = 2.4Hz, 1H), 7.06 (t, J = 7.2Hz, 1H), 7.04-6.94 (m, 2H), 3.02 (t, J = 7.4Hz, 2H), 8.05 (t, J = 7.4Hz, 2H). 13 C NMR (100MHz, DMSO-d6): δ171.1,139.4,136.3,128.7,127.1,123.0,122.2,121 .0,119.1,118.5,118.3,113.8,111.4,37.3,20.9.HRMS(ESI,m / z):Calcd.for C 17 H 16 N2O[M+Na] + 287.1155; found:287.1157.

[0126] Embodiment 29: Preparation of N-(3-bromophenyl)-3-(1H-indol-3-yl)propionamide (M2)

[0127] 3-(1H-indol-3-yl)propionic acid (500 mg, 2.64 mmol) was dissolved in DMF in a 25 mL round-bottom flask, HOAt (432 mg, 1.20 mmol) was added and reacted for 10 min, then m-bromoaniline (0.35 ml, 3.17 mmol) and NMM (206 μL, 1.85 mmol) were added under ice bath conditions and reacted for 10 min, and then EDCI (608 mg, 3.17 mmol) was added and reacted at room temperature for 4 h. After the reaction was completed as monitored by TLC, it was extracted 3 times with ethyl acetate (50 mL) and water (50 mL×3) to remove DMF, the organic phase was collected, dried over anhydrous sodium sulfate, and then the solvent was removed by rotary evaporation, and the crude product was purified by silica gel column chromatography, eluent [V (petroleum ether): V (ethyl acetate) = 10:1], and the crude product was obtained by column chromatography, and then the target compound M2 was obtained as a white solid with a yield of 89%. 1 HNMR (400MHz, DMSO-d6): δ10.81(s,1H),10.13(s,1H),8.02(s,1H),7.58(d,J=7.9Hz,1H),7.51(d,J=7.8Hz,1H),7.36(d,J=8.1Hz,1H ),7.29–7.19(m,2H),7.15(d,J=2.3Hz,1H),7.08(t,J=7.4Hz,1H),6.99(t,J=7.6Hz,1H),3.05(t,J=7.6Hz,2H),2.72(t,J=7.6Hz,2H). 13 C NMR (100MHz, DMSO-d6): δ171.4,140.9,136.2,130.7,127.0,125.6,122.2,121.6,121 .4,121.0,118.3,118.2,117.7,113.5,111.4,37.3,20.7.HRMS(ESI,m / z):Calcd.forC 17 H 15 BrN2O[M+Na] + 365.0260; found:365.0266.

[0128] Biological activity test

[0129] cAMP is a key signaling molecule for many G protein-coupled receptors. The accumulation level of cAMP is mainly tested using the GloSensor method, a bioluminescent cAMP biosensor (Promega). HEK293T cells in good condition were seeded in a 6-well plate or 35MM cell culture dish and placed in a 37°C, 5% CO2 incubator for 24 hours to allow the cells to adhere to the wall. The β2 adrenergic receptor plasmid and pGloSensor were then transfected with a transfection reagent. TM -22FcAMP plasmid was transferred into cells and placed in a 37℃ 5% CO2 incubator for 24 hours to allow transcription and expression of the target gene. Then, the cells transfected with the plasmid were evenly seeded in a 96-well plate and cultured in a 37℃ 5% CO2 incubator for 24 hours. Finally, the old culture medium in the 96-well plate was discarded, the cells were washed with new culture medium, and then the GloSensor was added. TM The balanced culture medium of cAMP Reagent, serum and CO2-independent medium was cultured in a 37°C, 5% CO2 incubator for 1-2 hours or until a stable background signal was obtained. Isoproterenol (ISO) was dissolved in DMSO, sterilized and filtered, and diluted with culture medium to different concentration gradients as the compound mother solution (the DMSO concentration in the prepared compound was less than or equal to 0.1%). The prepared compound was quickly added to a 96-well plate containing HEK293T cells to start drug administration stimulation. The rapid rise of the bioluminescent signal after the addition of the compound can be observed by a multifunctional microplate reader. When the signal value rises to the peak and no longer increases, a compound with a concentration of 50μM is immediately added. At the same time, the bioluminescent signal was immediately collected with a multifunctional microplate reader, and the bioluminescent signal value increased with the increase of the compound concentration. Finally, the data was sorted and processed with GraphPadPrism8 software to obtain the dose-effect curve of the allosteric modulator with concentration as the horizontal axis and signal value as the vertical axis.

