Application of diltiazem in preparation of medicine for treating autism

Abstract

The invention relates to application of diltiazem in preparation of a medicine for treating autism, belongs to the technical field of biological medicines, and selects a new medicine, namely diltiazem, which has a good treatment effect on autism through a constructed medicine-protein-disease multi-element and multi-layer interaction network. The diltiazem is further determined to be capable of relieving the autism-like manifestation of the mouse through tests, and particularly, the diltiazem can be used for reducing the repetitive self-combing behavior of the gene knockout mouse and reducing the repetitive behavior of the autism mouse; the social ability and the social novelty of the gene knockout mouse can be improved, and the social behavior disorder of the autistic mouse can be improved, so that the diltiazem has a treatment effect on the autistic mouse.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedicine, and in particular to application of diltiazem in preparing a medicine for treating autism. Background Art

[0002] Autism, also known as autism, is a group of neurodevelopmental disorders that begin in infancy and early childhood. Its main clinical symptoms include social disorders, repetitive stereotypes, and cognitive deficits. Social disorders involve defects in social interaction, lack of eye contact, reduced ability to continue conversations, and impaired daily interaction skills. Repetitive stereotypes include compulsive behaviors, unique attachment to objects, rigid adherence to routines or etiquette, and repetitive motor quirks and self-stimulatory behaviors. Cognitive deficits include language abnormalities. The main symptoms of autism are obvious in the infants and young children of patients, and even make patients lose the ability to take care of themselves, causing huge economic burdens and mental stress to the family. The incidence of autism is increasing year by year. The high incidence makes autism one of the most common developmental disabilities and a neurodevelopmental disease in children that cannot be ignored.

[0003] Studies have shown that the concentration of GABA(A) receptors in the cortical region is increased in patients with autism, and two SNPs in the GABA(A) receptor γ3 gene are significantly associated with autism. This gene or its proximal gene can cause the onset of autism. Glutamate dysfunction, manifested as reduced excitatory transmission. NMDA-mediated plasticity dysfunction and mGluR-mediated signal transduction play a role in the cognitive and behavioral pathology of autism. Neuronal acetylcholine receptor subunits are expressed at reduced levels in patients with autism, leading to cholinergic abnormalities. The cholinergic system mediates a series of brain processes, including attention, cognition, sensory and motor functions, and cholinergic abnormalities may lead to sensory or attention deficits. Currently, α7 and α4β2nAChR subtypes have become possible therapeutic targets for autism. The α7 agonist galantamine achieved good results in a randomized, double-blind, placebo-controlled trial. In a survey of patients with autism spectrum disorders, it was found that almost all pore-forming cells and some auxiliary subunits of voltage-gated calcium channels were mutated. These mutations may disrupt the closed conformation of the activation gate, thereby enhancing voltage-dependent channel activation. These activation effects may lead to an imbalance in excitatory and inhibitory neurotransmission in brain circuits, leading to epilepsy, intellectual disability, and autism-like behaviors. There are also a small number of functional targets affected by autism, such as 5-hydroxytryptamine receptors, acetylcholinesterase, catechol O-methyltransferase, cytochrome P450, and muscarinic acetylcholine receptors. Serotonin syndrome with increased levels of 5-HT in the blood is the earliest and most common manifestation of autism. In children with autism, there are increased levels of 5-hydroxytryptamine in the blood and decreased levels of 5-hydroxytryptamine in the brain. Abnormalities in the serotonergic system during development are an important pathogenic factor in autism and will have long-term effects on brain function in adulthood.

[0004] Reduced acetylcholinesterase activity in the fusiform gyrus is observed in adult patients with autism, and defects in cholinergic innervation in the fusiform gyrus may be associated with social dysfunction. Catechol-O-methyltransferase Val158Met (COMT Val158Met) polymorphism is associated with the pathogenesis of certain phenotypes in patients with autism, especially hyperactivity symptoms. Animal models also indicate that changes in COMT levels in the cerebral cortex can lead to changes in dopamine levels, and it is speculated that the COMT gene may be one of the genetic factors involved in neuropsychological function. Genetic polymorphisms of cytochrome P450 enzymes are also associated with autism, especially CYP27B1, which is essential for the normal metabolism of vitamin D. Autistic patients have defects or deficiencies in vitamin D metabolism, which may lead to abnormal neuronal growth and neurodevelopment in autistic patients. Reduced brain muscarinic cholinergic receptor activity may lead to autistic phenotypes, and treatment with M1 muscarinic receptor agonists can reduce repetitive behaviors and restricted interests in patients with autism by stimulating striatal M1 muscarinic receptors and regulating striatal dopamine signals.

