N-aryl indole derivative and application thereof

Abstract

The invention relates to the technical field of medicines, in particular to an N-arylindole derivative and application thereof in preparation of a medicine for resisting cerebral arterial thrombosis. The invention relates to an N-arylindole derivative. The N-arylindole derivative is a compound as shown in a formula I, a pharmaceutically acceptable salt and an optical active body or racemate thereof, and a diastereoisomer thereof, by deeply exploring the regulation mechanism of the cerebral arterial thrombosis, potential therapeutic targets are defined, and a new research direction is provided for developing the medicine for effectively treating the cerebral arterial thrombosis. The research further enriches the structure type of the small molecule inhibitor for resisting cerebral ischemic stroke, lays a foundation for the biological activity test of subsequent monomeric compounds, not only provides an active lead compound for the development of new drugs, but also provides theoretical support for the deep research and development of the GluN2B-NMDAR antagonist in the field of cerebral stroke treatment.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to an N-arylindole derivative and its application in the preparation of drugs for treating ischemic stroke. Background Technology

[0002] Stroke has become the second leading cause of death worldwide, affecting approximately 15 million people annually. This significant burden is attributed to its limited treatment window, high mortality rate, severe disability, and frequent recurrence. Stroke typically results in irreversible brain damage, usually caused by the sudden rupture or blockage of a blood vessel in the brain. Strokes are classified into ischemic and hemorrhagic types, with ischemic strokes accounting for 80% of cases. Ischemic strokes, primarily caused by blockage of blood vessels in the brain, are a leading cause of neurological deficits and death. Cerebrovascular diseases are a major cause of death and disability globally, resulting in millions of deaths and long-term functional impairments each year. In recent years, the incidence of ischemic stroke has been rising, showing a trend towards affecting younger people, seriously threatening human health. Therefore, developing drugs with neuroprotective effects and the ability to improve functional recovery after stroke is of great significance for improving the quality of life for patients. The etiology of ischemic stroke is complex, involving multiple molecular pathways and cellular damage mechanisms, and its specific pathogenesis is not yet fully understood. Currently, the number of drugs available for ischemic stroke in clinical practice is limited, and these drugs suffer from problems such as a short therapeutic window and narrow applicability. There is also a lack of effective drugs for repairing neurological damage after stroke. Ischemic brain injury involves multiple mechanisms, including excitotoxicity, oxidative stress, and neuroinflammation. During an ischemic stroke, excessive release of glutamate by neurons can lead to excitotoxicity, ultimately resulting in neuronal cell death.

[0003] Glutamate is a major excitatory neurotransmitter in the central nervous system (CNS), playing a crucial role in regulating neuronal activity. Glutamate receptors are mainly divided into two types: metabotropic glutamate receptors (mGluRs) and ionotropic glutamate receptors (iGluRs). Excessive accumulation of glutamate and overstimulation of its receptors can induce excitotoxicity, a process that promotes the production of calcium ions (Ca). 2+Excitotoxicity flows into neurons, leading to oxidative stress, mitochondrial dysfunction, and ultimately neuronal death. Excitotoxicity is a crucial factor in the pathogenesis of stroke. Ionotropic glutamate receptors include N-methyl-D-aspartate receptors (NMDARs), karyophyllate receptors (KARs), and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), with NMDARs being the primary mediators of excitotoxicity. NMDARs are heterotetrameric proteins composed of subunits from three gene families: GluN1 (GluN1a-4a, GluN1b-4b), GluN2 (GluN2A-D), and GluN3 (GluN3A-B). Structurally, functional NMDARs consist of two GluN1 and two GluN2 subunits. Although GluN1 is widely expressed in the central nervous system, the spatial distribution of the GluN2 subunits is determined by the expression patterns of the corresponding genes. NMDAR isotypes exhibit different biophysical, pharmacological, and signal transduction properties, contributing to different physiological and pathological functions. By promoting the influx of calcium ions into neurons, NMDARs mediate excitatory synaptic transmission and synaptic plasticity, both essential for learning and memory. Excitotoxicity is primarily triggered by the selective activation of NMDARs containing the GluN2B subunit. Overactivation of these GluN2B-containing NMDARs leads to excessive calcium influx into neurons, potentially contributing to neurodegenerative diseases such as Parkinson's and Alzheimer's. Growing evidence suggests that GluN2B-NMDARs have the potential to treat neurodegenerative diseases, including ischemic stroke, depression, Alzheimer's, and Parkinson's.

[0004] Non-selective NMDA receptor antagonists, such as MK-801 and the racemic mixture of ketamine, were initially developed for analgesia, but they have significant toxicity and potential addictive risks. In contrast, selective GluN2B receptor antagonists may offer stronger therapeutic effects and fewer adverse reactions. Ifenprodil was the first reported GluN2B-NMDAR antagonist. However, it has poor oral bioavailability and exhibits off-target activity against α1, σ1, σ2 receptors, and hERG ion channels. Despite these limitations, Ifenprodil remains an important lead compound for designing novel GluN2B-NMDAR antagonists. Structure-related compounds, such as Ro 25-6981 and CP-101,606, showed better selectivity and tolerability but were discontinued due to concerns about cardiovascular toxicity. CERC-301, another potent orally bioavailable GluN2B-NMDAR antagonist, failed to meet its primary efficacy endpoint in a large phase II clinical trial. A 2016 study reported the co-crystal structure of EVT-101 bound to a GluN1 / GluN2B-containing receptor. The results showed that the benzyl ring of EVT-101 occupies the same hydrophobic pocket as the benzylpiperidine moiety of Ifenprodil, while its pyrazine and imidazole groups bind to different sites. Furthermore, other GluN2B-NMDAR antagonists such as Neu2000, Radiproglucil, and MIJ-821 are currently in clinical development. These studies highlight the structural and functional characteristics of selective GluN2B-NMDAR antagonists, providing important insights for the development of next-generation compounds. Summary of the Invention

[0005] The purpose of this invention is to design and synthesize an N-arylindole derivative based on previous work and its application in the preparation of drugs for treating ischemic stroke.

[0006] Another object of the present invention is to provide a method for preparing N-arylindole derivatives.

[0007] In order to achieve the objective of this invention, the following technical solution is adopted:

[0008] An N-arylindole derivative, characterized in that: the N-arylindole derivative is a compound of Formula I, a pharmaceutically acceptable salt thereof, or its optically active form or racemate, or a diastereomer;

[0009]

[0010] in:

[0011] R1 and R2 may be the same or different and are independently selected from H, hydroxyl, halogen, cyano, nitro, unsubstituted or substituted with at least one R3, C1-6 alkyl, C3-8 cycloalkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkylsulfonylamino, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 alkoxy, C2-6 alkenyloxy, C2-6 alkynyloxy, C1-6 alkylthio, C2-6 alkenylthio, C2-6 alkynylthio, C1-6 alkylsulfonyl, C2-6 alkenylsulfonyl, C2-6 alkynylsulfonyl, C1-6 alkylsulfinyl, C2-6 alkenylsulfonyl, C1-6 alkylsulfinyl, C2 ... 2-6-Alkenylsulfinyl, C2-6-alkynylsulfinyl, C3-8-cycloalkenyl; unsubstituted or substituted carbonyl group with at least one R4, unsubstituted or substituted amino group with at least one R5, unsubstituted or substituted phenyl group with at least one R6, benzyl, benzyloxy, benzoyl, benzylsulfonyl, pyridine ring, pyrazole ring, pyrrole ring, pyrimidine ring, quinoline ring, isoquinoline ring, imidazole ring, morpholine ring, piperazine ring, pyridazine ring, pyrazine ring, piperidine ring, thiophene ring, thiazole ring, isothiazole ring, benzothiazole ring, pyran ring, indole ring, oxazole ring, isoxazole ring, triazole ring, benzotriazole ring, or furan ring;

[0012] R3 is selected from H, halogen, hydroxyl, cyano, nitro or amino;

[0013] R4 is selected from H, hydroxyl, C1-6 alkyl, amino, C1-6 alkylamino, C1-6 alkoxy, C1-6 alkylthio, and C3-8 cycloalkyl.

[0014] R5 is selected from H, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkylsulfonyl, C2-6 alkenylsulfonyl, C2-6 alkynylsulfonyl, C1-6 alkyl carbonyl, C2-6 alkenyl carbonyl, and C2-6 alkynyl carbonyl.

[0015] R6 is selected from H, halogen, C1-6 alkyl, C1-6 alkylamino, C3-7 cycloalkyl, C1-6 alkoxy, C1-6 alkylsulfonylamino, benzyloxycarbonyl, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 haloalkyl, hydroxyl, cyano, nitro, formyl, and amino.

[0016] n is selected from 0, 1, 2, 3, or 4; m is selected from 0, 1, 2, 3, 4, or 5;

[0017] X is selected from O, NH, or NHNHCO.

[0018] Preferably, the N-arylindole derivative is a compound of Formula I, a pharmaceutically acceptable salt, or its optically active form or racemate, or a diastereomer;

[0019] In the formula,

[0020] R1 and R2 may be the same or different and are independently selected from H, hydroxyl, halogen, cyano, nitro, unsubstituted or substituted with at least one R3 C1-6 alkyl, C3-8 cycloalkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkylsulfonylamino, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 alkoxy, C2-6 alkenyloxy, C2-6 alkynyloxy, C1-6 alkylthio, C2-6 alkenylthio; unsubstituted or substituted with at least one R6 phenyl, benzyl, benzyloxy, benzoyl;

[0021] R3 is selected from H, halogen, hydroxyl, cyano, nitro or amino;

[0022] R6 is selected from H, halogen, C1-6 alkyl, C1-6 alkylamino, C3-7 cycloalkyl, C1-6 alkoxy, C1-6 alkylsulfonylamino, benzyloxycarbonyl, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 haloalkyl, hydroxyl, cyano, nitro, formyl, and amino.

[0023] N is 0, 1, 2, 3, or 4; m is selected from 1, 2, 3, or 4;

[0024] X is selected from O, NH, or NHNHCO.

[0025] Further preferably, the N-arylindole derivative is a compound of Formula I, a pharmaceutically acceptable salt, or its optically active form or racemate, or a diastereomer;

[0026] In the formula,

[0027] R1 and R2 may be the same or different and are independently selected from H, hydroxyl, halogen, nitro, unsubstituted or substituted with at least one R3 C1-6 alkyl, C2-6 alkenyl, C2-6 alkoxy, C2-6 alkenyloxy, C2-6 alkoxy; unsubstituted or substituted with at least one R6 phenyl, benzyl;

[0028] R3 is selected from H, halogen, hydroxyl, cyano, nitro or amino;

[0029] R6 is selected from H, halogen, C1-6 alkyl, C1-6 alkylamino, C3-7 cycloalkyl, C1-6 alkoxy, C1-6 alkylsulfonylamino, benzyloxycarbonyl, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 haloalkyl, hydroxyl, cyano, nitro, formyl, and amino.

[0030] N is 0, 1, 2, 3, or 4; m is selected from 1, 2, 3, or 4;

[0031] X is selected from O, NH, or NHNHCO.

[0032] Further preferred, the N-arylindole derivative is:

[0033] N-(4-hydroxybenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI1);

[0034] N-Benzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI2);

[0035] 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylbenzyl)acetamide (NFI3);

[0036] N-(4-methoxybenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI4);

[0037] N-(4-fluorobenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI5);

[0038] N-(4-chlorobenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI6);

[0039] N-Benzyl-2-(5-bromo-1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI7);

[0040] 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylbenzyl)acetamide (NFI8);

[0041] 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methoxybenzyl)acetamide (NFI9);

[0042] 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxybenzyl)acetamide (NFI10);

[0043] N-(2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazine (NFI11);

[0044] 4-Methyl-N-(2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI12);

[0045] 4-Methoxy-N-(2-(1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI13);

[0046] 4-Hydroxy-N-(2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazine (NFI14);

[0047] Benzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI15);

[0048] 4-Methylbenzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI16);

[0049] 4-Chlorobenzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI17);

[0050] 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenylacetamide (NFI18);

[0051] N-(4-methoxyphenyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI19);

[0052] 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenylethylacetamide (NFI20);

[0053] N-(4-methoxyphenethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI21);

[0054] 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylphenethyl)acetamide (NFI22);

[0055] N-(4-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI23);

[0056] 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(3-phenylpropyl)acetamide (NFI24);

[0057] 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI25);

[0058] 2-(5-chloro-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI26);

[0059] 2-(5-fluoro-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI27);

[0060] N-(2-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI28);

[0061] N-(3-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI29);

[0062] N-(3,4-dihydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI30);

[0063] 2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenethylacetamide (NFI31);

[0064] 2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylphenethyl)acetamide (NFI32);

[0065] N-(4-methoxyphenethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI33);

[0066] N-(4-hydroxyphenylethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI34);

[0067] N-(4-fluorophenylethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI35).

