Biomarker composition for diagnosing or predicting prognosis of cranial nerve diseases and diagnostic method using same

Abstract

[Technical Field] The present invention relates to a biomarker composition for diagnosing or predicting prognosis of brain and nervous system diseases and a diagnostic method using the same, and more particularly, to a biomarker composition for diagnosing or predicting prognosis of brain and nervous system diseases and a diagnostic method using the same, which have been invented after confirming expression levels of 1500 proteins through proteomics of biological samples in patients with brain and nervous system diseases and confirming a pattern in which NRXN2 biomarkers are increased compared to normal patients.SOLUTION: The NRXN2 biomarker of the present invention can rapidly diagnose or predict the prognosis of a brain and nervous system disease without a test such as computer tomography (CT) or magnetic resonance imaging (MRI). In addition, since there is a significant difference in biomarker expression between normal patients and patients with brain and nervous system diseases, it can be usefully utilized for accurate diagnosis or prognosis prediction of brain and nervous system diseases.SELECTED DRAWING: Figure 2b

Description

[Technical Field]

[0001] The present invention relates to a biomarker composition for diagnosing or predicting the prognosis of cranial nervous system diseases, and a diagnostic method using the same. [Background technology]

[0002] The cranial nervous system refers to the body's regulatory system, which is composed of the brain, spinal cord, cranial nerves, spinal nerves, and autonomic nervous system. Specifically, cranial nervous system disorders are diverse, including cerebral palsy, brain injury, traumatic brain injury, ischemic brain injury, concussion, cerebral contusion, stroke, cerebral infarction, cerebral hemorrhage, subarachnoid hemorrhage, Parkinson's disease, Alzheimer's disease, Huntington's disease, stroke, dementia, Lu Gehrig's disease, Pick's disease, Creutzfeldt-Jakob disease, amyotrophic lateral sclerosis, primary lateral sclerosis, degenerative ataxia, multiple sclerosis, nervous system dysfunction, memory loss, epilepsy, encephalitis, prion disease, and neurological disorders. Brain injury refers to a condition in which abnormalities in the neural tissue of the brain occur for various internal or external reasons, resulting in behavioral or functional abnormalities. Brain injury can be caused by open head trauma, closed head trauma, deceleration injury, toxic exposure, oxygen deprivation, tumors, infections, stroke, and cerebrovascular diseases such as subarachnoid hemorrhage.

[0003] Subarachnoid hemorrhage (SHE) is a condition that occurs when arteries on the surface of the brain are damaged, potentially resulting in brain damage. Emergency CT and MRI scans can be performed to diagnose SHE. If symptoms are strongly suspected but the CT and MRI scans fail to detect the condition, a lumbar puncture can be performed to examine the spinal fluid. CT and MRI angiography and catheter-assisted cerebral angiography can be performed to examine the blood vessels in detail and detect aneurysms and cerebral vascular malformations. However, these diagnostic methods require expensive testing equipment and can be complex, making them inconvenient for quickly diagnosing SHE. While numerous studies have been conducted on predictors of surgical outcomes and prognosis in SHE patients, these studies have primarily focused on neurosurgical aspects, and there has been little research on diagnosis, both in Korea and abroad.

[0004] NRXNs (Neurexins) are a family of proteins that function as cell adhesion molecules and receptors in the vertebrate nervous system. They are encoded by various unlinked genes, of which NRXN1 and NRXN3 are among the largest known human genes. NRXN2 (Neurexin-2) uses two promoters. Most NRXN2 (Neurexin-2) transcripts are generated by the upstream promoter and encode the α-neurexin isoform, while a minority of transcripts are generated by the downstream promoter and encode the β-neurexin isoform. α-neurexins contain an epidermal growth factor-like (EGF-like) sequence and a laminin G domain and have been shown to interact with neurexophilins. β-neurexins lack the EGF-like sequence and contain fewer laminin G domains than α-neurexins.

[0005] There is a continuing need for new biomarkers that can be applied to the diagnosis of various nervous system diseases, but no new diagnostic method for nervous system diseases using NRXN2 (Neurexin-2) has yet been reported. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] Korean Patent Application Publication No. 10-2020-0180210 Summary of the Invention [Problem to be solved by the invention]

[0007] An object of the present invention is to provide a biomarker composition for diagnosing or predicting the prognosis of cranial nervous system diseases, which comprises, as an active ingredient, NRXN2 (Neurexin-2) protein or a gene encoding said protein.