[0005] Currently, there are limited treatments for autism, and more drugs that can cure autism have not yet been discovered. Summary of the invention

[0006] The purpose of the present invention is to overcome the shortcomings of the prior art and provide the use of diltiazem in the preparation of a drug for treating autism.

[0007] To achieve the above object, the technical solution adopted by the present invention is:

[0008] In a first aspect, the present invention provides use of diltiazem in preparing a medicament for treating autism.

[0009] The inventors of the present invention have discovered through a large number of experiments that an existing FDA-approved drug (diltiazem) has a new function. Diltiazem is originally a cardiovascular drug. L-type calcium channels (LTC) are mainly postsynaptic channels that play a fundamental role in regulating neuronal discharge and gene expression, as well as in important physiological and pathological processes (such as learning and memory, Parkinson's disease, autism, and the recently recognized pathophysiology of pain processes). However, the role of these channels in cardiovascular function limits the use of classical molecules to treat LTC-dependent diseases. The LTC pharmacological blocker diltiazem has been shown to effectively change short-term sensitivity to pain, inflammation-induced hyperexcitability, and neuropathy-induced allodynia,

[0010] In the technical solution of the present invention, a new drug, diltiazem, with good therapeutic effects on autism was selected through the constructed drug-protein-disease multivariate multi-layer interaction network. Experiments further confirmed that diltiazem can alleviate autism-like symptoms in mice, indicating that diltiazem has a therapeutic effect on autistic mice, which not only provides drug options for the clinical treatment of autism, but also enriches the application scope of diltiazem.

[0011] As a preferred embodiment of the application of the present invention, the diltiazem is dissolved in a solvent to prepare a diltiazem working solution, and the mass concentration of diltiazem in the diltiazem working solution is 5 to 10 mg / ml.

[0012] As a preferred embodiment of the application of the present invention, the solvent includes physiological saline.

[0013] The solvent may also be a commonly used solvent in the art, such as a buffer solution, etc., as long as it can be prepared into a working solution.

[0014] As a preferred embodiment of the use of the present invention, the dosage of diltiazem applied to animals is 10-30 mg / kg, preferably 20 mg / kg.

[0015] As a preferred embodiment of the application of the present invention, the medicine also includes excipients commonly used in pharmacy.

[0016] The auxiliary materials include at least one of fillers, binders, disintegrants and lubricants. The types of auxiliary materials are not limited to the above, and the auxiliary materials commonly used in the art are included in the present invention.

[0017] As a preferred embodiment of the use of the present invention, the drug treats autism by reducing repetitive behaviors and / or improving social behavior disorders.

[0018] Animal experiments have shown that the duration of repeated self-grooming behavior in gene knockout mice was significantly reduced after receiving intraperitoneal injection of diltiazem, indicating that intraperitoneal injection of diltiazem (20 mg / kg) restored the repetitive behavior of mice to normal levels.

[0019] In addition, diltiazem can improve the social skills deficiency and social novelty defects of gene knockout mice, and better achieve the technical effect of treating autism.

[0020] In a second aspect, the present invention provides the use of diltiazem in the preparation of a medicament for reducing repetitive behaviors in autism.

[0021] In a third aspect, the present invention provides the use of diltiazem in the preparation of a medicament for improving social behavior disorders in autism.

[0022] In a fourth aspect, the present invention provides the use of diltiazem in the preparation of a medicament for reducing repetitive self-grooming behavior in autism.

