Application of small molecule RNA (Ribonucleic Acid) in preparation of medicine for treating depression

Abstract

The invention discloses application of micromolecule RNA in preparation of a medicine for treating depression, and belongs to the technical field of biomedicine. The invention finds that the small molecule RNA comprising the nucleotide sequence shown as ACAUUCA has a remarkable anti-depression effect. The micromolecule RNA, especially miR-181a-5p, acts on a chronic social contusion stress (CSDS) model mouse of which the expression level of the mPFC brain region is remarkably reduced, and the miR-181a-5p level of the mPFC brain region of the CSDS model mouse can be improved, so that the pathological change and depression-like behaviors of the CSDS mouse can be remarkably improved. According to the application of the small-molecule RNA in preparation of the anti-depression medicine, the small-molecule RNA can recover mitochondrial functions of neurons in a CSDS model mouse mPFC brain region, repair neuronal synaptic injury and improve and recover depression-like behaviors of depression diseases.

Description

Technical Field

[0001] The present application relates to the field of biomedical technology, and specifically to the application of small molecule RNA in the preparation of drugs for treating depression. Background Art

[0002] Depression, also known as depressive disorder, is a neuropsychiatric disorder with high morbidity and recurrence. It is characterized by persistent low mood, loss of interest, and anhedonia. Some patients may engage in self-harm and suicidal behavior, as well as psychotic symptoms such as delusions and hallucinations. The pathogenesis of depression involves biochemistry, neuroendocrinology, neuroimmunology, sleep and electrophysiological abnormalities, brain imaging, genetics, and psychosocial factors, and is still not fully understood. Currently, the recommended antidepressants for clinical use include selective serotonin reuptake inhibitors (SSRIs), selective serotonin and norepinephrine reuptake inhibitors (SNRIs), norepinephrine and specific serotonin reuptake inhibitors (NaSSAs), and norepinephrine and dopamine reuptake inhibitors (NDRIs).

[0003] The number of people suffering from depression is increasing year by year worldwide, affecting more than 300 million people. However, existing antidepressants have many problems: their mechanism of action is single, with mainstream drugs primarily regulating the serotonin, norepinephrine, or dopamine systems, but failing to intervene in and regulate other multiple mechanisms involved in the pathology of depression, such as glutamate system imbalance, neurogenesis damage, neuroinflammation, and mitochondrial dysfunction; their onset of action is delayed, with most antidepressants (such as SSRIs and SNRIs) requiring 2-6 weeks of continuous treatment to be effective, which may lead to reduced patient compliance and even increased suicide risk; their clinical efficacy is low, with approximately 30%-50% of patients responding poorly to first-line drugs; there are large individual differences, with genetic, metabolic, and environmental factors leading to significant differences in drug responses among different patients; there are also many other problems, such as significant side effects, high relapse rates after discontinuation, and withdrawal reactions. The development of new antidepressants is urgently needed. Summary of the Invention

[0004] The present invention provides the use of small RNA molecules in the preparation of antidepressant drugs. Small RNA molecules can restore mitochondrial function in neurons in the mPFC brain region, repair synaptic damage, and improve and restore depressive-like behaviors in depression. The use of small RNA molecules in the preparation of antidepressant drugs of the present invention has an onset of effect within two weeks, a high overall efficacy rate, no need for long-term maintenance, and minimal side effects. This solves the technical problem of existing antidepressant drugs requiring long-term maintenance treatment and having severe side effects.

[0005] According to the purpose of the present invention, there is provided the use of a small molecule RNA in preparing a drug for treating depression, wherein the nucleotide sequence of the small molecule RNA includes the sequence ACAUUCA.

[0006] Preferably, the micromolecule RNA is miR-181a-5p, and the nucleotide sequence of the miR-181a-5p is shown in SEQ ID NO: 1.

[0007] Preferably, the vector of miR-181a-5p is a viral vector.

[0008] Preferably, the viral vector is an adenoviral vector or a lentiviral vector.

[0009] Preferably, the vector of miR-181a-5p is a non-viral vector.

[0010] Preferably, the non-viral vector is an extracellular vesicle, a liposome, a lipid complex, a cationic polymer, a chitosan polymer or a nanoparticle.

[0011] Preferably, the micromolecule RNA is used to restore mitochondrial function of neurons in the mPFC brain region.

[0012] Preferably, the micromolecule RNA is used to repair neuronal synaptic damage.

[0013] Preferably, the preparation of the small molecule RNA is nasal drops, aerosol, spray or injection.

[0014] Preferably, the excipients of the small molecule RNA preparation are water, propylene glycol, sucrose, mannitol, glucose, liquid paraffin or vegetable oil.