[0068] The structural formulas of the above compounds are as follows:

[0069]

[0070]

[0071]

[0072] The pharmaceutically acceptable salt is obtained by reacting the compound of Formula I with an acid to obtain the corresponding salt; wherein the acid is hydrochloric acid, sulfuric acid, phosphoric acid, tartaric acid, citric acid, methanesulfonic acid, hydrobromic acid, maleic acid, or benzenesulfonic acid.

[0073] A pharmaceutical composition comprising the aforementioned N-arylindole derivative, a pharmaceutically acceptable salt thereof, and its optically active form or racemate, or a diastereomer.

[0074] A pharmaceutical formulation comprising, as an active ingredient, a pharmaceutically acceptable salt thereof and its optically active form or racemate, diastereomer; or the pharmaceutical composition thereof, wherein the active ingredient is mixed with a pharmaceutically acceptable carrier, wherein the active ingredient comprises 0.01-99.9% of the formulation mass.

[0075] The formulation is a tablet, coated tablet, capsule, solution for injection, ampoule, suppository, patch, inhalable powder, suspension, emulsion or ointment.

[0076] One application of the compound of Formula I, a pharmaceutically acceptable salt thereof and its optically active form or racemate, diastereomer, pharmaceutical composition or pharmaceutical formulation, as a GluN2B-NMDAR antagonist.

[0077] The use of the compound of Formula I, its pharmaceutically acceptable salt and its optically active form or racemate, diastereomer, pharmaceutical composition or pharmaceutical preparation in the preparation of an anti-ischemic stroke drug.

[0078] The synthetic routes for N-arylindole derivatives of the above general formula I, or pharmaceutically acceptable salts thereof, and their optically active forms or racemates, and diastereomer mixtures, include the following:

[0079]

[0080] The synthetic routes and reaction conditions for compounds NFI1–NFI10 and NFI15–NFI30 are as follows: (a) 3-formylphenylboronic acid, copper acetate, triethylamine, pyridine, Molecular sieve, anhydrous dichloromethane, room temperature, 24 hours;

[0081] (b) Sodium borohydride, tetrahydrofuran, room temperature, 1 hour; (c) Triphenylphosphine, carbon tetrabromide, dichloromethane, room temperature, 1 hour; (d) 2-methylimidazole, sodium hydride, N,N-dimethylformamide, 0°C, 2 hours; (e) Sodium hydroxide / methanol system, reacted at 0°C for 2 hours, then treated with aqueous hydrochloric acid; (f) N,N,N',N'-tetramethylchloromethanesulfonyl hexafluorophosphate, N-methylimidazole, N,N-dimethylformamide, room temperature, 1.5 hours;

[0082] (g) 1-Ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, 4-dimethylaminopyridine, N,N-dimethylformamide, room temperature, 1 hour.

[0083] The synthesis of compounds NFI1–NFI10 and NFI15–NFI30 began with commercially available intermediate 1a. First, intermediate 2a was obtained via a Chan-Lam coupling reaction with 3-formylphenylboronic acid, followed by a reduction step to yield intermediate 3a. Next, 3a underwent an Appel reaction to generate intermediate 4a, which was then reacted with 2-methylimidazole to give intermediate 5a. Finally, 5a underwent alcoholysis to generate the key intermediate 6a, which was then condensed with aromatic amines and aromatic alcohols of varying substitutions to obtain the target products.

[0084]

[0085] Synthesis of compounds NFI11–NFI14: (a) N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate, N-methylimidazolium NMI, N,N-dimethylformamide, room temperature, 1.5 h.

[0086] The final products NFI11–NFI14 were synthesized by the condensation reaction of the key intermediate 6a with benzoylhydrazides of different substitutions.

[0087]

[0088] Synthesis of compounds NFI31–NFI35. Reagents and reaction conditions: (a) 2-formylphenylboronic acid, copper acetate, triethylamine, pyridine, (a) Molecular sieve, anhydrous dichloromethane, room temperature, 24 hours; (b) Sodium borohydride, tetrahydrofuran, room temperature, 1 hour; (c) Triphenylphosphine, carbon tetrabromide, dichloromethane, room temperature, 1 hour; (d) 2-methylimidazole, sodium hydride, N,N-dimethylformamide, 0°C, 2 hours; (e) Sodium hydroxide / methanol, reacted at 0°C for 2 hours, then treated with hydrochloric acid aqueous solution; (f) N,N,N',N'-tetramethylchloromethanesulfonyl hexafluorophosphate, N-methylimidazole, N,N-dimethylformamide, room temperature, 1.5 hours.

[0089] The synthetic route for compounds NFI31–NFI35 begins with commercially available ethyl-3-indoleacetic acid (1b) as the starting material, and proceeds via a Chan-Lam coupling reaction with 2-formylphenylboronic acid to obtain intermediate 2b. Subsequent reaction steps are the same as described above. Finally, the key intermediate 6b is condensed with phenethylamines of varying substitutions to yield the final product.

[0090] The beneficial effects of this invention are:

[0091] This invention relates to the use of N-arylindole derivatives of Formula I, or their isomers, or pharmaceutically acceptable salts, or pharmaceutical compositions thereof, in the preparation of drugs for treating ischemic stroke. This invention, through in-depth exploration of the regulatory mechanisms of ischemic stroke, clarifies potential therapeutic targets and provides a new research direction for the development of effective drugs for treating ischemic stroke. This research further enriches the structural types of small molecule inhibitors for treating ischemic stroke and lays the foundation for subsequent bioactivity testing of monomeric compounds. It not only provides active lead compounds for new drug development but also provides theoretical support for the in-depth research and development of GluN2B-NMDAR antagonists in the field of stroke treatment. Attached Figure Description

[0092] Figure 1 This is a graph showing the effect of NFI23 calcium ion concentration measurement provided in an embodiment of the present invention. In this graph, A represents representative green fluorescence images captured at different time points, showing intracellular calcium in PC12 cells. 2+ The concentration changes are shown in Figure B, which represents the fluorescence intensity curves of each group within the detection period.

[0093] Figure 2 The above is a graph showing the effect of NFI23 reactive oxygen species measurement provided in the embodiments of the present invention. In the graph, A is a flow cytometry analysis of ROS levels in PC12 cells, and B is a quantitative analysis of the percentage of ROS-positive cells.

[0094] Figure 3 The image shows the effect of NFI23 mitochondrial membrane potential measurement provided in the embodiment of the present invention. In the image, A is a flow cytometry analysis of MMPs in PC12 cells, and B is a quantitative analysis of MMP loss.

[0095] Figure 4 The above diagram shows the effect of NFI23 cell apoptosis assay provided in the embodiment of the present invention. In the diagram, A is a flow cytometry analysis of PC12 cell apoptosis and B is a quantitative analysis of the percentage of apoptotic cells.

[0096] Figure 5 The image shows the Western blot (WB) results of NFI23 provided in this embodiment of the invention. In the image, A represents the effect of NFI23 on the expression levels of p-ERK1 / 2 and total ERK1 / 2, B represents the quantitative analysis of the expression ratio of p-ERK1 / 2 to total ERK1 / 2, and C represents the quantitative analysis of the expression ratio of p-ERK1 / 2 to β-actin.

[0097] Figure 6 The diagram shows the patch-clamp effect of NFI23 provided in this embodiment of the invention. In this diagram, A represents the effect of Ifenprodil on the dielectric current of GluN1 / GluN2b, B represents the effect of NFI23 on the dielectric current of GluN1 / GluN2b, and C represents the IC of Ifenprodil. 50 Fitted curve, D is the IC of NFI23 50 Fitted curve.

[0098] Figure 7 The image shows the therapeutic effect of NFI23 on MCAO rats provided in this embodiment of the invention. In the image, A is a representative TTC-stained brain slice, B is a quantitative analysis of the cerebral infarction area, and C is a quantitative analysis of the neurological function score.

[0099] Figure 8 The molecular dynamics simulation diagram of NFI23 provided in the embodiments of the present invention is shown below. A is the RMSD curve of NFI23, GluN2B-NMDAR and NFI23-GluN2B-NMDAR complex, B is the RMSF curve of GluN2B-NMDAR, C is the Rg curve of GluN2B-NMDAR, D is the hydrogen bond number diagram of GluN2B-NMDAR-NFI23 complex, E is the 2D landscape diagram of Gibbs free energy of GluN2B-NMDAR-NFI23 complex, and F is the 3D landscape diagram of Gibbs free energy of GluN2B-NMDAR-NFI23 complex. Detailed Implementation

[0100] The compounds of the present invention and their preparation will be better understood in conjunction with the following examples, which are intended to illustrate rather than limit the scope of the invention.

[0101] Example 1: N-(4-hydroxybenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI1)

[0102] At room temperature, ethyl indole-3-acetate (1a, 10.00 g, 49.20 mmol) and 3-formylphenylboronic acid (14.75 g, 98.40 mmol) were dissolved in 100 mL of anhydrous dichloromethane. After complete dissolution, copper(II) acetate (17.87 g, 98.40 mmol), triethylamine (9.97 g, 98.40 mmol), and pyridine (7.78 g, 98.40 mmol) were added sequentially, followed by 2 g of [a specific ingredient / component - likely a specific ingredient or component]. Molecular sieves were used. The reaction mixture was stirred at room temperature for 24 hours, and the reaction progress was monitored by thin-layer chromatography (TLC). After the reaction was complete, the mixture was filtered to remove residual solids, and the filtrate was concentrated under reduced pressure to give a crude product. The crude product was purified by column chromatography (eluent: petroleum ether / ethyl acetate = 1:1) to give a white solid 2a (11.84 g), with a yield of 78.37%. The synthesis of intermediate 2b was carried out using the same method. Intermediate 2a (5.00 g, 16.27 mmol) was dissolved in 50 mL of anhydrous tetrahydrofuran (THF), and sodium borohydride (0.92 g, 24.40 mmol) was added. The reaction mixture was stirred for 1 hour, and the reaction progress was monitored by thin-layer chromatography (TLC). After the reaction was complete, 100 mL of water was added to quench the reaction, followed by extraction with ethyl acetate (30 mL × 3). The combined organic layers were washed with saturated brine (100 mL × 2) and dried over anhydrous sodium sulfate. The solvent was removed under reduced pressure to give a white solid 3a (4.70 g), with a yield of 93.47%. The synthesis of intermediate 3b was carried out using the same method. Intermediate 3a (3.00 g, 9.70 mmol) was dissolved in 30 mL of anhydrous dichloromethane, followed by the addition of triphenylphosphine (3.05 g, 11.64 mmol) and carbon tetrabromide (3.86 g, 11.64 mmol). The reaction mixture was stirred for 1 hour, and the reaction progress was monitored by thin-layer chromatography (TLC). After the reaction was complete, the solvent was removed under reduced pressure, and the crude product was purified by column chromatography (eluent: petroleum ether / ethyl acetate = 1:1) to give a white solid 4a (3.30 g), with a yield of 91.28%. The synthesis of intermediate 4b was carried out using the same method. 2-Methylimidazole (0.50 g, 6.09 mmol) was dissolved in 10 mL of anhydrous tetrahydrofuran (THF) in an ice bath at 0 °C. Sodium hydride (60%, 0.49 g, 20.30 mmol) was added, and the mixture was stirred for 30 min. Intermediate 4a (1.82 g, 4.87 mmol) was then added, and the reaction was continued at room temperature for 1 h, with the reaction progress monitored by thin-layer chromatography (TLC). After the reaction was complete, 20 mL of water was added to quench the reaction, and the mixture was extracted with ethyl acetate (8 mL × 3). The combined organic phases were washed twice with saturated brine (8 mL × 2), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (eluent: petroleum ether / ethyl acetate = 5:1) to give a white solid 5a (1.58 g) in 86.54% yield. The synthesis of intermediate 5b was carried out using the same method. Intermediate 5a (1.00 g, 2.68 mmol) was dissolved in 20 mL of 4N methanol-sodium hydroxide solution in an ice bath at 0 °C. The mixture was stirred for 2 hours, and the reaction progress was monitored by thin-layer chromatography (TLC). After the reaction was complete, methanol was evaporated under reduced pressure, and the residue was dissolved in 20 mL of water.Simultaneously, 2N hydrochloric acid solution was slowly added dropwise in an ice bath until the pH of the solution dropped to 3–4, forming a white precipitate. The precipitate was filtered and dried to obtain intermediate 6a (0.86 g), with a yield of 93.21%. The synthesis of intermediate 6b was carried out using the same method. At room temperature, intermediate 6a (0.50 g, 1.42 mmol) and 4-hydroxybenzylamine (0.19 g, 1.56 mmol) were dissolved in 5 mL of anhydrous DMF. Then, TCFH (0.48 g, 1.70 mmol) and NMI (0.14 g, 1.70 mmol) were added sequentially to the reaction mixture, and the reaction was carried out with stirring for 1.5 h, with the reaction progress monitored by thin-layer chromatography (TLC). After the reaction was complete, 30 mL of water was added, followed by extraction with ethyl acetate (10 mL × 3). The combined organic layers were washed with saturated brine (10 mL × 2) and then concentrated under reduced pressure. The crude product was purified by column chromatography (eluent: dichloromethane / methanol = 20:1) to give a yellow solid NFI1 (0.47 g), with a yield of 74.28%.