[0023] In a fifth aspect, the present invention provides the use of diltiazem in the preparation of a medicament for improving social ability and social novelty in autism.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] The present invention provides the use of diltiazem in the preparation of a drug for treating autism. By constructing a drug-protein-disease multivariate multi-layer interaction network, a new drug-diltiazem with good therapeutic effect on autism is selected. It is further determined through experiments that diltiazem can alleviate autism-like manifestations in mice, specifically reducing the repetitive self-grooming behavior of knockout mice and reducing the repetitive behavior of autistic mice; it can also improve the social ability and social novelty of knockout mice and improve the social behavior disorder of autistic mice, indicating that diltiazem has a therapeutic effect on autistic mice, which not only provides drug options for the clinical treatment of autism, but also enriches the application range of diltiazem. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1This is a subnetwork diagram of the interaction between diltiazem and its target protein in Example 1 of the present invention;

[0027] Figure 2 The timeline of the mouse behavior experiment in Example 2 of the present invention;

[0028] Figure 3 This is a graph showing the results of the repeated self-grooming experiment on mice in Example 3 of the present invention;

[0029] Figure 4 This is a diagram showing the results of the three-box social experiment on mice in Example 4 of the present invention. DETAILED DESCRIPTION

[0030] In order to better illustrate the purpose, technical solutions and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.

[0031] Example 1

[0032] The inventor team of the present invention previously constructed a drug-protein-disease multivariate multilayer interaction network through autism-related genes, including 6,906 proteins, 2,268 drugs and 21,878 diseases. And by restarting the random walk algorithm, starting from the drug or disease node, a biased random walk is used to calculate the access probability of each drug or disease to the node in the multivariate multilayer interaction network. At each step, the random walker can restart the walk or jump to the adjacent node according to the weight of different node types. Therefore, the proteins closer to the drug or disease are visited more frequently, and the diffusion matrix tends to converge as the number of walks increases. Proteins interact with other proteins to form a molecular network to function, so the target protein of the drug or the pathogenic protein of the disease will recursively affect other proteins in the molecular network. The drug or disease diffusion matrix reflects the propagation process of each drug or disease in the multivariate multilayer interaction network, and the Euclidean distance of the drug and disease diffusion matrix is ​​calculated to determine the degree of correlation between the drug and autism. According to the geometric mean of the Euclidean distance between the drugs and the autism records in the DisGeNET database, they were arranged in ascending order, and the seven cardiovascular drugs with the smallest distance were screened out as candidate drugs for autism. The drug information was annotated, including Deslanoside, Digitoxin, Felodipine, Gemfibrozil, Diltiazem, Amlodipine and Manidipine. Finally, a candidate drug for autism was screened out - the cardiovascular drug: Diltiazem (ID is DB00343).

[0033] In this study, the cardiovascular drug diltiazem was selected for experimental verification. L-type calcium channels (LTC) are mainly postsynaptic channels that play a fundamental role in regulating neuronal firing and gene expression, as well as in important physiological and pathological processes such as learning and memory, Parkinson's disease, autism, and the recently recognized pathophysiology of pain processes. However, the role of these channels in cardiovascular function limits the use of classical molecular treatments for LTC-dependent diseases. The LTC pharmacological blocker diltiazem has been shown to effectively alter short-term sensitivity to pain, inflammation-induced hyperexcitability, and neuropathy-induced allodynia. The interacting subnetworks of diltiazem and its target proteins are shown in Figure 2. Figure 1 shown.

[0034] Example 2

[0035] Large-scale sequencing has found that many autism susceptibility genes are associated with synapse formation. Synapses are key structures for transmitting information between neurons. Abnormal synapse formation and function may play an important role in the pathogenesis of autism. Studies have shown that autism may be caused by an imbalance between excitatory and inhibitory synaptic transmission. Excitatory synaptic transmission is usually mediated by glutamate as the main neurotransmitter, while inhibitory synaptic transmission mainly depends on neurotransmitters such as GABA and 5-hydroxytryptamine.

[0036] In order to further study the pathological mechanism of autism, the present invention has produced mouse models with typical autism-like symptoms targeting neurotransmitter receptors Gabra4, Gabrb1 and Htr3a. These mouse models can be used to study abnormalities in inhibitory synaptic transmission and their effects on behavior and cognitive functions.

[0037] First, the present invention selects C57BL / 6 male mice (8-12 weeks old), including Gabra4 - / - ,Gabrb1 - / - , Htr3a - / - Three kinds of knockout mice and wild-type (Wide type, WT) mice were injected intraperitoneally with drugs or saline according to their weight. After the injection, the mice were returned to the cage to wait for the drug to take effect. After the drug took effect, the behavioral experiment was performed. Each behavioral experiment was separated by one week to ensure that the drug was completely metabolized and would not affect the next behavioral experiment.

[0038] Rodent models of autism-related behaviors are important tools for understanding the genetic variation that contributes to the pathogenesis of autism. Mouse models of autism-like symptoms provide preclinical tools to evaluate the efficacy and safety of potential pharmacological therapies to reverse autism-related behaviors.