[0008] Another object of the present invention is to provide a composition for diagnosing or predicting the prognosis of a neurological disease, which comprises as an active ingredient a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein.

[0009] Yet another object of the present invention is to provide a kit for diagnosing or predicting the prognosis of a cranial nervous system disease, the kit including a composition for diagnosing or predicting the prognosis of a cranial nervous system disease, the composition containing as an active ingredient a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein, and instructions.

[0010] It is still another object of the present invention to provide a method for treating a subject, comprising the steps of: (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from a biological sample isolated from an individual; and (S2) A step of comparing the expression level of the protein or the gene with a control group; and (S3) A method for providing information necessary for diagnosing or predicting the prognosis of a neurological disease.

[0011] Another object of the present invention is to provide a method for treating a patient with a neurological disorder by administering a test substance to an animal model of the patient with the disorder. (S2) A method for screening therapeutic agents for cranial nervous system diseases, comprising the step of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein in an animal model treated with the test substance.

[0012] However, the technical problems that the present invention aims to achieve are not limited to those mentioned above, and other problems not mentioned will be clearly understood by those skilled in the art to which the present invention pertains from the following description. [Means for solving the problem]

[0013] The present invention provides a biomarker composition for diagnosing or predicting the prognosis of cranial nervous system diseases, which comprises, as an active ingredient, NRXN2 (Neurexin-2) protein or a gene encoding said protein.

[0014] In one embodiment of the present invention, the cranial nervous system disease may be any one selected from the group consisting of stroke, cerebral infarction, cerebral embolism, cerebral thrombosis, cerebral arteriosclerosis, cerebral hemorrhage, subarachnoid hemorrhage, Alzheimer's disease, and dementia, but is not limited thereto.

[0015] The present invention provides a composition for diagnosing or predicting the prognosis of cranial nervous system diseases, which comprises, as an active ingredient, a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein.

[0016] In one embodiment of the present invention, the preparation capable of measuring the expression level of the protein is at least one preparation selected from the group consisting of an antibody, a peptide, an aptamer, a protein, and a compound that specifically binds to the protein; The preparation capable of measuring the expression level of the gene may be any one or more preparations selected from the group consisting of a primer, an antisense oligonucleotide, and a probe that specifically binds to the mRNA of the gene, but is not limited thereto.

[0017] The present invention provides a kit for diagnosing or predicting the prognosis of a cranial nervous system disease, which includes a composition for diagnosing or predicting the prognosis of a cranial nervous system disease, which contains as an active ingredient a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein, and instructions.

[0018] In one embodiment of the present invention, the kit for diagnosing or predicting the prognosis of a cranial nervous system disease may be any one or more selected from the group consisting of a microarray, an aptamer chip kit, an ELISA (Enzyme Linked Immunosorbent Assay) kit, a blotting kit, an immunoprecipitation kit, an immunofluorescence test kit, a protein chip kit, a reverse transcription polymerase chain reaction (RT-PCR) kit, and a real-time polymerase chain reaction (qRT-PCR) kit, but is not limited thereto.

[0019] In another embodiment of the present invention, the instructions may instruct, but are not limited to, that if the expression level of NRXN2 (Neurexin-2) protein or a gene encoding the protein is higher in a patient than in a control group, the patient is determined to be a patient with a neurological disease.

[0020] The present invention relates to a method for detecting NRXN2 (Neurexin-2) protein in a biological sample isolated from an individual, comprising: (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from said biological sample; and (S2) a step of comparing the expression level of the protein or the gene with a control group; and (S3) a method of providing information necessary for diagnosing or predicting the prognosis of a cranial nervous system disease.

[0021] In one embodiment of the present invention, the method may further include, but is not limited to, (S3) determining that the patient is a patient with a cranial nervous system disease if the expression level of the protein or the gene in step (S2) is higher in the patient than in a control group.