[0103] 1 H NMR(600MHz,DMSO-d6)δ9.28(s,1H),8.42(t,J=5.9Hz,1H),7.65(d,J=7.9Hz,1H) ,7.59(t,J=7.8Hz,1H),7.53(d,J=8.1Hz,3H),7.49(s,1H),7.46(s,1H),7.26(s,1 H),7.20(dd,J=11.3,7.5Hz,2H),7.12(t,J=7.5Hz,1H),7.05(d,J=8.5Hz,2H),6. 67(d,J=8.4Hz,2H),5.39(s,2H),4.17(d,J=5.8Hz,2H),3.62(s,2H),2.48(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.39,156.70,144.81,139.95,138.29,135.44,130.93,130.11,129.37,129.12,127.17,125.43,12 3.41,123.02,122.81,122.51,121.96,120.47,120.16,115.42,112.43,110.73,49.52,42.33,32.74,12.05.ESI-HRMS:calcd for C 28 H 27 N4O2,[M+H] + ,451.2134; found 451.2133.

[0104] The compounds NFI2–NFI14 and NFI18–NFI35 were synthesized using the same method.

[0105] Example 2: N-Benzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI2)

[0106] When 4-hydroxybenzylamine is replaced with benzylamine in the above preparation process, NFI2 is obtained; a yellow solid with a yield of 80.26%.

[0107] 1 H NMR (600MHz, DMSO-d6) δ8.53(t,J=5.9Hz,1H),7.66(d,J=7.8Hz,1H),7.57(t,J=7.8Hz,1H) ,7.52(d,J=8.1Hz,2H),7.50(s,1H),7.42(s,2H),7.29(t,J=7.4Hz,2H),7.24(d,J=7.4Hz, 2H),7.22(d,J=6.9Hz,1H),7.19(d,J=8.5Hz,1H),7.18(d,J=7.8Hz,1H),7.13(d,J=7.2Hz, 1H),7.11(d,J=11.2Hz,1H),5.35(s,2H),4.29(d,J=6.0Hz,2H),3.66(s,2H),2.40(s,3H). 13 C NMR (150MHz, DMSO-d6) δ170.62,140.02,139.91,138.83,135.46,130.88,129.35,129.31,128.70,127.74,127.2 4,127.22,125.30,123.27,122.64,120.48,120.12,112.30,110.74,49.30,42.75,32.73,12.43.ESI-HRMS:calcd forC 28 H 27 N4O, [M+H] + ,435.2185; found 435.2192.

[0108] Example 3 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylbenzyl)acetamide (NFI3)

[0109] When 4-hydroxybenzylamine is replaced with 4-methylbenzylamine in the above preparation process, NFI3 is obtained; a yellow solid with a yield of 84.78%.

[0110] 1 H NMR (600MHz, DMSO-d6) δ8.51(t,J=5.9Hz,1H),7.67(d,J=7.9Hz,1H),7.61(t,J=7.8H z,1H),7.58(d,J=5.8Hz,2H),7.57–7.53(m,2H),7.51(d,J=3.9Hz,1H),7.33(d,J=7.9 Hz,1H),7.25–7.20(m,2H),7.15(d,J=4.0Hz,1H),7.13(d,J=4.9Hz,2H),7.10(d,J=7 .9Hz,2H),5.48(s,2H),4.24(d,J=5.9Hz,2H),3.65(s,2H),2.63(s,3H),2.26(s,3H). 13 C NMR (150MHz, DMSO-d6) δ170.62,140.02,139.91,138.83,135.46,130.88,129.35,129.31,128.70,127.74,127.2 4,127.22,125.30,123.27,122.64,120.48,120.12,112.30,110.74,49.30,42.75,32.73,12.43.ESI-HRMS:calcd for C 29 H 29 N4O, [M+H] + ,449.2341;found 449.2341.

[0111] Example 4: N-(4-methoxybenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI4)

[0112] When 4-hydroxybenzylamine is replaced with 4-methoxybenzylamine in the above preparation process, NFI4 is obtained; a white solid with a yield of 87.36%.

[0113] 1H NMR (600MHz, DMSO-d6) δ8.46(t,J=5.8Hz,1H),7.65(d,J=7.9Hz,1H),7.57(t,J=7.8Hz,1H),7.50(dd,J=16.3,7.4Hz,3H),7.39(s,1H),7.37(s, 1H),7.22–7.10(m,5H),6.99(d,J=8.6Hz,1H),6.85(d,J=8.6Hz,2H),5. 33(s,2H),4.21(d,J=5.9Hz,2H),3.72(s,3H),3.63(s,2H),2.37(s,3H). 13 C NMR(150MHz,DMSO-d6)δ158.67,139.90,139.15,130.92,130.83,129.36,129.12,127.18,125.20,124.78,123.1 6,123.01,122.52,120.46,120.12,114.50,114.11,110.72,55.53,49.14,42.23,32.75,12.63.ESI-HRMS:calcd forC 29 H 29 N4O2,[M+H] + ,465.2291; found 465.2294.

[0114] Example 5: N-(4-fluorobenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI5)

[0115] When 4-hydroxybenzylamine is replaced with 4-fluorobenzylamine in the above preparation process, NFI5 is obtained; a yellow solid with a yield of 88.47%.

[0116] 1H NMR (600MHz, DMSO-d6) δ8.55(t,J=6.0Hz,1H),7.65(d,J=7.9Hz,1H),7.59(t,J= 7.8Hz,1H),7.55–7.52(m,3H),7.50(s,1H),7.48(s,1H),7.30–7.29(m,1H),7.28 (d,J=5.6Hz,2H),7.22(d,J=6.0Hz,1H),7.20(d,J=7.2Hz,1H),7.13(s,1H),7.1 1(d,J=8.9Hz,2H),5.40(s,2H),4.27(d,J=5.9Hz,2H),3.66(s,2H),2.48(s,3H). 13 C NMR (150MHz, DMSO-d6) δ170.66,138.24,136.24,131.77,131.72,130.94,129.70,129.35,127.26,125.47 ,123.45,123.05,122.84,120.09,115.48,115.34,112.24,110.77,49.54,42.06,12.02.ESI-HRMS:calcd for C 28 H 26 N4OF,[M+H] + ,453.2091; found 453.2095.

[0117] Example 6: N-(4-chlorobenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI6)

[0118] When 4-hydroxybenzylamine is replaced with 4-chlorobenzylamine in the above preparation process, NFI6 is obtained; a white solid with a yield of 86.57%.

[0119] 1 H NMR (600MHz, DMSO-d6) δ8.56(t,J=6.0Hz,1H),7.64(d,J=7.9Hz,1H),7.58(t,J=7.8Hz,1H),7.54–7.46(m,4H),7.43(s,1H),7.36–7. 34(m,2H),7.28–7.25(m,2H),7.22–7.17(m,2H),7.13(t,J=7.4Hz,2H),5.36(s,2H),4.27(d,J=6.0Hz,2H),3.66(s,2H),2.42(s,3H). 13C NMR(150MHz,DMSO-d6)δ170.75,139.91,139.15,138.74,135.48,131.75,131.33,130.88,129.61,129.33,129.10,128.62 ,127.28,125.34,123.75,123.29,123.04,122.67,120.51,120.08,112.18,110.76,49.36,32.73,12.37.ESI-HRMS:calcd forC 28 H 26 N4OCl,[M+H] + ,469.1795; found 469.1796.

[0120] Example 7: N-Benzyl-2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI7)

[0121] By replacing 4-hydroxybenzylamine with benzylamine and indole-3-ethyl acetate with 5-bromo-indole-3-ethyl acetate in the above preparation process, NFI7 is obtained; a yellow solid with a yield of 81.24%.

[0122] 1 H NMR (600MHz, DMSO-d6) δ8.58(t,J=5.9Hz,1H),7.89(s,1H),7.59–7.55(m,2H),7.50(dd,J=8.1,2.1Hz,1H),7.47–7.43(m,2H),7.37(s,1H),7.30 (t,J=7.9Hz,3H),7.24(dd,J=16.3,7.5Hz,3H),7.17(d,J=7.6Hz,1H),6. 98(s,1H),5.31(s,2H),4.29(d,J=5.8Hz,2H),3.65(s,2H),2.35(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.41,139.96,139.36,134.32,131.12,130.89,129.31,129.15,129.02,128.81,127.70,12 7.24,125.62,125.39,123.28,122.67,122.57,113.03,112.86,111.94,49.02,42.76,32.58,12.77.ESI-HRMS:calcd for C 28 H 26N4OBr,[M+H] + ,513.1290; found 513.1293.

[0123] Example 8 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylbenzyl)acetamide (NFI8)

[0124] When 4-hydroxybenzylamine was replaced with 4-methylbenzylamine and indole-3-ethyl acetate was replaced with 5-bromo-indole-3-ethyl acetate in the above preparation process, NFI8 was obtained; a yellow solid with a yield of 79.45%.

[0125] 1 H NMR (600MHz, DMSO-d6) δ8.53(t,J=5.8Hz,1H),7.88(d,J=1.7Hz,1H),7.57( dd,J=15.5,7.7Hz,2H),7.52–7.48(m,1H),7.46(d,J=8.8Hz,1H),7.39(s,1H ),7.35–7.30(m,2H),7.18(d,J=7.6Hz,1H),7.16–7.09(m,4H),7.02(s,1H) ,5.32(s,2H),4.24(d,J=5.9Hz,2H),3.63(s,2H),2.37(s,3H),2.26(s,3H). 13 C NMR (150MHz, DMSO-d6) δ170.32,139.37,139.30,136.90,134.31,131.12,130.91,129.64,129.29,128.78,127.70,125.6 5,125.39,124.92,123.31,122.72,122.57,121.36,113.02,112.85,49.08,42.52,32.58,21.14,12.67.ESI-HRMS:calcd for C 29 H 28 N4OBr,[M+H] + ,527.1446; found 527.1451.

[0126] Example 9 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methoxybenzyl)acetamide (NFI9)

[0127] When 4-hydroxybenzylamine was replaced with 4-methoxybenzylamine and indole-3-ethyl acetate was replaced with 5-bromo-indole-3-ethyl acetate in the above preparation process, NFI9 was obtained; a yellow solid with a yield of 80.27%.

[0128] 1 H NMR (600MHz, DMSO-d6) δ8.52(t,J=5.9Hz,1H),7.87(d,J=1.5Hz,1H),7.58(t,J =7.8Hz,1H),7.55(s,1H),7.52(d,J=8.2Hz,1H),7.49–7.42(m,3H),7.31(dd,J =8.8,1.9Hz,1H),7.22(d,J=7.6Hz,1H),7.18(d,J=8.7Hz,3H),6.89–6.85(m,2 H),5.36(s,2H),4.22(d,J=5.8Hz,2H),3.72(s,3H),3.62(s,2H),2.44(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.24,158.67,144.76,139.40,134.29,131.87,131.13,130.96,129.08,128.77,125.82,125.4 0,123.48,122.93,122.58,121.72,114.16,113.04,112.87,112.02,55.53,49.32,42.23,32.57,12.27.ESI-HRMS:calcd for C 29 H 28 N4O2Br,[M+H] + ,543.1396; found543.1403.

[0129] Example 10 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxybenzyl)acetamide (NFI10)

[0130] Replacing ethyl indole-3-acetate in the above preparation process with ethyl 5-bromo-indole-3-acetate yields NFI10; a yellow solid with a yield of 76.14%.