[0015] In general, the above technical solutions conceived by the present invention have the following technical advantages compared with the existing technology:

[0016] (1) The present invention discovered that small molecule RNA has a significant antidepressant effect. This small molecule RNA, especially miR-181a-5p, acts on chronic social defeat stress (CSDS) model mice in which the expression level of the mPFC brain region is significantly reduced, and can increase the level of miR-181a-5p in the mPFC brain region of CSDS model mice, thereby significantly improving the pathological changes and depressive-like behaviors of CSDS mice.

[0017] (2) The antidepressant effect of the small molecule RNA of the present invention, especially miR-181a-5p, has the characteristics of rapid onset, high overall efficacy, no need for long-term maintenance and low side effects, which is more advantageous than traditional antidepressants.

[0018] (3) The use of small molecule RNA in the present invention, especially miR-181a-5p, in the preparation of antidepressant drugs. Small molecule RNA, especially miR-181a-5p, can restore the mitochondrial function of neurons in the mPFC brain region of CSDS model mice, repair neuronal synaptic damage, and improve and recover depressive-like behaviors of depression.

[0019] (4) miRNA is a type of non-coding small RNA molecule whose main function is post-transcriptional translation inhibition and mRNA degradation. The post-transcriptional inhibition of the present invention only requires that 7 nucleotide sequences of miRNA (nucleotide sequences at positions 2-8 at the 5' end of the miR-181a-5p sequence) bind to the target gene mRNA to achieve the effect of translation inhibition of the target gene expression process, thereby exerting its effect. The nucleotide sequences at positions 2-8 at the 5' end of the miR-181a-5p sequence are the core elements of its targeted regulation of mRNA translation process. They bind to the 3' untranslated region (3'UTR) of the target mRNA through complementary base pairing, determining target specificity.

[0020] (5) The social contact experiment showed that the social contact ratio of mice in the chronic social frustration stress model decreased from 1.264±0.123 to 0.354±0.065 (p<0.001). Compared with the control group, miR-181a-5p significantly increased the social contact ratio of mice in the chronic social frustration stress model group, increasing it to 1.058±0.124 (p<0.001).

[0021] (6) The sugar water preference experiment showed that after the chronic social frustration stress model was established, the sugar water preference ratio of mice decreased from 85.7%±1.2% to 77.8%±1.5% (p<0.001). Compared with the control group, miR-181a-5p significantly increased the sugar water preference ratio of mice in the stress model group, restoring it to 84.0±1.2% (p<0.01). BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 is the expression level of miR-181a-5p in the mPFC of normal mice and CSDS model mice.

[0023] Figure 2 The effect of overexpression of miR-181a-5p in the mPFC brain region on the social contact ratio of CSDS model mice.

[0024] Figure 3 This is the effect of overexpression of miR-181a-5p in the mPFC brain region on the sugar-water preference ratio in social contact of CSDS model mice. DETAILED DESCRIPTION

[0025] In order to make the objectives, technical solutions and advantages of the present invention more clearly understood, the present invention is further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely for the purpose of explaining the present invention and are not intended to limit the present invention. In addition, the technical features involved in the various embodiments of the present invention described below may be combined with each other as long as they do not conflict with each other.

[0026] The dosage in the present invention varies depending on the patient's age and weight, the nature and severity of the disease, and the route of administration. The results of animal experiments and various circumstances can be referenced, and the total dosage should not exceed a certain range. In animal experiments, the dosage was 10 nmol per 23g C57 mouse. When administered to animals, including humans, the dosage can be converted based on the above experimental data, depending on the patient's age and weight, the nature and severity of the disease, and the route of administration, and should not exceed three times the converted dosage.

[0027] The nasal drops of the present invention may be added with various pharmaceutical excipients, including but not limited to water, propylene glycol, sucrose, mannitol, glucose, liquid paraffin, vegetable oil, etc. The various excipients are used to promote absorption, prevent mucosal edema, and regulate osmotic pressure, pH value and viscosity.

[0028] 1. Experimental Animals

[0029] Seventy male C57BL / 6J mice (6-7 weeks old, 20-22 g) were purchased from Hunan Slake Jingda Laboratory Animal Co., Ltd., and 20 male CD1 mice (6-7 months old) were purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd. Mice were housed at a rate of 4-5 per cage for one week in a standard animal environment (temperature: 22 ± 1°C; humidity: 50% ± 8%; daylight cycle: 12 h light / 12 h dark) prior to the formal experiments, with free access to adequate food and water.