[0022] In another embodiment of the present invention, the expression level of the protein is measured by one or more methods selected from the group consisting of Western blot, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), radial immunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunohistochemistry (IHC), immunoprecipitation assay, complement fixation assay, flow cytometry (Fluorescence Activated Cell Sorter, FACS), and protein chip; The mRNA level of the gene may be measured by one or more methods selected from the group consisting of reverse transcription polymerase chain reaction (RT-PCR), competitive reverse transcription polymerase chain reaction (competitive RT-PCR), real-time quantitative RT-PCR, multiplex reverse transcription polymerase chain reaction (multiplex PCR), real-time polymerase chain reaction (qRT-PCR), RNase protection method, Northern blotting, DNA chip technology assay, methylated DNA binding domain sequencing (MBD-seq) assay, and reduced representation bisulfite sequencing (RRBS) assay, but is not limited thereto.

[0023] In yet another embodiment of the present invention, the biological sample may be at least one selected from the group consisting of cerebrospinal fluid, exosomes, nerve cells, immune cells, blood, serum, whole blood, plasma, urine, saliva, tissue, cells, organs, bone marrow, fine needle aspiration specimen, core needle biopsy specimen, and vacuum aspiration biopsy specimen, but is not limited thereto.

[0024] The present invention provides a method for treating a patient group of a cranial nervous system disease with a test substance, comprising: (S1) treating the patient group with an animal model of a cranial nervous system disease with a test substance; and (S2) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein in an animal model treated with the test substance;

[0025] In one embodiment of the present invention, the method may further include, but is not limited to, (S3) a step of selecting the test substance as a candidate therapeutic substance if the expression level of the protein or the gene is decreased.

[0026] Furthermore, the present invention provides a diagnostic use of a composition containing, as an active ingredient, NRXN2 (Neurexin-2) protein or a gene encoding said protein for diseases of the central nervous system.

[0027] Furthermore, the present invention provides use of a composition containing, as an active ingredient, NRXN2 (Neurexin-2) protein or a gene encoding said protein for predicting the prognosis of nervous system diseases.

[0028] Furthermore, the present invention provides a use of a composition containing, as an active ingredient, NRXN2 (Neurexin-2) protein or a gene encoding said protein for producing a diagnostic agent for a central nervous system disease.

[0029] Furthermore, the present invention provides use of a composition containing, as an active ingredient, NRXN2 (Neurexin-2) protein or a gene encoding said protein for producing a drug for predicting the prognosis of a neurological disease.

[0030] Furthermore, (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from a biological sample isolated from the individual; and (S2) a step of comparing the expression level of the protein or the gene with a control group.

[0031] Furthermore, (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from a biological sample isolated from the individual; and (S2) a step of comparing the expression level of the protein or the gene with a control group.

[0032] Furthermore, (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from a biological sample isolated from the individual; (S2) comparing the expression level of the protein or the gene with a control group; (S3) determining that the patient is a patient with a cranial nervous system disease if the expression level of the protein or the gene is higher in the patient than in a control group in the step (S2); and (S4) a step of treating a patient with the neurological disease.

[0033] Furthermore, (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from a biological sample isolated from the individual; (S2) comparing the expression level of the protein or the gene with a control group; (S3) determining that the patient is a patient with a cranial nervous system disease when the expression level of the protein or the gene is higher in the patient than in a control group in the step (S2); and (S4) a step of treating a patient with the neurological disease. [Effects of the Invention]

[0034] The present invention was developed after confirming the expression levels of 1,500 proteins through proteomics of biological samples from patients with cranial nervous system diseases and identifying a pattern in which the NRXN2 biomarker is increased compared to normal patients. The NRXN2 biomarker of the present invention can rapidly diagnose or predict the prognosis of cranial nervous system diseases without the need for tests such as computed tomography (CT) or magnetic resonance imaging (MRI). Furthermore, because there is a significant difference in biomarker expression between normal patients and patients with cranial nervous system diseases, it is expected that accurate diagnosis or prognosis prediction of cranial nervous system diseases will be possible. [Brief explanation of the drawings]

[0035] [Figure 1] FIG. 1 shows the results of electron microscopy (TEM) imaging of exosomes in the cerebrospinal fluid of a patient.