[0131] 1H NMR (600MHz, DMSO-d6) δ9.30 (s, 1H), 8.45 (t, J = 5.8Hz, 1H), 7.88 (s, 1H), 7 .59–7.52(m,2H),7.50–7.47(m,1H),7.44(d,J=8.7Hz,1H),7.34–7.22(m, 3H),7.15(d,J=7.7Hz,1H),7.04(d,J=8.0Hz,2H),6.90(s,1H),6.68(d,J= 8.1Hz,2H),5.29(s,2H),4.16(d,J=5.7Hz,2H),3.61(s,2H),2.30(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.18,156.71,139.79,139.35,134.30,131.11,130.86,130.02,129.03,128.73,126.19,12 5.50,125.37,123.16,122.59,122.52,115.48,113.00,112.83,112.03,55.39,42.32,32.61,12.99.ESI-HRMS:calcd for C 28 H 26 N4O2Br,[M+H] + ,529.1239; found 529.1238.

[0132] Example 11: N-(2-(1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI11)

[0133] When 4-hydroxybenzylamine is replaced with benzoylhydrazine in the above preparation process, NFI11 is obtained; a white solid with a yield of 74.48%.

[0134] 1 H NMR (600MHz, DMSO-d6) δ10.37(s,1H),10.19(s,1H),7.88(d,J=7.3Hz,2H),7.77(d,J=7.8Hz,1H),7.59(s,1H),7.57(t,J=7.7Hz,2H) ,7.50(dd,J=8.1,4.1Hz,4H),7.34(s,1H),7.25–7.18(m,2H),7.18–7.10(m,2H),6.81(s,1H),5.28(s,2H),3.72(s,2H),2.28(s,3H). 13C NMR (150MHz, DMSO-d6) δ168.93,138.94,138.76,131.86,131.19,129.73,128.17,127.84,126.82,126.22, 125.89,123.97,123.86,121.98,121.86,121.18,119.44,119.18,110.58,109.59,29.80.ESI-HRMS:calcd for C 28 H 26 N5O2,[M+H] + ,464.5410; found464.2089.

[0135] Example 12 4-Methyl-N-(2-(1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI12)

[0136] When 4-hydroxybenzylamine is replaced with 4-methylbenzoylhydrazine in the above preparation process, NFI12 is obtained; a white solid with a yield of 76.67%.

[0137] 1 H NMR (600MHz, DMSO-d6) δ10.30(s,1H),10.17(s,1H),7.78(d,J=8.2Hz,2H),7.73(s,1H),7.64–7.61(m,2H),7.60–7.56(m,3H),7.55( d,J=4.0Hz,2H),7.29(d,J=8.7Hz,3H),7.24–7.21(m,1H),7.17(t,J=7.4Hz,1H),5.48(s,2H),3.72(s,2H),2.61(s,3H),2.36(s,3H). 13 C NMR(150MHz,DMSO-d6)δ168.91,141.19,136.18,134.36,130.00,129.06,128.36,128.25,126.84,126.26,124.73,12 2.68,122.16,122.02,121.53,119.53,119.20,118.60,110.71,109.67,50.02,29.73,19.29,10.71.ESI-HRMS:calcd forC 29 H 28 N5O2,[M+H] + ,478.5680; found 478.2247.

[0138] Example 13 4-Methoxy-N-(2-(1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI13)

[0139] When 4-hydroxybenzylamine was replaced with 4-methoxybenzoylhydrazine in the above preparation process, NFI13 was obtained; a white solid with a yield of 79.48%.

[0140] 1 H NMR (600MHz, DMSO-d6) δ10.23 (s, 1H), 10.14 (s, 1H), 7.86 (d, J = 8.9Hz, 2H), 7.7 8(d,J=2.4Hz,1H),7.62(dd,J=4.8,2.5Hz,3H),7.61(d,J=3.1Hz,2H),7.58(t, J=8.1Hz,2H),7.31(d,J=7.3Hz,1H),7.23(t,J=7.0Hz,1H),7.17(t,J=7.4Hz,1 H),7.02(d,J=8.9Hz,2H),5.49(s,2H),3.82(s,3H),3.71(s,2H),2.65(s,3H). 13 C NMR (150MHz, DMSO-d6) δ168.95,163.59,160.31,143.96,139.38,136.44,134.36,130.65,128.72,128.25,126.25,124.81,123.9 9,122.76,122.25,122.03,121.69,119.54,119.21,117.37,112.21110.11,109.68,55.52,48.31,27.53,10.07.ESI-HRMS:calcd for C 29 H 28 N5O3,[M+H] + ,494.2114; found 494.2196.

[0141] Example 14 4-Hydroxy-N-(2-(1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI14)

[0142] When 4-hydroxybenzylamine is replaced with 4-hydroxybenzoylhydrazine in the above preparation process, NFI14 is obtained; a white solid with a yield of 73.47%.

[0143] 1 H NMR(600MHz,DMSO-d6)δ10.10(d,J=10.0Hz,3H),7.76(t,J=8.4Hz,3H),7.62–7.57(m,2H),7.55–7.50 (m,2H),7.44(s,2H),7.24–7.11(m,4H),6.81(d,J=8.7Hz,2H),5.37(s,2H),3.70(s,2H),2.43(s,3H). 13 C NMR(150MHz,DMSO-d6)δ171.42,165.15,160.00,144.25,139.63,137.69,135.07,130.37,128.84,127.50,126.22,124.29,12 2.46,122.21,121.99,121.61,121.03,120.58,119.46,119.20,114.33,110.71,109.61,53.47,29.77,12.33.ESI-HRMS:calcd for C 28 H 26 N5O3,[M+H] + ,480.1957; found 480.2040.

[0144] Example 15: Benzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI15)

[0145] Intermediate 6a (0.50 g, 1.42 mmol) was dissolved in 5 mL of anhydrous dimethylformamide (DMF) at room temperature. EDCI (0.33 g, 1.70 mmol) and DMAP (0.21 g, 1.70 mmol) were added, and the mixture was stirred for 30 minutes. Then, benzyl alcohol (0.17 g, 1.56 mmol) was added, and the mixture was stirred for another hour. The reaction was monitored by thin-layer chromatography (TLC). After the reaction was complete, 30 mL of water was added to quench the reaction, followed by extraction with ethyl acetate (10 mL × 3). The combined organic layers were washed with saturated brine (10 mL × 2) and concentrated under reduced pressure. The crude product was purified by column chromatography (eluent: dichloromethane / methanol = 40:1) to give a yellow oil NFI15 (0.47 g), with a yield of 74.28%.

[0146] 1H NMR (600MHz, DMSO-d6) δ7.62(d,J=7.9Hz,1H),7.56(s,1H),7.54(t,J=7.8Hz,1H),7.51(d,J=8.3Hz,1H),7.46(dd,J=7.7,2.1Hz,1H),7.3 8–7.31(m,6H),7.31–7.23(m,1H),7.23–7.19(m,1H),7.15–7.09(m,2 H),6.87(s,1H),5.26(s,2H),5.16(s,2H),3.92(s,2H),2.26(s,3H). 13 C NMR(150MHz,DMSO-d6)δ171.53,140.02,139.71,136.62,135.53,130.75,129.13,128.88,128.50,128.46,12 7.49,125.11,123.14,122.37,120.64,120.02,110.80,110.38,66.29,48.87,30.91,13.28.ESI-HRMS:calcd for C 28 H 26 N3O2, [M+H] + ,436.1947; found 436.2029.

[0147] The compounds NFI16 and NFI17 were synthesized using the same method.

[0148] Example 16: 4-Methylbenzyl-2-(1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI16)

[0149] When benzyl alcohol was replaced with 4-methylbenzyl alcohol in the preparation process of Example 15 above, NFI16 was obtained; a yellow solid with a yield of 74.34%.

[0150] 1H NMR (600MHz, DMSO-d6) δ7.60(d,J=7.9Hz,1H),7.54(d,J=8.1Hz,2H),7.50(d,J= 8.3Hz,1H),7.47(d,J=1.9Hz,1H),7.33(s,1H),7.25(d,J=7.7Hz,2H),7.20(t,J =7.7Hz,1H),7.15(d,J=7.7Hz,2H),7.13(d,J=7.5Hz,1H),7.11(d,J=7.8Hz,2H) ,6.87(s,1H),5.27(s,2H),5.10(s,2H),3.89(s,2H),2.29(s,3H),2.25(s,3H). 13 C NMR(150MHz,DMSO-d6)δ171.51,140.02,139.70,137.82,135.51,133.59,130.75,129.11,128.62,127.45,125.1 1,123.12,123.00,122.37,120.62,120.00,110.78,110.39,66.21,55.38,30.92,21.22,14.55.ESI-HRMS:calcd for C 29 H 28 N3O2, [M+H] + ,450.2182; found 450.2189.

[0151] Example 17 4-Chlorobenzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI17)

[0152] When benzyl alcohol was replaced with 4-chlorobenzyl alcohol in the preparation process of Example 15 above, NFI17 was obtained; a yellow solid with a yield of 65.49%.

[0153] 1 H NMR (600MHz, DMSO-d6) δ7.59(d,J=7.9Hz,1H),7.57–7.52(m,2H),7.51–7.46(m,2H),7.41(q,J=8.5Hz,4H),7.32(d,J=2.2Hz ,1H),7.21(ddd,J=8.2,7.0,1.2Hz,2H),7.15–7.09(m,2H),6.82(s,1H),5.27(s,2H),5.14(s,2H),3.91(s,2H),2.26(s,3H). 13C NMR(150MHz,DMSO-d6)δ171.46,139.68,135.68,135.51,133.12,130.77,130.34,129.08,128.87,127.50,12 5.13,123.14,123.02,122.36,120.63,119.99,110.79,110.29,65.46,55.39,30.85,13.26.ESI-HRMS:calcd for C 28 H 25 N3O2Cl,[M+H] + ,470.1635; found470.1637.

[0154] Example 18 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenylacetamide (NFI18)

[0155] When 4-hydroxybenzylamine was replaced with aniline in the preparation process of Example 1 above, NFI18 was obtained; a white solid with a yield of 62.37%.

[0156] 1 H NMR(600MHz,DMSO-d6)δ10.19(s,1H),7.72(d,J=7.8Hz,1H),7.62(s,1H),7.61(s,2 H),7.58(d,J=7.6Hz,1H),7.57(d,J=4.3Hz,2H),7.54(d,J=8.8Hz,1H),7.48(s,1H) ,7.40(ddt,J=31.9,16.9,8.0Hz,1H),7.30(t,J=7.7Hz,2H),7.21(t,J=8.0Hz,2H), 7.15(t,J=7.4Hz,1H),7.04(t,J=7.4Hz,1H),5.40(s,2H),3.83(s,2H),2.46(s,3H). 13 CNMR(150MHz,DMSO-d6)δ138.82,138.67,137.21,129.86,128.29,128.10,126.23,124.44,122.56,12 2.41,122.00,121.84,119.56,119.01,118.52,110.94,109.73,48.66,32.69,11.08.ESI-HRMS:calcd for C 27 H 25 N4O, [M+H] +,421.2028; found 421.2032.

[0157] Example 19: N-(4-methoxyphenyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI19)

[0158] When 4-hydroxybenzylamine was replaced with 4-methoxyaniline in the preparation process of Example 1 above, NFI19 was obtained; a white solid with a yield of 74.57%.

[0159] 1 H NMR (600MHz, DMSO-d6) δ10.05(s,1H),7.72(d,J=7.8Hz,1H),7.60–7.50(m,7H),7.46(s,1H),7.35(s,1H),7.21(t ,J=7.6Hz,2H),7.15(t,J=7.4Hz,1H),6.87(d,J=8.7Hz,2H),5.38(s,2H),3.79(s,2H),3.71(s,3H),2.43(s,3H). 13 C NMR(150MHz,DMSO-d6)δ169.07,155.62,139.90,138.59,135.48,130.91,129.37,127.26,125.42,123.39,12 3.06,122.80,121.16,120.61,120.11,114.30,112.15,110.79,55.62,49.57,33.65,12.31.ESI-HRMS:calcd for C 28 H 27 N4O2,[M+H] + ,451.2134; found 451.2133.

[0160] Example 20 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenethylacetamide (NFI20)

[0161] When 4-hydroxybenzylamine was replaced with phenethylamine in the preparation process of Example 1 above, NFI20 was obtained; a white solid with a yield of 85.97%.