[0130] Table 2 Statistics of functional activities of new indole derivatives

[0131]

[0132]

[0133]

[0134] Note: "+" indicates corresponding activity and "-" indicates no corresponding activity

[0135] Table 3 Comparison of the activity of the new derivatives with allosteric antagonistic activity and Cmpd-15

[0136]

[0137]

[0138] Note: a value is expressed relative to the blocking activity of Cmpd-15

[0139] In the Glosensor cAMP accumulation test results, new compounds L1, L2, L4, L6, L7, L8, L9, L 10 , L 11 , L 12 , L 14 , L 15 , L 16 , L 23 , L 24 , L 25 , L 26 , L 27 , M1 and M2 all have β2AR allosteric antagonism, and the other new compounds have no allosteric antagonism activity, as shown in Table 2. Most of the above compounds with allosteric antagonism have better or equivalent activities than Cmpd-15. Among them, the compound with the best activity is L 10 and L 11 The allosteric antagonist activity was significantly improved compared with Cmpd-15. Compounds L1, L4, L7, L6, L9, L 14 , L 16 , L 26 The allosteric antagonist activity of L 23 , L 24 The allosteric antagonist activity of M2 is comparable to that of Cmpd-15, while the activities of the remaining compounds are inferior to those of Cmpd-15 (see Table 3).

[0140] The present invention designs and synthesizes L and M compounds, wherein L is a trans-indole propenyl substituted at the 3-position of indole, while M series compounds have no double bond. As can be seen from Table 3, among all the synthesized indoleamide compounds, compound L 10 and L 11 The allosteric antagonist activity of Cmpd-15 is the best, which is about 2.65 times that of Cmpd-15. The common points are that the trans-indole propenyl on the left is at the 3-position of indole, and both are substituted by Br at the 4-position, which is an electron-withdrawing substitution; the cyclohexylamine and piperidine on the right are both fatty amines.

[0141] Most of the L-type compounds have allosteric antagonistic activity. When trans-3-indoleacrylic acid substituted with a bromine atom at position 4 of R1 is coupled with aliphatic amines such as cyclohexylamine and piperidine, the allosteric activity is the strongest, which is about 2.65 times that of Cmpd-15. When ① trans-3-indoleacrylic acid substituted with a bromine atom, chlorine atom, or methyl atom at position 4 of R1 is coupled with aniline or m-fluoroaniline, ② trans-3-indoleacrylic acid substituted with a bromine atom at position 7 of R1 is coupled with aniline, and ③ trans-3-indoleacrylic acid substituted with a H atom at position 7 of R1 is coupled with aniline or m-fluoroaniline, the allosteric activity is the strongest. The antagonistic activity is strong, about 1.6 times that of Cmpd-15; when trans-3-indoleacrylic acid substituted with bromine or chlorine at position 6 is coupled with aniline or m-bromoaniline, or 3-indoleacrylic acid substituted with H at position 6 is coupled with m-bromoaniline, the allosteric activity is equivalent to that of Cmpd-15; when trans-3-indoleacrylic acid substituted with H at position 7 is coupled with 3-aminobenzonitrile, or trans-3-indoleacrylic acid substituted with bromine at position 7 is coupled with m-bromoaniline, the allosteric antagonistic activity is weaker, not as good as that of Cmpd-15. Among them, the inventors found that when trans-3-indoleacrylic acid substituted with an electron-withdrawing group at position 6 or 7 is coupled with aniline, its antagonistic activity is equivalent to or stronger than Cmpd-15; and when R1 is at position 4, whether trans-3-indoleacrylic acid substituted with an electron-withdrawing group or an electron-donating group is coupled with aliphatic amines or aromatic amines, its antagonistic activity is significantly better than Cmpd-15, so that a compound with better activity can be obtained.

[0142] In addition, in the M-type compounds, the indole propyl group is substituted at the 3-position of indole. When 3-indolepropionic acid is coupled with aniline, its antagonistic activity is only about half of that of Cmpd-15, while when coupled with m-bromoaniline, its antagonistic activity is equivalent to that of Cmpd-15, which is about 1.15 times that of Cmpd-15. It can be seen that the antagonistic activity of indole derivatives without double bonds is inferior to or equivalent to that of Cmpd-15, thus indicating the necessity of double bonds.