[0036] [Figure 2a] FIG. 1 shows the results of comparing the expression levels of NRXN2 for the diagnosis of subarachnoid hemorrhage.

[0037] [Figure 2b] This figure shows the difference in NRXN2 expression in cerebrospinal fluid between non-SAH and SAH patients, and shows the average values ​​for each non-SAH group and SAH group (the Y axis shows the p-value of protein expression). DETAILED DESCRIPTION OF THE INVENTION

[0038] The present invention provides a biomarker composition for diagnosing or predicting the prognosis of cranial nervous system diseases, which comprises, as an active ingredient, NRXN2 (Neurexin-2) protein or a gene encoding said protein.

[0039] In one embodiment of the present invention, the cranial nervous system disease may be any one selected from the group consisting of stroke, cerebral infarction, cerebral embolism, cerebral thrombosis, cerebral arteriosclerosis, cerebral hemorrhage, subarachnoid hemorrhage, Alzheimer's disease, and dementia, but is not limited thereto.

[0040] The present invention provides a composition for diagnosing or predicting the prognosis of cranial nervous system diseases, which comprises, as an active ingredient, a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein.

[0041] In one embodiment of the present invention, the preparation capable of measuring the expression level of the protein may be any one or more preparations selected from the group consisting of antibodies, peptides, aptamers, proteins, and compounds that specifically bind to the protein, but is not limited thereto.

[0042] In another embodiment of the present invention, the preparation capable of measuring the expression level of the gene may be any one or more preparations selected from the group consisting of a primer, an antisense oligonucleotide, and a probe that specifically binds to the mRNA of the gene, but is not limited thereto.

[0043] In the present invention, NRXN2 (Neurexin-2) can be a protein series that functions as a cell adhesion molecule and receptor in the nervous system of vertebrates. Also, in the present invention, NRXN2 can refer to the protein encoded by the NRXN2 gene, and can be used as a term to refer to all gene or protein forms depending on the context.

[0044] In one embodiment of the present invention, the NRXN2 protein may have, but is not limited to, the amino acid sequence represented by SEQ ID NO: 1. In addition, in one embodiment of the present invention, specific information regarding the NRXN2 protein can be found in, but is not limited to, the National Center for Biotechnology Information (NCBI) (NCBI GenBank: AAI50276.1).

[0045] In one embodiment of the present invention, the gene encoding NRXN2 may be, but is not limited to, the nucleotide sequence represented by SEQ ID NO: 2. In addition, in one embodiment of the present invention, specific information regarding the gene encoding NRXN2 can be found in, but is not limited to, NCBI (National Center for Biotechnology Information) (NCBI Gene ID: 9379).

[0046] In the present invention, the cranial nervous system disease may be, but is not limited to, cerebral palsy, brain injury, traumatic brain injury, ischemic brain injury, cerebral concussion, cerebral contusion, stroke, cerebral infarction, cerebral embolism, cerebral thrombosis, cerebral arteriosclerosis, cerebral hemorrhage, subarachnoid hemorrhage, Parkinson's disease, Alzheimer's disease, Huntington's disease, stroke, dementia, Lu Gehrig's disease, Pick's disease, Creutzfeldt-Jakob disease, amyotrophic lateral sclerosis, primary lateral sclerosis, degenerative ataxia, multiple sclerosis, nervous system dysfunction, memory loss, epilepsy, encephalitis, prion disease, or neurological disorders.

[0047] In the present invention, brain injury may refer to a condition in which abnormalities occur in the neural tissue of the brain due to various internal or external reasons, resulting in behavioral or functional abnormalities, and brain injury may be caused by, but is not limited to, open head injury, closed head injury, deceleration injury, exposure to toxic substances, oxygen deficiency, tumor, infection, stroke, and cerebrovascular diseases such as subarachnoid hemorrhage.

[0048] In the present invention, subarachnoid hemorrhage (SHA) refers to a disease in which bleeding occurs in the space between the arachnoid membrane, one of the three meninges that cover the human brain parenchyma. The arachnoid membrane, which has a shape similar to a spider's web, is called the arachnoid membrane. Furthermore, in the present invention, the arachnoid membrane may refer to the space through which most of the large blood vessels that supply blood to the brain communicate with cerebrospinal fluid, but is not limited thereto.