[0162] 1H NMR (600MHz, DMSO-d6) δ8.06(t,J=5.7Hz,1H),7.63–7.55(m,2H),7.54–7.49(m,2H),7.44(s,1H),7.41(s,2H),7.25–7.21(m,2H),7.19(t,J=6.8H z,2H),7.17–7.14(m,3H),7.12(d,J=7.4Hz,1H),7.09(s,1H),5.35(s,2H ),3.57(s,2H),3.30(q,J=6.8Hz,2H),2.71(t,J=7.3Hz,2H),2.40(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.49,139.92,138.94,135.45,130.86,129.35,129.14,129.11,128.71,127.16,126.49,125.27,1 24.22,123.25,122.99,122.58,121.54,120.50,120.09,112.31,110.70,49.24,40.81,35.57,32.79,12.49.ESI-HRMS:calcd for C 29 H 29 N4O, [M+H] + ,449.2341; found 449.2339.

[0163] Example 21: N-(4-methoxyphenethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI21)

[0164] When 4-hydroxybenzylamine was replaced with 4-methoxyphenylethylamine in the preparation process of Example 1 above, NFI21 was obtained; a white solid with a yield of 81.21%.

[0165] 1H NMR(600MHz,DMSO-d6)δ8.03(t,J=5.7Hz,1H),7.72–7.57(m,3H),7.57–7.53(m,2H), 7.51(s,1H),7.47(s,1H),7.40(h,J=9.7,9.1Hz,1H),7.24(d,J=7.6Hz,1H),7.23–7.1 9(m,1H),7.14(t,J=7.4Hz,1H),7.09–7.04(m,2H),6.78(d,J=8.5Hz,2H),5.43(s,2H) ,3.68(s,3H),3.57(s,2H),3.26(q,J=6.8Hz,2H),2.64(t,J=7.2Hz,2H),2.52(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.44,158.09,139.98,137.85,135.44,131.77,130.97,130.19,130.05,129.38,127.19,125.56,1 23.54,123.01,122.96,120.52,120.12,114.53,114.14,112.39,110.73,55.40,49.81,41.06,34.68,32.76.ESI-HRMS:calcd forC 30 H 31 N4O2,[M+H] + ,479.2447; found 479.2441.

[0166] Example 22 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylphenethyl)acetamide (NFI22)

[0167] When 4-hydroxybenzylamine was replaced with 4-methylphenethylamine in the preparation process of Example 1 above, NFI22 was obtained; a white solid with a yield of 90.31%.

[0168] 1H NMR(600MHz,DMSO-d6)δ8.04(t,J=5.7Hz,1H),7.61(dd,J=7.8,6.3Hz,2H),7 .58(d,J=5.3Hz,1H),7.54(d,J=8.2Hz,2H),7.50(s,1H),7.46(s,1H),7.33( s,1H),7.25–7.19(m,2H),7.15–7.13(m,1H),7.03(s,4H),5.41(s,2H),3.56 (s,2H),3.29–3.24(m,2H),2.66(t,J=7.3Hz,2H),2.51(s,3H),2.22(s,3H). 13 C NMR (150MHz, DMSO-d6) δ138.03,136.79,135.44,130.95,129.66,129.38,129.30,128.98,127.17,125.51,123.49,123. 00,122.91,122.10,121.84,120.51,120.11,112.38,110.72,49.64,40.90,35.17,32.75,21.08,11.88.ESI-HRMS:calcd for C 30 H 31 N4O, [M+H] + ,463.2498; found 463.2498.

[0169] Example 23: N-(4-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI23)

[0170] When 4-hydroxybenzylamine was replaced with 4-hydroxyphenylethylamine in the preparation process of Example 1 above, NFI23 was obtained; a white solid with a yield of 90.01%.

[0171] 1H NMR(600MHz,DMSO-d6)δ9.19(s,1H),7.99(t,J=5.7Hz,1H),7.61(d,J=7.8Hz,1H),7.56 (t,J=7.8Hz,1H),7.51–7.47(m,2H),7.45(s,1H),7.32(s,1H),7.26(s,1H),7.19(t,J= 7.6Hz,1H),7.16–7.10(m,2H),6.94(d,J=8.1Hz,2H),6.86(s,1H),6.63(d,J=8.1Hz,2H ),5.29(s,2H),3.56(s,2H),3.22(q,J=6.8Hz,2H),2.59(t,J=7.3Hz,2H),2.29(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.41,156.08,139.86,139.82,135.45,130.78,129.94,127.16,124.98,122.97,1 22.96,122.23,120.46,120.12,115.52,112.30,110.67,55.39,41.19,34.79,32.81,13.11.ESI-HRMS:calcd for C 29 H 29 N4O2,[M+H] + ,465.2291; found 465.2295.

[0172] Example 24 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(3-phenylpropyl)acetamide (NFI24)

[0173] When 4-hydroxybenzylamine was replaced with amphetamine in the preparation process of Example 1 above, NFI24 was obtained; a white solid with a yield of 74.11%.

[0174] 1H NMR (600MHz, DMSO-d6) δ8.07(t,J=5.6Hz,1H),7.68(d,J=7.9Hz,1H),7.57(t,J=7.8H z,1H),7.52(dd,J=7.9,2.5Hz,2H),7.50(s,1H),7.48–7.43(m,2H),7.25(t,J=7.5Hz, 2H),7.21(dd,J=13.3,6.9Hz,3H),7.14(dd,J=7.7,4.3Hz,4H),5.36(s,2H),3.59(s, 2H), 3.09 (q, J = 6.6Hz, 2H), 2.56 (t, J = 7.8Hz, 2H), 2.44 (s, 3H), 1.70 (p, J = 7.2Hz, 2H). 13 C NMR (150MHz, DMSO-d6) δ170.47,144.73,142.19,139.94,138.61,135.48,130.89,128.93,128.73,127.15,126.56,126. 18,125.35,123.31,123.01,122.70,121.74,120.50,110.75,49.38,38.79,33.00,32.85,31.45,12.26.ESI-HRMS:calcd for C 30 H 31 N4O, [M+H] + ,463.2498; found 463.2495.

[0175] Example 25 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI25)

[0176] When 4-hydroxybenzylamine was replaced with 4-hydroxyphenylethylamine and indole-3-ethyl acetate was replaced with 5-bromo-indole-3-ethyl acetate in the preparation process of Example 1 above, NFI25 was obtained; a yellow solid with a yield of 73.24%.

[0177] 1H NMR(600MHz,DMSO-d6)δ9.20(s,1H),8.06(t,J=5.8Hz,1H),7.90–7.81(m,1H),7.56(t, J=7.8Hz,1H),7.51–7.45(m,2H),7.44(d,J=8.8Hz,1H),7.31(dd,J=8.2,1.9Hz,2H),7.2 1(s,1H),7.13(d,J=7.6Hz,1H),6.94(d,J=8.0Hz,2H),6.81(s,1H),6.63(d,J=8.0Hz,2 H),5.27(s,2H),3.55(s,2H),3.22(q,J=6.8Hz,2H),2.59(t,J=7.3Hz,2H),2.27(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.23,156.09,144.54,140.09,139.34,134.27,131.15,130.84,129.94,129.92,128.68,126.96,1 25.40,125.36,123.08,122.54,122.39,115.54,113.00,112.81,111.96,48.73,41.25,34.84,32.60,13.18.ESI-HRMS:calcd forC 29 H 28 N4O2Br,[M+H] + ,543.1396; found 543.1396.

[0178] Example 26 2-(5-chloro-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI26)

[0179] When 4-hydroxybenzylamine was replaced with 4-hydroxyphenylethylamine and indole-3-ethyl acetate was replaced with 5-chloro-indole-3-ethyl acetate in the preparation process of Example 1 above, NFI26 was obtained; a yellow solid with a yield of 74.24%.

[0180] 1H NMR (600MHz, DMSO-d6) δ9.32 (s, 1H), 8.09 (t, J = 5.6Hz, 1H), 7.74 (d, J = 1.9 Hz,1H),7.61(s,1H),7.59(d,J=7.7Hz,1H),7.55–7.50(m,3H),7.46(s,1H) ,7.43–7.14(m,3H),6.95(d,J=8.4Hz,2H),6.64(d,J=8.4Hz,2H),5.42(s,2 H), 3.56 (s, 2H), 3.23 (q, J = 6.7Hz, 2H), 2.59 (t, J = 7.3Hz, 2H), 2.43 (s, 3H). 13 C NMR(150MHz,DMSO-d6)δ156.09,138.18,133.99,131.00,130.57,129.93,128.87,123.60,12 3.06,122.85,119.50,115.54,112.44,112.12,41.26,40.53,34.84,32.56.ESI-HRMS:calcd for C 29 H 28 N4O2Cl,[M+H] + ,499.1901; found 499.1905.

[0181] Example 27 2-(5-fluoro-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI27)

[0182] When 4-hydroxybenzylamine was replaced with 4-hydroxyphenylethylamine and indole-3-ethyl acetate was replaced with 5-fluoro-indole-3-ethyl acetate in the preparation process of Example 1 above, NFI27 was obtained; a yellow solid with a yield of 73.47%.

[0183] 1H NMR (600MHz, DMSO-d6) δ9.15 (s, 1H), 8.07 (t, J = 5.6Hz, 1H), 7.67 (s, 1H), 7.60 (t ,J=7.7Hz,1H),7.54(dd,J=9.3,5.3Hz,4H),7.47(s,1H),7.45–7.41(m,1H),7.28 (d,J=7.6Hz,1H),7.06–7.03(m,1H),6.95(d,J=8.4Hz,2H),6.63(d,J=8.4Hz,2H ),5.44(s,2H),3.54(s,2H),3.22(q,J=6.7Hz,2H),2.58(dd,J=13.6,6.2Hz,5H). 13 C NMR(150MHz,DMSO-d6)δ170.23,157.06,137.55,132.16,130.09,129.93,129.04,125.92,123.69,123.19,115.5 2,112.45,112.42,112.08,112.02,111.06,110.89,105.15,105.00,41.24,40.53,34.82,32.65.ESI-HRMS:calcd for C 29 H 28 N4O2F, [M+H] + ,483.2196; found483.2200.

[0184] Example 28: N-(2-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI28)

[0185] When 4-hydroxybenzylamine was replaced with 2-hydroxyphenylethylamine in the preparation process of Example 1 above, NFI28 was obtained; a white solid with a yield of 73.78%.

[0186] 1H NMR (600MHz, DMSO-d6) δ9.33 (s, 1H), 8.05 (t, J = 5.7Hz, 1H), 7.64 (d, J = 7.8Hz, 1H), 7.58(t,J=7.8Hz,1H),7.54–7.49(m,2H),7.44(d,J=19.1Hz,3H),7.22–7.17(m,2H) ,7.15–7.07(m,2H),7.00–6.94(m,2H),6.77(d,J=9.1Hz,1H),6.64(t,J=7.4Hz,1H ),5.36(s,2H),3.56(s,2H),3.28–3.24(m,2H),2.66(t,J=7.4Hz,2H),2.41(s,3H). 13 C NMR (150MHz, DMSO-d6) δ170.43,153.88,138.85,137.72,134.78,130.51,129.79,129.60,128.29,126.56,126.08,124.81,124.20,122.2 0,121.90,121.56,119.78,119.41,119.04,118.53,118.18,114.24,111.78,109.61,47.53,38.21,32.77,29.39,11.35.ESI-HRMS:calcd for C 29 H 29 N4O2,[M+H] + ,465.2212; found 465.2294.

[0187] Example 29: N-(3-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI29)

[0188] When 4-hydroxybenzylamine was replaced with 3-hydroxyphenylethylamine in the preparation process of Example 1 above, NFI29 was obtained; a white solid with a yield of 68.44%.

[0189] 1H NMR (600MHz, DMSO-d6) δ9.28(s,1H),8.06(t,J=5.7Hz,1H),7.61(d,J=7.8Hz,1H),7.56(t,J =7.8Hz,1H),7.50(d,J=8.0Hz,2H),7.44(s,1H),7.35(s,1H),7.29(s,1H),7.19(t,J=7.6Hz ,1H),7.16–7.10(m,2H),7.02(t,J=7.7Hz,1H),6.91(s,1H),6.60(s,1H),6.58(dd,J=8.2,2 .5Hz,2H),5.30(s,2H),3.56(s,2H),3.28–3.24(m,2H),2.62(t,J=7.4Hz,2H),2.32(s,3H). 13 C NMR (150MHz, DMSO-d6) δ169.37,157.95,143.49,140.19,138.79,138.51,134.36,129.72,128.58,128.26,126.06,124.86,123.98,121.9 6,121.89,121.27,120.05,119.41,119.02,118.63,114.84,112.44,111.20,109.59,47.86,40.80,34.61,31.20,10.82.ESI-HRMS:calcd for C 29 H 29 N4O2,[M+H] + ,465.2212; found 465.2293.