[0030] 2. Construction of a Chronic Social Defeat Stress Depression Model in Mice

[0031] The chronic social defeat stress mouse depression model simulates social stress between C57BL / 6J and CD1 mice, causing the socially frustrated C57BL / 6J mice to exhibit stressful states such as social avoidance. It is currently the depression model that most closely resembles the pathogenesis of human depression. CD1 mice, which are highly aggressive, were screened for the chronic social defeat stress model. C57BL / 6J mice (7-8 weeks old) were placed in a cage housing CD1 mice. Due to their strong territorial awareness, CD1 mice will persistently attack the C57BL / 6J mice. When the aggression persisted for 5-10 minutes and the C57BL / 6J mice displayed fearful behaviors such as avoidance and freezing, and their fur became matted, they were separated by a perforated transparent acrylic partition, preventing physical contact between them and the CD1 mice, but allowing them to see and smell each other. The same procedure was repeated with different CD1 mice for 10 consecutive days. Twenty-four hours after the modeling, social contact and sugar water preference tests were performed. In this study, the social contact ratio in the social contact experiment was less than 1 and the sugar water preference ratio was less than 75% as the conditions for successful depression modeling.

[0032] 3. Experimental Process

[0033] (1) Mice that had successfully undergone chronic social defeat stress modeling and control male C57BL / 6J mice were selected, and the expression level of miR-181a-5p in the mPFC brain region was detected by qPCR;

[0034] (2) Mice with successful chronic social defeat stress modeling and control male C57BL / 6J mice were selected, and miR-181a-5p overexpressing adeno-associated virus or its control virus were injected into the mPFC on both sides of the mice using a stereotaxic apparatus. The virus titer was 1×10 9 The injection dose was 200 nanoliters. Behavioral testing was performed 3 weeks after virus injection.

[0035] Statistical Analysis

[0036] Statistical analysis of the experimental data was performed using Graphpad Prism 9 software. Two-way analysis of variance was performed with Bonferroni's post hoc analysis. All data are presented as mean ± standard error. A p value < 0.05 was considered statistically significant.

[0037] 5. Experimental Results

[0038] By the attached Figure 1 It can be seen that the level of miR-181a-5p in the mPFC brain region of mice in the chronic social frustration stress model group was significantly reduced, and the relative level decreased from 1.000±0.074 to 0.586±0.038 (p<0.01);

[0039] By the attached Figure 2 It can be seen that the social contact experiment showed that the social contact ratio of mice decreased from 1.264±0.123 to 0.354±0.065 (p<0.001) after chronic social frustration stress modeling. Compared with the control group, miR-181a-5p significantly increased the social contact ratio of mice in the chronic social frustration stress model group, increasing it to 1.058±0.124 (p<0.001).

[0040] By the attached Figure 3 It can be seen that the sugar water preference experiment showed that after the chronic social frustration stress model was established, the sugar water preference ratio of mice decreased from 85.7%±1.2% to 77.8%±1.5% (p<0.001). Compared with the control group, miR-181a-5p significantly increased the sugar water preference ratio of mice in the stress model group, restoring it to 84.0±1.2% (p<0.01).

[0041] It will be easily understood by those skilled in the art that the above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. Use of a small molecule RNA in the preparation of a drug for treating depression, wherein the nucleotide sequence of the small molecule RNA includes the sequence ACAUUCA.

2. The use according to claim 1, characterized in that The micromolecule RNA is miR-181a-5p, and the nucleotide sequence of the miR-181a-5p is shown in SEQ ID NO:

1.

3. The use according to claim 1 or 2, characterized in that The vector of the miR-181a-5p is a viral vector.

4. The use according to claim 3, characterized in that The viral vector is an adenoviral vector or a lentiviral vector.

5. The use according to claim 1 or 2, characterized in that The miR-181a-5p vector is a non-viral vector.

6. The use according to claim 5, characterized in that The non-viral vector is an extracellular vesicle, a liposome, a lipid complex, a cationic polymer, a chitosan polymer or a nanoparticle.

7. The use according to claim 1 or 2, characterized in that The small molecule RNA is used to restore the mitochondrial function of neurons in the mPFC brain region.

8. The use according to claim 1 or 2, characterized in that The small molecule RNA is used to repair neuronal synaptic damage.

9. The use according to claim 1 or 2, characterized in that The preparation of the small molecule RNA is nasal drops, aerosols, sprays or injections.

10. The use according to claim 9, characterized in that The auxiliary materials of the small molecule RNA preparation are water, propylene glycol, sucrose, mannitol, glucose, liquid paraffin or vegetable oil.

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