Application of amino-acid ester derivatives in treatment of anxiety

Abstract

The invention relates to application of amino-acid ester derivatives in drugs for treating anxiety. The amino-acid ester derivative provided by the invention has nNOS-CAPN protein-protein interaction inhibitory activity, and has a good application prospect in preparation of anti-anxiety drugs.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceuticals and provides a novel use of amino acid ester compounds. These compounds can inhibit nNOS-CAPON protein interactions and have application prospects in the preparation of antianxiety drugs. The anxiety disorders include generalized anxiety disorder, panic attacks, panic disorder, and social phobia. Background Art

[0002] Anxiety disorders are the most common mental disorders worldwide. According to the World Health Organization (WHO), anxiety disorders can be categorized as generalized anxiety disorder, panic disorder, social anxiety disorder, agoraphobia, separation anxiety disorder, specific anxiety disorders, and selective mutism. Research indicates that people may suffer from more than one anxiety disorder simultaneously, with symptoms typically beginning in childhood or adolescence and persisting into adulthood. Due to its high prevalence, chronicity, and comorbidities, the WHO has listed anxiety disorders as the ninth leading cause of health-related disability. Therefore, the prevention and treatment of anxiety disorders have become a key focus in psychiatry.

[0003] Currently, the main medications for anxiety disorders include selective serotonin reuptake inhibitors (SSRIs), serotonin-norepinephrine reuptake inhibitors (SNRIs), monoamine oxidase inhibitors (MAOIs), and benzodiazepines (BZDs). Although BZDs have been used to treat anxiety disorders, they are generally not used as first-line medications due to their limited long-term efficacy and the tendency to become addictive [5]. In the treatment of major anxiety disorders, SSRIs and SNRIs are the first-line drugs. They inhibit the reuptake of serotonin and norepinephrine by transporters in the presynaptic membrane, thereby increasing the concentration of neurotransmitters in the synapse. However, there are still a considerable number of patients who do not respond to these first-line treatment drugs, and there is a lack of guidance on the effectiveness of treating anxiety disorders. Therefore, the development of new anti-anxiety drugs has become an urgent issue.

[0004] Over the past few decades, a growing body of research has revealed that the key molecules regulating anxiety-like behaviors include γ-aminobutyric acid (GABA), N-methyl-D-aspartate (NMDA), and serotonin (5-HT). Studies of the upstream and downstream pathways of these molecules have revealed that NMDAR-mediated activation of neuronal nitric oxide synthase (nNOS) plays a crucial role in regulating anxiety. Enhanced interaction between hippocampal nNOS and its carboxy-terminal PDZ ligand (CAPON) elicits anxiety-like behaviors. Therefore, modulating nNOS-CAPON coupling holds promise for the development of novel anxiolytic drugs. Summary of the Invention

[0005] The purpose of the present invention is to provide a drug for treating anxiety through a new mechanism, wherein the drug is an amino acid ester derivative or a pharmaceutically acceptable salt thereof.

[0006] The above-mentioned drug is characterized in that by inhibiting the protein-protein interaction of nNOS-CAPON, it alleviates anxiety-like symptoms, exerts the effect of treating anxiety, and finally achieves the purpose of treating anxiety.

[0007] Therefore, the present invention provides a new use of an amino acid ester derivative or a pharmaceutically acceptable salt thereof and a pharmaceutical composition thereof for preparing an anti-anxiety drug.

[0008] Technical problem to be solved:

[0009] The present invention provides the use of an amino acid ester compound having inhibitory activity on the protein-protein interaction of nNOS-CAPON in the preparation of a drug for treating anxiety.

[0010] Technical solution:

[0011] A class of compounds shown by Formula I and Formula II or a pharmaceutically acceptable salt, hydrate or solvate thereof,

[0012]

[0013] wherein, R1 is a C1-C3 alkyl group;

[0014] R2 and R3 are independently selected from hydrogen or a C1-C4 alkyl group;

[0015] n is taken from the numbers 1, 2, 3 or 4.

[0016] Preferably, the compound is selected from:

[0017]

[0018] Compound 1, as shown in N1:

[0019]

[0020] Compound 2, as shown in N2:

[0021]

[0022] Compound 3, as shown in N3:

[0023]

[0024] Compound 4, as shown in N4:

[0025]

[0026] Compound 5, as shown in N5:

[0027]

[0028] Compound 6, as shown in N6:

[0029]

[0030] Compound 7, as shown in N7:

[0031]

[0032] Compound 8, as shown in N8:

[0033]

[0034] Compound 9, as shown in N9:

[0035]

[0036] Compound 10, as shown in N10:

[0037]

[0038] Compound 11, as shown in N11:

[0039]

[0040] Compound 12, as shown in N12:

[0041]

[0042] Compound 13, as shown in N13:

[0043]

[0044] Compound 14, as shown in N14:

[0045]

[0046] Compound 15, as shown in N15:

[0047]

[0048] Compound 16, as shown in N16:

[0049]

[0050] Compound 17, as shown in N17:

[0051]

[0052] Compound 18, as shown in N18:

[0053]

[0054] Compound 19, as shown in N19:

[0055]

[0056] Compound 20, as shown in N20:

[0057]

[0058] Compound 21, as shown in N21:

[0059]

[0060] Compound 22, as shown in N22:

[0061]

[0062]

[0063] Compound 23, as shown in N23.