[0190] Example 30: N-(3,4-dihydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI30)

[0191] When 4-hydroxybenzylamine was replaced with 3,4-dihydroxyphenylethylamine in the preparation process of Example 15 above, NFI30 was obtained; a white solid with a yield of 73.47%.

[0192] 1H NMR (600MHz, DMSO-d6) δ8.69(d,J=49.0Hz,2H),8.05(t,J=5.7Hz,1H),7.74(d,J=2.1Hz ,1H),7.61(dd,J=8.7,7.4Hz,2H),7.59–7.53(m,4H),7.49(s,1H),7.29(d,J=9.1Hz,1H) ,7.21(t,J=7.7Hz,1H),7.14(t,J=7.0Hz,1H),6.62–6.56(m,2H),6.40(dd,J=8.0,2.4H z,1H),5.48(s,2H),3.57(s,2H),3.21(q,J=6.9Hz,2H),2.62(s,3H),2.55–2.52(m,2H). 13 C NMR(150MHz,DMSO-d6)δ169.28,144.46,143.92,142.91,138.94,136.01,134.35,129.98,129.56,128.33,126.10,124.68,122.71,122.1 7,121.93,121.60,119.97,119.06,118.61,118.27,115.72,114.81,111.35,109.66,47.87,40.14,34.05,31.63,10.85.ESI-HRMS:calcd for C 29 H 29 N4O3,[M+H] + ,481.2161; found 481.2243.

[0193] Example 31 2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenethylacetamide (NFI31)

[0194] When 4-hydroxybenzylamine was replaced with phenethylamine and 3-iodobenzyl bromide was replaced with 2-iodobenzyl bromide in the preparation process of Example 1 above, NFI31 was obtained; a white solid with a yield of 74.46%.

[0195] 1H NMR(600MHz,DMSO-d6)δ8.09(t,J=5.7Hz,1H),7.63–7.57(m,3H),7.40(ddd,J =12.5,7.5,1.8Hz,2H),7.29–7.25(m,3H),7.23(d,J=7.5Hz,2H),7.19–7.17( m,2H),7.15(d,J=7.3Hz,1H),7.13–7.10(m,2H),6.89–6.86(m,1H),5.06(d,J =21.9Hz,2H),3.58(s,2H),3.35–3.32(m,2H),2.74–2.72(m,2H),2.00(s,3H). 13 C NMR(150MHz,DMSO-d6)δ169.47,138.85,136.52,136.12,131.86,129.33,128.46,128.26,128.10,128.06,128.04,12 7.90,127.66,127.19,126.98,125.42,121.87,119.16,118.85,110.72,108.89,45.80,34.52,31.71.ESI-HRMS:calcd for C 29 H 29 N4O, [M+H] + ,449.5700; found 449.2344.

[0196] Example 32 2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylphenethyl)acetamide (NFI32)

[0197] When 4-hydroxybenzylamine was replaced with 4-methylphenethylamine and 3-iodobenzyl bromide was replaced with 2-iodobenzyl bromide in the preparation process of Example 1 above, NFI32 was obtained; a white solid with a yield of 87.24%.

[0198] 1H NMR(600MHz,DMSO-d6)δ8.06(t,J=5.7Hz,1H),7.64–7.55(m,3H),7.40(ddd,J=1 8.3,7.3,1.9Hz,2H),7.29(s,1H),7.14(s,1H),7.12(d,J=2.1Hz,1H),7.10(dd, J=6.4,4.5Hz,2H),7.05(d,J=4.1Hz,4H),6.90–6.86(m,1H),5.06(d,J=21.3Hz, 2H),3.57(s,2H),3.31(s,2H),2.68(t,J=7.3Hz,2H),2.23(s,3H),1.99(s,3H). 13 C NMR(150MHz,DMSO-d6)δ136.51,136.13,135.72,135.23,134.33,133.45,131.91,129.35,129.29,128.59,128.45,128.23,1 27.94,127.90,127.19,126.98,121.87,119.13,110.74,108.90,43.45,40.33,34.11,31.72,18.13,10.48.ESI-HRMS:calcd for C 30 H 31 N4O, [M+H] + ,463.5970; found 463.2502.

[0199] Example 33: N-(4-methoxyphenethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI33)

[0200] When 4-hydroxybenzylamine was replaced with 4-methoxyphenethylamine and 3-iodobenzyl bromide was replaced with 2-iodobenzyl bromide in the preparation process of Example 1 above, NFI33 was obtained; a white solid with a yield of 76.32%.

[0201] 1H NMR(600MHz,DMSO-d6)δ8.05(t,J=5.7Hz,1H),7.64–7.56(m,3H),7.41(td,J=7 .9,1.9Hz,2H),7.31(s,1H),7.16(d,J=6.7Hz,1H),7.13–7.10(m,2H),7.10–7.0 6(m,3H),6.89–6.86(m,1H),6.79(d,J=8.6Hz,2H),5.07(d,J=25.7Hz,2H),3.69 (s,3H),3.57(s,2H),3.28(q,J=6.8Hz,2H),2.66(t,J=7.3Hz,2H),2.00(s,3H). 13 C NMR(150MHz,DMSO-d6)δ170.49,158.10,137.61,137.20,132.86,131.77,130.48,130.45,130.06,129.54,129.34,1 28.08,122.94,120.22,119.95,114.14,111.83,109.97,55.42,46.93,41.07,34.71,32.79,10.94.ESI-HRMS:calcd for C 30 H 31 N4O2,[M+H] + ,479.2447; found 479.2437.

[0202] Example 34: N-(4-hydroxyphenylethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI34)

[0203] When 4-hydroxybenzylamine was replaced with 4-hydroxyphenylethylamine and 3-iodobenzyl bromide was replaced with 2-iodobenzyl bromide in the preparation process of Example 1 above, NFI34 was obtained; a white solid with a yield of 71.01%.

[0204] 1H NMR (600MHz, DMSO-d6) δ9.16 (s, 1H), 8.06 (t, J = 5.6Hz, 1H), 7.64–7.57 (m, 3H), 7. 44(ddd,J=21.3,7.4,1.9Hz,2H),7.32(s,1H),7.22(s,1H),7.11(dd,J=5.9,2.9H z,3H),6.96(d,J=8.4Hz,2H),6.88–6.82(m,1H),6.64(d,J=8.4Hz,2H),5.18–5.0 4(m,2H),3.57(s,2H),3.25(q,J=5.7Hz,2H),2.61(t,J=7.4Hz,2H),2.02(s,3H). 13 C NMR(150MHz,DMSO-d6)δ137.70,137.19,132.56,130.68,130.64,129.96,129.93,129.57,129.44,128.27,128.0 6,122.94,121.90,120.24,119.95,115.53,111.88,109.91,41.22,40.53,34.83,32.76,10.68.ESI-HRMS:calcd for C 29 H 29 N4O2,[M+H] + ,465.2291; found 465.2294.

[0205] Example 35: N-(4-fluorophenylethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI35)

[0206] When 4-hydroxybenzylamine was replaced with 4-fluorophenylethylamine and 3-iodobenzyl bromide was replaced with 2-iodobenzyl bromide in the preparation process of Example 1 above, NFI35 was obtained; a white solid with a yield of 79.55%.

[0207] 1H NMR(600MHz,DMSO)δ8.10(t,J=5.7Hz,1H),7.64–7.58(m,3H),7.51(dd,J=7.4,1 .9Hz,1H),7.42(dd,J=7.3,1.8Hz,1H),7.30(s,2H),7.21(dd,J=8.6,5.7Hz,3H), 7.10(dd,J=6.1,3.0Hz,2H),7.05(t,J=8.9Hz,2H),6.86–6.83(m,1H),5.21–5.0 6(m,2H),3.56(s,2H),3.33(d,J=6.5Hz,2H),2.73(t,J=7.1Hz,2H),2.03(s,3H). 13 C NMR (151MHz, DMSO) δ162.07,160.47,137.18,136.07,136.05,130.93,130.88,130.86,130.75,129.61,129.50,128 .25,128.02,122.95,120.24,119.90,115.45,115.31,109.89,47.37,40.78,34.66,32.73,10.49.ESI-HRMS:calcd for C 29 H 28 N4OF,[M+H] + ,467.2247; found 467.2252.

[0208] Pharmacological studies of the product of this invention

[0209] Pharmacological tests have demonstrated that the compounds involved in this invention possess anti-ischemic stroke activity.

[0210] Neuroprotective activity in PC12 cell line

[0211] The neuroprotective activity of compounds NFI1-NFI35 in the PC12 cell line was evaluated using the MTT assay. A brief procedure is as follows: PC12 cells (1×10⁻⁶ cells) were... 4 Cells (per well) were seeded in 96-well plates containing 100 μL of RPMI 1640 complete medium and incubated for 24 hours at 37°C with 5% CO2. After incubation, different concentrations (0.05, 0.5, and 5 μM) of compounds NFI1-NFI35 and control Ifenprodil were added to the cells, and each group was incubated for another 6 hours. Subsequently, the medium was replaced with HEPES buffer (1×HMF, MgSO4-free). 2+Cells were cultured with NMDA (500 μM) with or without the addition of product number CC0074 for 30 minutes. Afterward, the medium was replaced with fresh medium, and cells were incubated for another 12 hours. At the end of the treatment, 20 μL of MTT reagent (product number M-2128, Sigma) was added to each well, and the cells were incubated for another 4 hours. The medium was then removed, and 150 μL of DMSO was added to dissolve the formazan violet crystals. The absorbance was measured at 490 nm, and cell viability (expressed as a percentage) was calculated, as shown in Table 1. Control group was the blank control group (no cells added), Model group was the NMDA damage model group, and Ifenprodil group was the positive control group.

[0212] Table 1

[0213]

[0214]

[0215] Data are expressed as the mean ± standard deviation of three independent experiments.

[0216] As shown in Table 1, all the compounds of the general formula of this invention have a certain degree of neuroprotective activity, and most of the drugs have excellent neuroprotective activity, which is superior to the positive control drug Ifenprodil. Among the compounds in this series, NFI23 has the best neuroprotective activity.

[0217] Measurement of intracellular calcium ion concentration

[0218] PC12 cells (7.5 × 10⁻⁶) 4 Cells (number per plate) were seeded in 35mm glass-bottomed cell culture dishes and cultured at 37°C in a 5% CO2 incubator for 24 hours in 1640 medium containing 10% fetal bovine serum. Cells were then treated with or without Ifenprodil or the aforementioned compound NFI23 (5μM) for 6 hours. After treatment, the medium was removed, and fresh 1640 medium containing Fluo-4 AM (2.5μM, S1060) was added, followed by incubation for another 30 minutes. The medium was then removed, and the cells were rinsed with HEPES buffer (1×HMF, Mg2+-free). 2+ Wash twice with product number CC0074. After adding 1 mL of HEPES buffer, scan for 30 seconds using a laser scanning confocal microscope (LSM 710, Zeiss, Germany) until the baseline fluorescence signal stabilizes. Then, add NMDA (final concentration: 500 μM) or an equal volume of HEPES buffer and continue scanning for 5 minutes to monitor changes in fluorescence intensity. Figure 1 As shown.

[0219] Depend on Figure 1It can be seen that, compared with the control group, the model group showed lower intracellular Ca2+ levels after NMDA treatment. 2+ The concentration increased significantly. Pretreatment with NFI23 significantly reduced intracellular Ca2+. 2+ It exhibits a similar effect to Ifenprodil. As shown in B, NFI23 can reduce Ca to a greater extent compared to Ifenprodil. 2+ Inflow. Therefore, the results indicate that NFI23 can significantly inhibit NMDA-induced Ca2+ flow. 2+ internal flow.

[0220] Measurement of intracellular ROS generation

[0221] Intracellular ROS levels were measured using the fluorescent probe 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA). The simplified procedure is as follows: PC12 cells were seeded in 6-well plates (2 × 10⁻⁶ cells / well). 5 Cells were cultured in wells (number cells / well) for 24 hours, followed immediately by the addition of 10 μL of Ifenprodil or NFI23 (5 μM). After 6 hours of pretreatment with the test compound, cell damage was induced using NMDA. Following treatment, cells were collected and incubated at 37°C for 20 minutes in the dark with DCFH-DA working solution (1:1000 diluted in serum-free 1640 medium). After incubation, cells were washed three times with PBS to remove excess extracellular DCFH-DA. The washed cells were resuspended in PBS, and the fluorescence intensity of dichlorofluorescein (DCF) was analyzed by flow cytometry to quantify ROS levels, such as... Figure 2 As shown.