[0143] The cAMP accumulation test was used to test whether the target compounds could allosterically regulate the functional activity of the β2AR endogenous ligand ISO. From the results in Table 3, it can be seen that among the new compounds, L1, L2, L4, L6, L7, L8, L9, L 10 , L 11 , L 12 , L 14 , L 15 , L 16 , L 23 , L 24 , L 25 , L 26 , L 27 , M1 and M2 are all negative allosteric regulators of β2AR. 10 and L 11The pharmacological activity of the lead compound (Cmpd-15) is 2.6 times that of the lead compound. In addition, the active compounds with multiple concentrations were added to further confirm whether the compounds are negative allosteric modulators of β2AR. The following are two compounds with good activity (example L 10 , L 11 ) and the allosteric antagonism mechanism diagram of L8. Taking the indoleamide compound L8 as an example, Figure 5 As shown in Figure 1, when the compound concentration increases to a certain level, the curve of ISO shows a significant downward shift and reaches the maximum downward shift limit of the dose-effect regulation of ISO functional activity. Further increase in the compound concentration will hardly cause the dose-effect curve of ISO to shift downward. Therefore, the IC 50 The value may be between 30μM and 60μM, which is manifested as a sudden drop in the concentration curve, that is, the newly synthesized indoleamide derivatives are negative allosteric antagonists of β2-adrenaline receptors with good activity. This allosteric regulation phenomenon is consistent with the reported allosteric antagonism regulation mechanism.

[0144] In summary, these results show that replacing the peptide core structure of Cmpd-15 with an indole skeleton can obtain new compounds with better allosteric antagonistic activity, which provides a direction for the skeleton transition of lead compounds, structural simplification and new drug development, especially when R1 is connected to a fatty amine on the trans-3-indole acrylic acid ring substituted at the 4th position, the antagonistic activity is greatly improved; if it is connected to an aromatic amine, R1 at the 4th position, whether it is an electron-withdrawing group or an electron-donating group, has a stronger antagonistic activity than Cmpd-15. Therefore, it is of great significance to expand the structural types and structural simplification of lead compounds, explore more active β2-adrenaline receptor allosteric antagonists, and develop new drugs.

[0145] Those skilled in the art can understand that the above-mentioned embodiments are specific examples for implementing the present application, and in practical applications, various changes can be made to them in form and detail without departing from the spirit and scope of the present application. Any person skilled in the art can make their own changes and modifications without departing from the spirit and scope of the present application, so the scope of protection of the present application shall be based on the scope defined in the claims.

Claims

1. An indole acrylic acid amide derivative, characterized in that: The structure of the indole acrylic acid amide derivative is shown in Formula 1: R1=H,F,Cl,Br,CN,Me,OCH3; 2. The method for preparing the indole acrylic acid amide derivatives according to claim 1, characterized in that: The preparation method comprises: dissolving trans-indole acrylic acid with different substituents in N,N-dimethylformamide, adding 1-hydroxy-7-azabenzotriazole, stirring at room temperature for 10 minutes, adding N-methylmorpholine and aniline, aliphatic amine or heterocyclic amine with different substituents at 0°C, adding 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride, and reacting at room temperature for several hours to obtain the indole acrylic acid amide derivatives.

3. The method for preparing the indole acrylic acid amide derivatives according to claim 2, characterized in that: The trans-indoleacrylic acid with different substituents includes (E3)-3-(4-bromo-1H-indol-3-yl)acrylic acid, (E3)-3-(5-bromo-1H-indol-3-yl)acrylic acid, and (E3)-3-(6-bromo-1H-indol-3-yl)acrylic acid.

4. The method for preparing the indole acrylic acid amide derivatives according to claim 2, characterized in that: The anilines with different substituents include m-bromoaniline, m-chloroaniline and m-fluoroaniline; the aliphatic amines with different substituents include cyclohexylamine; and the heterocyclic amines with different substituents include piperidine and piperazine.

5. The method for preparing the indole acrylic acid amide derivatives according to claim 2, characterized in that: The preparation method specifically comprises: dissolving 1 mmol of trans-indole propene with different substituents in N,N-dimethylformamide, adding 1.20 mmol of N-hydroxy-7-azabenzotriazole, 1.20 mmol of aniline, aliphatic amine or heterocyclic amine with different substituents and 0.70 mmol of N-methylmorpholine respectively under ice bath conditions, reacting for 10 minutes, then adding 1.20 mmol of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, and then reacting at room temperature for 4 hours; after the reaction is completed, diluting the reaction solution with ethyl acetate, washing with water, and then drying with anhydrous sodium sulfate; after rotary evaporation to remove the solvent, the obtained solid crude product is purified by silica gel column chromatography, eluted with an eluent, and the product after the column chromatography is concentrated, and then filtered with dichloromethane to obtain the described new indole acrylic acid amide derivative.

6. The method for preparing the indole acrylic acid amide derivatives according to claim 5, characterized in that: The eluent is prepared from petroleum ether and ethyl acetate in a volume ratio of 1:

1.

7. Use of the indole acrylic acid amide derivatives as claimed in claim 1 as β2 adrenergic receptor allosteric antagonists.

8. The use according to claim 7, characterized in that: The indole acrylic acid amide derivatives are used to exert an allosteric antagonistic effect on the G-protein signaling pathway of β2AR, and negatively regulate the agonistic effect of endogenous agonist isoproterenol on β2AR.

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