[0039] 1. Mouse model:

[0040] (1)Gabra4 - / - Mouse:

[0041] The generation of Gabra4 knockout C57BL / 6 mice by transcription activator-like (TAL) effector nuclease (TALEN) technology was performed at Cyagen Biosciences Co., Ltd. (China). Exon 1 of the mouse Gabra4 gene (GenBank accession number, NM_010251.2) was selected as the target site, and then the TALEN mRNA produced by in vitro transcription was injected into fertilized eggs for knockout mouse production, and the product was subjected to Sanger sequencing to confirm the deletion. Gabra4 knockout mice showed autism-like behavior, enhanced spatial memory, and reduced susceptibility to pentylenetetrazol-induced seizures.

[0042] (2)Gabrb1 - / - Mouse:

[0043] Gabrb1 knockout C57BL / 6J mice were generated by TALEN (transcription activator-like effector nuclease). Exon1 of the mouse Gabrb1 gene (chr5:71857359-72294152, NCBI reference sequence: NC_000071.7) was selected as the TALEN target site, and TALEN mRNAs were generated by in vitro transcription and then injected into fertilized eggs. The DNA fragments containing the target site were PCR typed and then Sanger sequenced. The primers used for genotyping included: 5'-AGGTCCATTCGGGAATTACTGCC-3' (SEQ ID NO: 1) and 5'-GGCACACCTACCCCCAAATAACA-3' (SEQ ID NO: 2). The Gabrb1 knockout mouse model showed three phenotypes: autistic-like behaviors (including impaired social behavior, cognitive deficits and increased repetitive self-grooming), impaired memory and reduced sensitivity to pentylenetetrazol-induced epileptic seizures.

[0044] (3)Htr3a - / - Mouse:

[0045] Htr3a knockout C57BL / 6N mice generated by transcription activator-like effector nuclease (TALEN) technology were generated by Cyagen Biosciences (China). Briefly, 11 bases (GGGGAAGgtaa) in exon 1 of the mouse Htr3a gene (Ensemble ID: ENSMUSG00000032269; Gene ID: 15561) were selected as TALEN target sites. The 11-base deletion at the junction of the first exon and intron disrupted the reading frame after the 16th codon in all isoforms according to the UCSC genome browser (https: / / genome.ucsc.edu / ). TALEN mRNAs were transcribed in vitro and injected into fertilized eggs for the production of knockout mice. The genomic region spanning the deletion was PCR amplified using the forward primer 5'-AGTTGGAAAAGCAGTCTGCCTGG-3' (SEQ ID NO: 3) and the reverse primer 5'-TTGACCCCACACCTCAGAATCCT-3' (SEQ ID NO: 4), followed by Sanger sequencing for genotyping. The Htr3a knockout mouse model exhibited three phenotypes: autistic-like behaviors (including impaired social behavior, cognitive deficits, and increased repetitive self-grooming), impaired memory, and reduced sensitivity to pentylenetetrazol-induced seizures.

[0046] 2. Therapeutic experiments:

[0047] (1) Drug treatment:

[0048] Diltiazem was purchased from SIGMA (USA) and dissolved in physiological saline to prepare diltiazem working solution (5 mg / ml). Diltiazem working solution (20 mg / kg) or the corresponding diltiazem working solution solvent (physiological saline) was administered to mice by intraperitoneal injection (ip), and behavioral tests were performed 30 minutes later.

[0049] (2) Mouse selection and grouping

[0050] Select C57BL / 6 male mice (8-12 weeks old), including Gabra4 - / - ,Gabrb1 - / - , Htr3a - / -Three gene knockout mice and wild-type (Wide type, WT) mice, homozygous knockout mice and wild-type mice littermate controls. Mice were group-housed (3-4 mice per cage) under specific pathogen-free (SPF) conditions, given a 12h:12h light-dark cycle, and allowed free access to food and water. Before behavioral testing, mice were handled for 5 days, animal handling and habituation, mainly stroking the back of mice to reduce the stress response of mice during the experiment, and sent to the test room within 30-60 minutes. Male mice were used in the study.