[0222] Depend on Figure 2 As can be seen, in sample A, the level of reactive oxygen species (ROS) in the NMDA group was significantly increased after NMDA treatment, while Ifenprodil and NFI23 were able to inhibit this increase. NFI23 was able to inhibit NMDA-induced excessive ROS production. In sample B, it can be seen that NFI23 and Ifenprodil have comparable effects. Therefore, the results demonstrate that NFI23 can inhibit NMDA-induced excessive ROS production.

[0223] Measurement of MMP (mitochondrial membrane potential)

[0224] Mitochondrial membrane potential (MMP) was assessed using the fluorescent probe JC-1. Cell culture was performed using the same methods as the ROS assay. After the same pretreatment, PC12 cells were collected, washed once with PBS, and incubated with a working solution of 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazole carbonyl cyanine iodide (JC-1) at 37°C in the dark for 20 minutes. During incubation, the cell suspension was gently mixed every 5 minutes to ensure uniform staining. After staining, the cells were washed twice with PBS to remove excess dye, resuspended in PBS, and immediately analyzed by flow cytometry. JC-1 fluoresced green in monomeric form, indicating low MMP, and red in aggregate form, indicating high MMP. The mitochondrial membrane potential was quantified by calculating the red-green fluorescence ratio, such as... Figure 3 As shown.

[0225] Depend on Figure 3 As can be seen, in Figure A, cells in the upper right quadrant (Q2) of the JC-1 fluorescence dot plot represent healthy mitochondria with preserved MMPs, while cells in the lower right quadrant (Q3) represent mitochondrial depolarization. NMDA stimulation significantly increased the proportion of cells in the Q3 region, indicating MMP loss, and pretreatment with NFI23 effectively mitigated this phenomenon and preserved MMPs. Quantitative results in Figure B show that NFI23 has a slightly better protective effect than Ifenprodil. Therefore, the results indicate that NFI23 has a protective effect against NMDA-induced mitochondrial dysfunction, possibly by preventing mitochondrial depolarization.

[0226] Assay for apoptosis

[0227] Apoptosis analysis was performed using Annexin V-FITC / propidium iodide (PI) staining and detected by flow cytometry. PC12 cells were treated using the same method as described above, collected, and washed twice with ice-cold PBS. The cell pellet was resuspended in 100 μL binding buffer, followed by the addition of 5 μL Annexin V-FITC and 5 μL PI. The mixture was gently vortexed and incubated at room temperature in the dark for 15 minutes. After incubation, 400 μL binding buffer was added to each sample, and the samples were immediately analyzed by flow cytometry. Early apoptotic cells were Annexin V positive and PI negative, while late apoptotic or necrotic cells were double-positive for Annexin V and PI. Figure 4 As shown.

[0228] Depend on Figure 4As can be seen, after NMDA stimulation in sample A, a significant increase in cells was observed in the Q3 and Q2 regions, corresponding to early and late apoptosis, respectively. Compared with the control group, NFI23 pretreatment effectively reduced cell apoptosis. Quantitative analysis in sample B showed that NFI23 had an effect comparable to Ifenprodil. Therefore, the results indicate that NFI23 has a significant neuroprotective effect and can significantly inhibit NMDA-induced apoptosis of PC12 cells.

[0229] Western blot analysis

[0230] After culturing PC12 cells for 24 hours, they were treated with NFI23 at different concentrations (0.05 μM, 0.5 μM, and 5 μM) prepared in 10 μL DMSO for 6 hours. Subsequently, the medium was replaced with HEPES buffer (with or without NMDA, 500 μM), and the cells were treated for 30 minutes, followed by replacement with fresh medium and incubation for another 6 hours. Cells were then collected for protein extraction. Total cellular protein was extracted using RIPA lysis buffer, and protein concentration was determined using the BCA method. Equal volumes of protein samples were separated by SDS-PAGE under reducing conditions and transferred to a PVDF membrane. After blocking, the membrane was incubated with primary antibodies against p-ERK1 / 2 (Cell Signaling Technology, 4370S, USA) and ERK1 / 2 (Cell Signaling Technology, 4695S, USA). After washing, secondary antibody was added to the membrane. Protein bands were detected using ECL assay reagents, and images were acquired using a Tanon imaging system. The intensity of the protein bands was quantitatively analyzed using ImageJ software. Figure 5 As shown.

[0231] Depend on Figure 5 It is evident that NFI23 may exert a neuroprotective effect by activating the ERK1 / 2 pathway and counteracting GluN2B-mediated NMDAR signaling, thereby reducing excitotoxicity.

[0232] hERG channel inhibition experiment

[0233] This experiment used HEK293 cells stably expressing the hERG(NM_000238) gene. After removing the waste culture medium from the 6cm T-flask (cell density 30–70%), the cells were gently washed with 1 mL of PBS. Then, 1 mL of LTrypLe was added. TM Express, gently shake the culture flask to detach the cells, and incubate at 37°C for 2–3 minutes. After collecting the cells by gently tapping the culture flask, distribute them at 3 × 10⁶ cells per well. 4Cells were seeded at a density of [number] cells per culture dish with coverslips for subsequent analysis. The external solution consisted of (mM): 137NaCl, 4KCl, 2CaCl2, 1MgCl2, 10g glucose, and 10HEPES (pH 7.4); the internal solution consisted of (mM): 20KCl, 120K-aspartate, 10HEPES, 10EGTA, and 5Mg-ATP (pH 7.4). Before whole-cell patch-clamp recording, glass electrodes were fabricated using a Sutter P-1000 needle puller and brought close to the target cells under a 10× objective. Once contact was made and a resistance rise to 1 / 3 to 2 / 3 of the original was detected, a gigaseal was established. A short pulse was then applied to rupture the membrane, establishing whole-cell recording mode. hERG current recordings were performed using a holding potential of -80 mV, followed by membrane depolarization to +30 mV for 4 seconds to measure the tail current. A 15-second interval was maintained between each recording to ensure sufficient recovery of the current from the inactivated state. Current recordings were performed using an EPC-10 amplifier and PatchMaster (HEKA) software. After a stable baseline current was recorded, drug administration was initiated at the concentrations shown in Table 2. The percentage of current inhibition was calculated using the following formula: Tail current inhibition rate (%) = [1 - (Tail current after compound addition / Tail current under blank buffer conditions)] × 100. The dose-response curves of the test compounds were finally plotted using GraphPad Prism 8.0 software, as shown in Table 2.

[0234] Table 2

[0235]

[0236] a Data are expressed as mean ± standard deviation of three independent experiments.

[0237] As shown in Table 2, NFI23 has a low risk of arrhythmia and good cardiac safety, supporting further research.

[0238] Radioligand binding experiment

[0239] The cerebral cortex of male Wistar rats (175±25g) was used to prepare glutamate NMDA polyamine receptors using Tris-HCl buffer (pH 7.4). The test compound (NFI23) was diluted to final concentrations of 0.3, 1, 3, 10, 30, and 100 nM, and then mixed with 2 nM […]. 3H]Ifenprodil and 5 μM GBR 12909 were co-incubated to block non-polyamine-sensitive binding sites. Incubation conditions were 4 °C for 120 minutes. GF / B filter pads were pre-soaked in 0.5% polyethyleneimine (PEI) solution for 2 hours at room temperature. After incubation, the samples were filtered and washed five times with 300 μL of ice-cold distilled water. The filter pads were then dried at 95 °C. Solid scintillator was coated onto the dried filter pads and heated at 95 °C for 5 minutes, then cooled and cured at room temperature. Finally, radioactivity was measured to determine […]. 3 Specific binding of H]Ifenprodil. Non-specific binding was determined by means of 10 μM of label-free Ifenprodil, as shown in Table 3.

[0240] Table 3

[0241]

[0242] As can be seen from Table 3, NFI23 and [ 3 H]Ifenprodil effectively competes for membrane receptor binding, indicating a high binding affinity for GluN2B-NMDAR.

[0243] Patch clamp experiment

[0244] Whole-cell patch-clamp recordings were performed using HEK293 cells stably expressing GluN1 / GluN2A, GluN1 / GluN2B, GluN1 / GluN2C, and GluN1 / GluN2D receptor subtypes. Approximately 14 mm glass coverslips were placed in 24-well plates, and cells were grown at a rate of 3 × 10⁶ cells / well. 4 Seeding density was 1 cell / well. The external solution contained 137 mM NaCl, 4 mM KCl, 1.8 mM CaCl2, 10 mM glucose, and 10 mM HEPES, adjusted to pH 7.4 with NaOH. The intracellular microelectrode solution consisted of 140 mM CsCl, 5 mM EGTA, and 10 mM HEPES, adjusted to pH 7.2 with CsOH. The test compound (NFI23) was initially dissolved in DMSO to prepare a 30 mM stock solution. This was then serially diluted in DMSO to obtain intermediate concentrations of 30, 10, 3.0, 1.0, 0.3, and 0.1 mM. Subsequently, the intermediate solutions were diluted 1000-fold with the external solution to prepare final working solutions at concentrations of 100, 30, 10, 3.0, 1.0, 0.3, and 0.1 μM. The final DMSO concentration in all working solutions was maintained at 0.1%. Five concentrations were selected for NMDA current suppression and IC50 testing. 50Value determination. Glass microelectrodes were fabricated using a Sutter P-1000 puller. Recordings were performed at a holding potential of -70 mV. NMDA receptor-mediated currents were induced by the application of 100 μM NMDA and 10 μM glycine. Test compounds were applied in increments of concentration, each application lasting 10–15 seconds. Peak and steady-state currents were recorded. The currents induced by the test compounds were normalized to the reaction induced by 100 μM NMDA + 10 μM glycine. IC50 50 The value is calculated through nonlinear regression using the following equation:

[0245] Y=Bottom+(Top–Bottom) / [1+10^((LogIC 50 –X)×HillSlope)],where IC 50 This is the concentration observed at half of the maximum reaction value, where h is the Hill coefficient. Curve fitting and IC50 50 Value computation was performed using GraphPad Prism 8.0 software, such as... Figure 6 As shown in Table 4.

[0246] Depend on Figure 6 As shown in Table 4, treatment with glycine and NMDA significantly increased GluN1 / GluN2B receptor-mediated currents. Subsequently, NFI23 decreased these currents in a concentration-dependent manner. At a concentration of 10 μM, NFI23 produced only a minimal inhibitory effect on these GluN2A / GluN2B receptor-mediated currents, indicating its high selectivity for GluN1 / GluN2B receptors.

[0247] Table 4

[0248]

[0249] Plasma stability test

[0250] Blank human plasma was added to each incubation well, followed by a working solution containing the test or control compound Ifenprodil (prepared by diluting the stock solution to 10 μM with phosphate buffer, the diluent being the compound dissolved in DMSO), to a final concentration of 1 μM. All incubation mixtures were maintained in a 37°C water bath throughout the experiment. At 0, 5, 15, 30, 60, and 120 minutes, 50 μL of sample was removed from the reaction mixture and immediately quenched with 500 μL of ice-cold acetonitrile / methanol (1:1, v / v) containing internal standards: labetalol (50 ng / mL) and thiophanate-methyl (50 ng / mL). The sample was centrifuged at 4000 rpm for 10 minutes. 100 μL of the supernatant was mixed with 100 μL of ultrapure water and analyzed by LC-MS / MS. The in vitro half-life (T0) was determined. 1 / 2The remaining percentage (%) is calculated using the first-order elimination equation: 100 × (PAR at a certain time point / PAR at the beginning), where PAR is the peak area ratio of the analyte to the internal standard (IS).

[0251] The elimination rate constant (k) was determined by linear regression of the natural logarithm of the remaining percentage against incubation time, from which the in vitro half-life (T) was derived. 1 / 2 As shown in Table 5.

[0252] Table 5

[0253]

[0254] a Ifenprodil has a plasma stability half-life greater than 4.8 h.

[0255] As shown in Table 5, NFI23 exhibits good plasma stability.

[0256] Pharmacokinetic studies

[0257] Single-dose pharmacokinetic studies were conducted in Sprague Dawley rats (n=3) administered NFI23 via intravenous injection (2 mg / kg) and oral administration (10 mg / kg). The intravenous formulation was prepared by dissolving NFI23 in a mixture of 10% DMSO and 90% saline to a clear solution; the oral formulation was prepared by suspending NFI23 in a 0.5% sodium carboxymethyl cellulose aqueous solution, both freshly prepared for use. Approximately 0.05 mL of blood was collected from the buccal vein at 5, 15, and 30 minutes post-administration; and at 1, 2, 4, 6, 8, and 24 hours post-administration. Samples were collected using EDTA-K2 anticoagulant tubes and placed on wet ice for no more than 15 minutes. Plasma was then separated by centrifugation at 6000 rpm for 3 minutes. Plasma concentrations were determined using a validated LC-MS / MS method. Pharmacokinetic parameters were calculated using Phoenix WinNonlin 8.2 software based on non-zonal analysis and are shown in Table 6.