[0064] The compounds provided by the present invention further include pharmaceutically acceptable equivalents of the compound or mixtures of two or more thereof.

[0065] Preferably, the compounds provided by the present invention may include one or a mixture of two or more of pharmaceutically acceptable salts, hydrates, solvates, metabolites, prodrugs.

[0066] Preferably, the compounds provided by the present invention include acid salts or base salts of the compounds provided by the present invention. The pharmaceutically acceptable salts have the pharmaceutical activity of the compound and meet the requirements both biologically and in practical applications.

[0067] The present invention provides a class of amino acid ester compounds or pharmaceutically acceptable salts thereof for treating diseases such as anxiety and the like.

[0068] Beneficial effects:

[0069] The class of amino acid ester compounds described in this application is characterized by having inhibitory activity on the interaction between nNOS-CAPON proteins and can be used for preparing drugs for treating anxiety. Detailed implementation manners

[0070] The present invention discloses a class of amino acid ester derivatives and their uses. Those skilled in the art can draw on the content of this article and appropriately improve the process parameters to achieve. The methods and applications of the present invention have been described through preferred embodiments. Relevant personnel can obviously make changes or appropriate alterations and combinations to the methods and applications described in this article without departing from the content, spirit, and scope of the present invention to implement and apply the technology of the present invention.

[0071] To enable those skilled in the art to better understand the technical solution of the present invention, the present invention will be further described in detail below in conjunction with specific embodiments.

[0072] Example 1: Screening for the inhibitory activity of amino acid ester derivatives on the nNOS-CAPON protein-protein interaction;

[0073] 1 Materials and methods

[0074] 1.1 Recombinant proteins and antibodies

[0075]

[0076] 1.2 PPI detection buffer (Assay buffer)

[0077] Formulation: 20 mM PB, pH 7.4, 1 mM EDTA, 50 mM NaCl, and 0.05% pluronic F-68

[0078] 1.3 Main instruments and consumables

[0079] Name Manufacturer Model Specification Multi-channel Pipette (8 channels) Rainin 0.5 - 10 μL Microplate Reader MD SpectraMax i3x 96-well plate Greiner 784075

[0080] 1.4 Numbering and structures of amino acid ester derivatives

[0081]

[0082]

[0083]

[0084] 1.5 Experimental system

[0085] In this experiment, a 20 μL reaction system was used. Before the experiment, each group was diluted to 4 times the working solution concentration with Assay Buffer to prepare 4×His-nNOS(1-299) reaction solution, 4×FITC-CAPON-C20 reaction solution, 4×Tb-anti-hisantibody reaction solution, and 4×test sample reaction solution. Then, 5 μL was added to a 384-well plate in sequence and reacted at 25 °C for 6 h, and the fluorescence was detected under an enzyme-labeled instrument: 337 nm - 488 nm - 520 nm.

[0086] 1.6 Data analysis

[0087] The emission light ratio of the FRET system (337 - 488 - 520 nm) is Ratio = F520 nm / F490 nm.

[0088] The inhibition rate of the compound inhibiting PPI is calculated according to the formula Inhibition(%) = 100% × (Ratio sample -Ratio 0％inhibition ) / (Ratio 100％inhibition -Ratio 0％inhibition ); where Ratio sample is the emission light ratio of the compound well, Ratio 0％inhibition is the emission light ratio of the DMSO control well, and Ratio 100％inhibition is the emission light ratio of the well without his-NOS1-299.

[0089] Use Prism 7 (GraphPad Software) log(inhibitor) vs. response--Normalized response-Variable slope to fit the S curve of the compound concentration-inhibition rate, and calculate the IC 50 value of the compound inhibiting PPI.

[0090] 2 Experimental results

[0091] All the measured amino acid ester derivatives showed certain PPI inhibitory activities (Table 1).

[0092] Table 1 Summary of the IC 50 values of the compounds inhibiting PPI in the PPI reaction system incubation

[0093]

[0094]

[0095] Example 2: Elevated plus-maze experiment of compound N10

[0096] 1.1 Experimental animals

[0097] C57BL / 6J mice, male, SPF grade, weighing 20-30 g, Beijing Vital River Laboratory Animal Technology Co., Ltd.