[0049] In the present invention, subarachnoid hemorrhage may be caused by, but is not limited to, head trauma, and natural bleeding that can cause subarachnoid hemorrhage includes, but is not limited to, sudden rupture of an aneurysm located in an intracerebral artery or rupture of an abnormal connection between an artery and a vein (arteriovenous malformation) in or around the brain.

[0050] In the present invention, subarachnoid hemorrhage may be a concept that includes both subarachnoid hemorrhage caused by head trauma and stroke caused by spontaneous bleeding, but is not limited thereto.Furthermore, in the present invention, subarachnoid hemorrhage patients may be a concept that includes both subarachnoid hemorrhage patients caused by head trauma and subarachnoid hemorrhage (stroke) patients caused by spontaneous bleeding, but is not limited thereto.

[0051] In the present invention, subarachnoid hemorrhage is generally known to have a poor prognosis, with approximately one-third of patients dying immediately after rupture of the cerebral artery, and another one-third dying on the way to the hospital or at the hospital. It is known that only the remaining patients receive treatment, and approximately half of them may be left with a serious neurological prognosis. Therefore, rapid progress from diagnosis to treatment is required.

[0052] In the present invention, in conventional methods for diagnosing subarachnoid hemorrhage, a patient visits a nearby hospital after the onset of symptoms and undergoes a neurological examination, including computed tomography (CT) and magnetic resonance imaging (MRI), and a neuroimaging procedure to make a diagnosis. However, if a diagnosis is not made or a diagnosis is suspected after the above-mentioned tests, the presence or absence of bleeding must be confirmed by draining cerebrospinal fluid, which has the drawback of taking a long time to make a final diagnosis.

[0053] In the present invention, conventional methods for diagnosing subarachnoid hemorrhage have had several limitations: 1) most diagnostic tests must be performed within one hour of the onset of symptoms, 2) they are expensive, 3) diagnosis can only be made at a specialized or higher-level hospital, 4) even when patients visit a hospital, there is a risk of missing the appropriate timing for treatment due to the considerable waiting time, and 5) there are difficulties due to the physiological burden on the patient.

[0054] In one embodiment of the present invention, efforts were made to identify specific diagnostic biomarkers for patients with subarachnoid hemorrhage using the patient's cerebrospinal fluid, and as a result, NRXN2 (Neurexin-2) was selected as a biomarker. In the case of NRXN2, a pattern of specifically increased expression in the cerebrospinal fluid of patients with subarachnoid hemorrhage was confirmed, but the present invention is not limited to this.

[0055] In the present invention, diagnosis means to confirm the existence or characteristics of a pathological state. For the purposes of the present invention, diagnosis refers to determining the existence or non-existence of a neurological disease, the onset of the disease, and the possibility of onset (risk of onset), but is not limited thereto and also includes determining the severity of a neurological disease.

[0056] More specifically, the term "diagnosis" as used herein includes determining a subject's susceptibility to a particular disease or disorder, determining whether a subject currently has a particular disease or disorder, determining the prognosis of a subject with a particular disease or disorder, or therametrics (e.g., monitoring a subject's condition to provide information on the efficacy of treatment).

[0057] In the present invention, the term "severity" may refer to the degree of increase or decrease in the risk of developing a cranial nervous system disease, the degree of progression of a lesion, and the degree of increase or decrease in the risk of recurrence. The degree of increase or decrease in the risk of developing a cranial nervous system disease refers to the degree to which there is a risk of developing a cranial nervous system disease or the degree to which the disease may have already developed. The degree of progression of a lesion refers to the degree to which the lesion progresses after the onset of a cranial nervous system disease and the degree to which the cranial nervous system disease is mild or severe. Furthermore, the degree of increase or decrease in the risk of recurrence refers to the degree to which there is a risk or possibility of recurrence after the cranial nervous system disease has been diagnosed as being completely cured. The severity of the cranial nervous system disease can be expressed qualitatively and / or quantitatively, and can be classified according to its level.