[0258] Table 6

[0259]

[0260] As shown in Table 6, NFI23 exhibits rapid onset of action, wide tissue distribution, and effective central nervous system penetration.

[0261] Pharmacodynamic evaluation in MCAO rat model

[0262] Thirty-six Sprague-Dawley (SD) rats (weighing 200–220 g) were randomly divided into six groups. Prior to surgery, the rats were fasted overnight and anesthetized with chloroform hydrate (350 mg / kg, intravenously). MCAO surgery was then performed. A brief description of the procedure is as follows: the right common carotid artery (CCA) was exposed, and a 0.30–0.35 mm diameter nylon monofilament suture was inserted to occlude cerebral blood flow. After 2 hours of ischemia, the suture was withdrawn to restore perfusion. Subsequently, NFI23 (2, 5, and 10 mg / kg) and Ifenprodil (10 mg / kg) were administered intravenously (using the same methods as for pharmacokinetic assays). Neurological deficits were assessed using the Longa score 24 hours postoperatively. The rats were then euthanized, and the brains were rapidly harvested and placed at -20°C for 30 minutes. Each brain was cut into five coronal sections approximately 2 mm thick. Sections were incubated in 1% triphenyltetrazolium chloride (TTC, pH 7.4) at 37°C for 30 minutes, followed by fixation with 4% formalin for 30 minutes. Infarcted areas were observed and photographed. White (ischemic) areas and total brain area were quantified using image analysis software. Infarct volume was expressed as a percentage using the following formula: Infarct volume (%) = (Infarcted area / Total brain area) × 100%. All experimental protocols were performed in accordance with the National Institutes of Health's "Guidelines for the Care and Use of Laboratory Animals" and were approved by the Animal Experimentation Committee of Shenyang Pharmaceutical University. Figure 7 As shown.

[0263] Statistical analysis

[0264] All experimental data were collected, processed, and analyzed using GraphPad Prism 8.0. All data are expressed as mean ± standard deviation (SD). Unpaired t-tests were used to analyze statistical differences between two groups, while one-way ANOVA was used to compare multiple groups. A p-value < 0.05 was considered statistically significant.

[0265] Molecular dynamics (MD) simulations

[0266] Molecular dynamics (MD) simulations were performed using GROMACS 2022.3. For small molecule preparation, GAFF force field parameters were assigned using AmberTools22. Hydrogenation and RESP charge calculations were performed using Gaussian 16W. The generated potential energy data were incorporated into the topology file of the MD system. Simulations were conducted under constant temperature (300 K) and atmospheric pressure (1 bar). An AMBER99SB-ILDN force field was used, with TIP3P water molecules as the solvent. System neutrality was achieved by adding an appropriate amount of Na+ ions. Energy minimization was performed using the steepest descent method, followed by equilibration under NVT and NPT clusters, 100,000 steps per cluster. The two clusters used a coupling constant of 0.1 ps for a total duration of 100 ps. Subsequently, a 100 ns production MD simulation was performed with a time step of 2 fs, for a total of 5,000,000 steps. After the simulations, trajectory analysis was performed using GROMACS built-in tools. Key metrics such as root mean square deviation (RMSD), root mean square fluctuation (RMSF), and radius of gyration (Rg) of amino acid residues were calculated. Furthermore, molecular mechanical / generalized Born surface area (MM / GBSA) free energy and free energy landscape analysis were performed, such as... Figure 8 As shown in Table 7.

[0267] Table 7

[0268]

[0269] Depend on Figure 8 As can be seen from Table 7, the combination of NFI23 with GluN2B-NMDAR is thermodynamically favorable, mainly driven by hydrophobic effects and electrostatic interactions.

Claims

1. An N-arylindole derivative, characterized in that: N-arylindole derivatives are compounds of Formula I, pharmaceutically acceptable salts and their optically active forms or racemates, and diastereomers. in: R1 and R2 may be the same or different and are independently selected from H, hydroxyl, halogen, cyano, nitro, unsubstituted or substituted with at least one R3, C1-6 alkyl, C3-8 cycloalkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkylsulfonylamino, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 alkoxy, C2-6 alkenyloxy, C2-6 alkynyloxy, C1-6 alkylthio, C2-6 alkenylthio, C2-6 alkynylthio, C1-6 alkylsulfonyl, C2-6 alkenylsulfonyl, C2-6 alkynylsulfonyl, C1-6 alkylsulfinyl, C2-6 alkenylsulfonyl, C1-6 alkylsulfinyl, C2 ... 2-6-Alkenylsulfinyl, C2-6-alkynylsulfinyl, C3-8-cycloalkenyl; unsubstituted or substituted carbonyl group with at least one R4, unsubstituted or substituted amino group with at least one R5, unsubstituted or substituted phenyl group with at least one R6, benzyl, benzyloxy, benzoyl, benzylsulfonyl, pyridine ring, pyrazole ring, pyrrole ring, pyrimidine ring, quinoline ring, isoquinoline ring, imidazole ring, morpholine ring, piperazine ring, pyridazine ring, pyrazine ring, piperidine ring, thiophene ring, thiazole ring, isothiazole ring, benzothiazole ring, pyran ring, indole ring, oxazole ring, isoxazole ring, triazole ring, benzotriazole ring, or furan ring; R3 is selected from H, halogen, hydroxyl, cyano, nitro or amino; R4 is selected from H, hydroxyl, C1-6 alkyl, amino, C1-6 alkylamino, C1-6 alkoxy, C1-6 alkylthio, and C3-8 cycloalkyl. R5 is selected from H, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkylsulfonyl, C2-6 alkenylsulfonyl, C2-6 alkynylsulfonyl, C1-6 alkyl carbonyl, C2-6 alkenyl carbonyl, and C2-6 alkynyl carbonyl. R6 is selected from H, halogen, C1-6 alkyl, C1-6 alkylamino, C3-7 cycloalkyl, C1-6 alkoxy, C1-6 alkylsulfonylamino, benzyloxycarbonyl, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 haloalkyl, hydroxyl, cyano, nitro, formyl, and amino. n is selected from 0, 1, 2, 3, or 4; m is selected from 0, 1, 2, 3, 4, or 5; X is selected from O, NH, or NHNHCO.

2. The N-arylindole derivative according to claim 1, characterized in that: The N-arylindole derivatives are compounds of formula I, pharmaceutically acceptable salts, and their optically active forms or racemates, or diastereomers; wherein, R1 and R2 may be the same or different and are independently selected from H, hydroxyl, halogen, cyano, nitro, unsubstituted or substituted with at least one R3 C1-6 alkyl, C3-8 cycloalkyl, C2-6 alkenyl, C2-6 alkynyl, C1-6 alkylsulfonylamino, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 alkoxy, C2-6 alkenyloxy, C2-6 alkynyloxy, C1-6 alkylthio, C2-6 alkenylthio; unsubstituted or substituted with at least one R6 phenyl, benzyl, benzyloxy, benzoyl; R3 is selected from H, halogen, hydroxyl, cyano, nitro or amino; R6 is selected from H, halogen, C1-6 alkyl, C1-6 alkylamino, C3-7 cycloalkyl, C1-6 alkoxy, C1-6 alkylsulfonylamino, benzyloxycarbonyl, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 haloalkyl, hydroxyl, cyano, nitro, formyl, and amino. N is 0, 1, 2, 3, or 4; m is selected from 1, 2, 3, or 4; X is selected from O, NH, or NHNHCO.

3. The N-arylindole derivative according to claim 2, characterized in that: The N-arylindole derivatives are compounds of Formula I, pharmaceutically acceptable salts, and their optically active forms or racemates, or diastereomers. In the formula, R1 and R2 may be the same or different and are independently selected from H, hydroxyl, halogen, nitro, unsubstituted or substituted with at least one R3 C1-6 alkyl, C2-6 alkenyl, C2-6 alkoxy, C2-6 alkenyloxy, C2-6 alkoxy; unsubstituted or substituted with at least one R6 phenyl, benzyl; R3 is selected from H, halogen, hydroxyl, cyano, nitro or amino; R6 is selected from H, halogen, C1-6 alkyl, C1-6 alkylamino, C3-7 cycloalkyl, C1-6 alkoxy, C1-6 alkylsulfonylamino, benzyloxycarbonyl, C1-18 alkoxycarbonyl, C1-18 alkoxysulfonyl, C1-6 haloalkyl, hydroxyl, cyano, nitro, formyl, and amino. N is 0, 1, 2, 3, or 4; m is selected from 1, 2, 3, or 4; X is selected from O, NH, or NHNHCO.

4. The N-arylindole derivative according to claim 3, characterized in that: The N-arylindole derivatives are: N-(4-hydroxybenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI1); N-Benzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI2); 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylbenzyl)acetamide (NFI3); N-(4-methoxybenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI4); N-(4-fluorobenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI5); N-(4-chlorobenzyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI6); N-Benzyl-2-(5-bromo-1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI7); 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylbenzyl)acetamide (NFI8); 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methoxybenzyl)acetamide (NFI9); 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxybenzyl)acetamide (NFI10); N-(2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazine (NFI11); 4-Methyl-N-(2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI12); 4-Methoxy-N-(2-(1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazide (NFI13); 4-Hydroxy-N-(2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetyl)benzoylhydrazine (NFI14); Benzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI15); 4-Methylbenzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI16); 4-Chlorobenzyl-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetate (NFI17); 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenylacetamide (NFI18); N-(4-methoxyphenyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI19); 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenylethylacetamide (NFI20); N-(4-methoxyphenethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI21); 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylphenethyl)acetamide (NFI22); N-(4-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI23); 2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(3-phenylpropyl)acetamide (NFI24); 2-(5-bromo-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI25); 2-(5-chloro-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI26); 2-(5-fluoro-1-(3-(((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-hydroxyphenylethyl)acetamide (NFI27); N-(2-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI28); N-(3-hydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI29); N-(3,4-dihydroxyphenylethyl)-2-(1-(3-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI30); 2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-phenethylacetamide (NFI31); 2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)-N-(4-methylphenethyl)acetamide (NFI32); N-(4-methoxyphenethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI33); N-(4-hydroxyphenylethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI34); N-(4-fluorophenylethyl)-2-(1-(2-((2-methyl-1H-imidazol-1-yl)methyl)phenyl)-1H-indol-3-yl)acetamide (NFI35).

5. The N-arylindole derivative according to any one of claims 1-4, characterized in that: The pharmaceutically acceptable salt is obtained by reacting the compound of Formula I with an acid to obtain the corresponding salt; wherein the acid is hydrochloric acid, sulfuric acid, phosphoric acid, tartaric acid, citric acid, methanesulfonic acid, hydrobromic acid, maleic acid, or benzenesulfonic acid.

6. A pharmaceutical composition, characterized in that, Contains the N-arylindole derivatives of claim 1, pharmaceutically acceptable salts and their optically active forms or racemates, and diastereomers.

7. A pharmaceutical preparation, characterized in that, The formulation uses the compound of Formula I as claimed in claim 1, a pharmaceutically acceptable salt and its optically active form or racemate, diastereomer; or the pharmaceutical composition of claim 6 as the active ingredient, wherein the active ingredient is mixed with a pharmaceutically acceptable carrier, wherein the active ingredient accounts for 0.01-99.9% of the mass of the formulation.

8. The pharmaceutical preparation according to claim 7, characterized in that, The formulation is a tablet, coated tablet, capsule, solution for injection, ampoule, suppository, patch, inhalable powder, suspension, emulsion or ointment.

9. An application according to claim 1, claim 6, or claim 7, characterized in that: The use of the compound of Formula I as claimed in claim 1, a pharmaceutically acceptable salt thereof and its optically active form or racemate, diastereomer, pharmaceutical composition of claim 6, or pharmaceutical formulation of claim 7 as a GluN2B-NMDAR antagonist.

10. The application according to claim 9, characterized in that: The use of the compound of Formula I as claimed in claim 1, a pharmaceutically acceptable salt thereof and its optically active form or racemate, diastereomer, pharmaceutical composition of claim 6, or pharmaceutical formulation of claim 7 in the preparation of an anti-ischemic stroke drug.

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