[0098] 1.2 Experimental equipment

[0099] Equipment Name Manufacturer Model Analytical Balance Mettler toledo XA105 Electronic Scale Changshu Shuangjie Test Instrument Factory Model T1000 Elevated Plus Maze Advance EPMM Disposable Sterile Syringe Shanghai Misawa Medical Industry Co., Ltd. 1 mL

[0100] 1.3 Experimental methods

[0101] 1.3.1 Model establishment method

[0102] Normal C57BL / 6J mice have anxiety-like symptoms and do not require model establishment.

[0103] 1.3.2 Animal grouping and drug administration

[0104] This experiment was divided into 5 groups, namely the solvent control group, N10 administration group (10 mg / kg), N10 administration group (20 mg / kg), FLX administration group (5 mg / kg), and FLX administration group (5 mg / kg). C57BL / 6J mice were randomly and blindly assigned to each group with equal probability. Mice in each group were accurately administered the corresponding control preparation, N10 preparation, or FLX solution by oral gavage using a 1 mL syringe. N10 was administered twice a day (with an interval of about 8 hours), and FLX and the solvent control group were administered once a day for 8 consecutive days.

[0105] 1.3.3 Elevated Plus Maze test (EPM)

[0106] After continuous administration for 8 days according to body weight, the elevated plus maze test was performed 1.0 h after administration on the 8th day.

[0107] A plus maze with a height of 50 cm from the ground and an arm length of 30 cm was used as the test tool. The maze consisted of two open arms (30×5 cm), two closed arms (30×5 cm). The periphery and the end of the closed arms were surrounded by a fence 15 cm high. The open arms and the closed arms were connected through a central area (5×5 cm). At the start of the test, the mouse was placed in the central area of the maze, facing the open arm and allowed to move freely. The activities of the animal within 5 min were recorded.

[0108] Observation indicators included: the number of entries into the open arm and the closed arm (counting as 1 time when all four limbs fully entered), the residence time in the open arm, and the residence time in the closed arm. The proportion of the number of entries into the open arm, the proportion of the residence time in the open arm, and the total number of entries into the maze were calculated.

[0109] 1.4 Data statistical analysis

[0110] Quantitative results were expressed as Mean±SEM. One-way ANOVA was performed on each behavioral test index using Prism6 software. After ANOVA showed differences, Fisher's LSD test was performed between two groups. A P<0.05 was defined as having a significant difference.

[0111] 2 Experimental results

[0112] Through one-way ANOVA, there were statistically significant differences in the residence time in the open arm among groups (F(4,65)=0.7125, **P=0.0072). Compared with the control group, the N10 (20 mg / kg) dose group could significantly increase the residence time (p<0.01) and the number of entries (p<0.05) of mice in the open arm. The results are shown in Table 2.

[0113] Table 2 Results of compound N10 in the EPM experiment

[0114]

[0115] Compared with the model group, *P < 0.05, **P < 0.01;

[0116] FLX: Fluoxetine Hydrochloride (Kaike).

Claims

1. A class of compounds represented by Formula I and Formula II or pharmaceutically acceptable salts, hydrates or solvates thereof, characterized in that, wherein, R1 is a C1-C3 alkyl group; R2 and R3 are independently selected from hydrogen or a C1-C4 alkyl group; n is taken from the numbers 1, 2, 3 or 4.

2. Any compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, The compounds are selected from: Compound 1, as shown in N1: Compound 2, as shown in N2: Compound 3, as shown in N3: Compound 4, as shown in N4: Compound 5, as shown in N5: Compound 6, as shown in N6: Compound 7, as shown in N7: Compound 8, as shown in N8: Compound 9, as shown in N9: Compound 10, as shown in N10: Compound 11, as shown in N11: Compound 12, as shown in N12: Compound 13, as shown in N13: Compound 14, as shown in N14: Compound 15, as shown in N15: Compound 16, as shown in N16: Compound 17, as shown in N17: Compound 18, as shown in N18: Compound 19, as shown in N19: Compound 20, as shown in N20: Compound 21, as shown in N21: Compound 22, as shown in N22: Compound 23, as shown in N23.

3. Any compound according to claims 1 to 2 or a pharmaceutically acceptable salt thereof, characterized in that Use of the compound in the preparation of a medicament for treating generalized anxiety disorder.

4. Any compound according to claims 1 to 2 or a pharmaceutically acceptable salt thereof, characterized in that, Use of the compound in the preparation of a medicament for treating panic attacks and panic disorder.

5. Any compound according to claims 1 to 2 or a pharmaceutically acceptable salt thereof, characterized in that, Use of the compound in the preparation of a medicament for treating social phobia.

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