[0051] The experimental mice were divided into four groups: gene knockout mouse drug treatment group, gene knockout mouse saline treatment group, wild-type drug treatment group and wild-type saline treatment group. Each group consisted of 8-12 mice. The experiment adopted a between-subject design with a one-week drug washout period, that is, each mouse received a drug or saline injection and then underwent a behavioral task test once a week. The schedule of behavioral tests is given in the figure. The tests were carried out in the following order: self-grooming test and three-box social test.

[0052] By comparing the behavioral experimental results of the four groups of mice, if there is a statistical difference between the experimental results of the gene knockout mouse saline treatment group and the wild-type saline treatment group, it means that the gene knockout mice have abnormal behavior; if there is a statistical difference between the experimental results of the gene knockout mouse drug treatment group and the gene knockout mouse saline treatment group, it means that the drug has an effect on the abnormal behavior of the gene knockout mice; if there is no statistical difference between the experimental results of the wild-type drug treatment group and the wild-type saline treatment group, it means that the drug has no effect on the behavior of the mice; if there is no statistical difference between the gene knockout mouse drug treatment group and the wild-type saline treatment group, it means that the drug has slowed down or even cured the abnormal behavior of the gene knockout mice.

[0053] Example 3

[0054] In this example, a self-grooming experiment was conducted to detect whether diltiazem has an improving effect on the repetitive behavior of gene knockout mice.

[0055] Self-grooming behavior experiment:

[0056] Each mouse was placed individually in a self-grooming experimental apparatus without bedding, and a camera was placed 15 cm from the cage. The behavior of the mice was recorded for 20 min and used for manual scoring of repetitive self-grooming during the last 10 min of the test session. Self-grooming behavior was defined as stroking or scratching the body or face, or licking a body part. The cumulative time spent grooming all body areas was assessed using a stopwatch as described previously.

[0057] like Figure 3 As shown, Gabra4 - / - , Gabrb1 - / - and Htr3a - / - (gene knockout mouse saline treatment group) compared with wild type (wild type saline treatment group), the self-grooming time increased, and after intraperitoneal injection of diltiazem, it could reduce Gabra4 - / - , Gabrb1 - / - The self-grooming time of gene knockout mice, and the difference was statistically significant (n = 10).

[0058] Through experiments, it was found that the gene knockout mouse saline treatment group showed a higher self-grooming time compared with the wild type saline treatment group, that is, the gene knockout mice showed repetitive behaviors. And there was no statistical difference in the self-grooming time between the wild type diltiazem treatment group and the wild type saline treatment group, indicating that intraperitoneal injection of diltiazem (20 mg / kg) had no effect on the behavior of mice.

[0059] The experimental results showed that Gabra4 - / - (n = 10, **p < 0.01) and Gabrb1 - / - (n = 10, ****p < 0.0001) mice showed a significant decrease in the time of repetitive self-grooming behavior after intraperitoneal injection of diltiazem. For Htr3a1 - / - mice, there was a tendency for the time of repetitive self-grooming behavior to decrease, but the difference was not statistically significant (n = 10, 0.05 < p < 0.1), that is, diltiazem could improve the repetitive behaviors of Gabra4 - / - and Gabrb1 - / - mice, and there was no statistical difference between the gene knockout mouse diltiazem treatment group and the wild type saline treatment group, indicating that intraperitoneal injection of diltiazem (20 mg / kg) restored the repetitive behavior of mice to the normal level.

[0060] Example 4

[0061] In this example, a three-chamber social experiment was used to detect whether diltiazem had an improving effect on the social ability and social novelty of gene knockout mice.

[0062] Wild-type and knockout mice exhibit different investigative behaviors when encountering unfamiliar mice. The social approach test was performed to determine behavioral correlates of autism in mouse models. The social apparatus (40 cm × 60 cm × 20 cm) was a rectangular three-chamber box made of transparent polycarbonate, with each chamber containing an empty round metal cage (12 cm high, 11 cm diameter) allowing nose-to-nose contact between mice. Retractable doorways within two partition walls allowed mice to enter the side chambers. A top-mounted infrared camera was placed directly above the box to record sessions for scoring purposes. The apparatus was cleaned with 70% ethanol and water between tests. A minimum of 5 min was allowed for the ethanol to evaporate and the odor to dissipate between cleaning and the start of the next test. The test was divided into two 10-min phases: habituation and social approach (unfamiliar mouse with object). During the 10-min habituation period, mice were free to explore all three chambers. Next, a strange mouse was placed in a metal cage on one side of the room, and the room on the other side was empty. The mouse was allowed to explore all three rooms again to test whether it preferred to interact with the object (O) or the strange mouse (S1). The sniffing time was plotted as social preference index = TS1 / (TS1+TO), TS1: the time the test mouse interacted with the new mouse (S1, strange mouse 1), TO: the time the test interacted with the empty cage. In order to assess the preference for the new strange mouse, the mouse was then tested in a second 10-minute session, in which a new strange mouse (S2) was contained in the relative wire cage. The sniffing time was plotted as social preference index = TS2 / (TS1+TS2), TS1: the time the test mouse interacted with the familiar mouse (S1, strange mouse 1), TS2: the time the test mouse interacted with the new mouse (S2, strange mouse 2). The duration of the sniff was measured using the software EthoVision XT11.5 (Noldus), defined as positioning the nose of the test mouse within 2.5 cm of the cage. Stranger mice were between 8 and 12 weeks old, matched for breed and sex, and habituated to plastic cages the day before (30 min).

[0063] like Figure 4 As shown, Gabra4 - / - ,Gabrb1 - / - and Htr3a - / - Sociability deficits compared to wild-type, increased Gabra4 after intraperitoneal diltiazem administration - / - ,Gabrb1 - / - and Htr3a - / - Social preference index, the time spent sniffing stranger mouse 1 was longer than that of the empty cage, and the difference was statistically significant (n = 10) ( Figure 4 center left).

[0064] Gabra4 - / - ,Gabrb1- / - and Htr3a - / - Sociability deficits compared to wild-type, increased Gabra4 after intraperitoneal diltiazem administration - / - ,Gabrb1 - / - and Htr3a - / - Social preference index: the time spent sniffing stranger mouse 2 was longer than that of stranger mouse 1, and the difference was statistically significant (n=10) ( Figure 4 center right).

[0065] The experimental results show that in the 10-minute social phase of the social approach task, Gabra4 - / - ,Gabrb1 - / - and Htr3a - / - After intraperitoneal injection of diltiazem, mice spent significantly more time in the unfamiliar mouse chamber compared with the saline-treated group, indicating that diltiazem can improve Gabra4 - / - ,Gabrb1 - / - and Htr3a - / - The mice had deficient social abilities, and there was no statistical difference between the knockout mice treated with diltiazem and the wild-type mice treated with saline, indicating that intraperitoneal injection of diltiazem (20 mg / kg) reversed the Gabra4 - / - ,Gabrb1 - / - and Htr3a - / - During the 10-min social phase of social novelty, Gabra4 - / - ,Gabrb1 - / - and Htr3a - / - After intraperitoneal injection of diltiazem, mice spent significantly more time in the unfamiliar mouse chamber than the gene knockout mice treated with saline, indicating that diltiazem improved the Gabra4 - / - ,Gabrb1 - / - and Htr3a - / - There was no statistical difference between the diltiazem-treated group and the wild-type saline-treated group in the social novelty deficit of the knockout mice, indicating that intraperitoneal injection of diltiazem (20 mg / kg) can reverse the Gabra4 - / - ,Gabrb1 - / - and Htr3a - / - The social novelty deficit.

[0066] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention rather than to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the essence and scope of the technical solution of the present invention.

Claims

1. Use of diltiazem in the preparation of drugs for treating autism.

2. The use according to claim 1, characterized in that When used, the diltiazem is dissolved in a solvent to prepare a diltiazem working solution, and the mass concentration of the diltiazem in the diltiazem working solution is 5-10 mg / ml.

3. The use according to claim 1, characterized in that The solvent includes physiological saline.

4. The use according to claim 1, characterized in that The dosage of diltiazem applied to animals is 10-30 mg / kg.

5. The use according to claim 1, characterized in that The medicine also includes excipients commonly used in pharmacy.

6. The use according to claim 5, characterized in that The drug treats autism by reducing repetitive behaviors and / or improving social behavior disorders.

7. Use of diltiazem in the preparation of drugs for reducing repetitive behaviors in autism.

8. Use of diltiazem in the preparation of drugs for improving social behavioral disorders in autism.

9. Use of diltiazem in the preparation of a drug for reducing repetitive self-grooming behavior in autism.

10. Use of diltiazem in the preparation of drugs for improving social ability and social novelty in autism.

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