[0058] In the present invention, prognosis prediction may mean predicting the degree of progression of a cranial nervous system disease, or may mean predicting the probability of progression, worsening, recurrence, maintenance, etc. of the condition of a cranial nervous system disease through an increase or decrease in the level of a biomarker of the present invention.

[0059] In the present invention, a biomarker is a marker that can be measured in an objective state such as a pathological difference or a response to a drug, and may be a diagnostic technique that can provide a measure of what kind of disease a patient has, and if a patient has a disease, how far the disease has progressed, but is not limited thereto.

[0060] The exosomes utilized in the present invention are a type of extracellular endoplasmic reticulum that are produced within cells and then released to the outside. They may be approximately 50 nm to 150 nm in size and can transmit various secreted cellular information, such as lipids, proteins, mRNA, and microRNA, but are not limited thereto. They can also play an important role in intercellular communication and the removal of cellular waste products, and can cross the BBB. Furthermore, exosomes have a double membrane structure that protects them from ribonucleases in the blood, making them highly stable and resistant to degradation, but are not limited thereto. Therefore, exosomes can be used as ideal biomarkers for circulating body fluids, but are not limited thereto.

[0061] In the present invention, the expression of exosomes within a size spectrum can be confirmed through nanoparticle tracking analysis, and exosome-specific markers can be confirmed and verified through NanoFC, but the present invention is not limited thereto.

[0062] In the present invention, exosomes may contain exosome membrane proteins, which may be used interchangeably with exosome-specific markers, but are not limited thereto.

[0063] In the present invention, the exosomal membrane protein may be, but is not limited to, any one or more selected from the group consisting of CD9, CD63, CD81, and CD90. In one embodiment of the present invention, the exosomal membrane protein may be, but is not limited to, any one or more selected from the group consisting of CD9 and CD63.

[0064] In the present invention, the exosome may be an exosome containing one or more membrane proteins selected from the group consisting of CD9 and CD63, but is not limited thereto.

[0065] The present invention provides a kit for diagnosing or predicting the prognosis of a cranial nervous system disease, which includes a composition for diagnosing or predicting the prognosis of a cranial nervous system disease, which contains as an active ingredient a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein, and instructions.

[0066] In one embodiment of the present invention, the kit for diagnosing or predicting the prognosis of a cranial nervous system disease of the present invention may be any one or more selected from the group consisting of a microarray, an aptamer chip kit, an ELISA (Enzyme Linked Immunosorbent Assay) kit, a blotting kit, an immunoprecipitation kit, an immunofluorescence test kit, a protein chip kit, a reverse transcription polymerase chain reaction (RT-PCR) kit, and a real-time polymerase chain reaction (qRT-PCR) kit, but is not limited thereto.

[0067] In another embodiment of the present invention, the instructions of the present invention may teach, but are not limited to, that if the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein is higher in a patient than in a control group, the patient is determined to be a patient with a neurological disease.

[0068] In yet another embodiment of the present invention, the instructions of the present invention may teach, but are not limited to, that if the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein is higher in a patient than in a control group, the patient is determined to be a patient with a poor prognosis for a neurological disease.

[0069] In yet another embodiment of the present invention, the instructions of the present invention may disclose, but are not limited to, that if the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein is higher in a patient than in a control group, the patient is determined to be a patient with a neurological disease.

[0070] In yet another embodiment of the present invention, the instructions of the present invention may disclose, but are not limited to, that if the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein is higher in a patient than in a control group, the patient is determined to be a patient with a poor prognosis for a neurological disease.

[0071] In the present invention, the kit may include a container; instructions; instructions; etc. The container may serve to package, store, and secure the formulation. The container may be, for example, a bottle, a tube, a sachet, an envelope, a tube, an ampoule, etc., which may be partially or entirely made of plastic, glass, paper, foil, wax, etc. The container may be equipped with a fully or partially detachable stopper that may initially be part of the container or that may be attached to the container by mechanical, adhesive, or other means, and may also be equipped with a stopper that allows the contents to be accessed by a syringe needle. The kit may include an outer package, which may include instructions or instructions regarding the use of each component.

[0072] The present invention relates to a method for detecting NRXN2 (Neurexin-2) protein in a biological sample isolated from an individual, comprising: (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from said biological sample; and (S2) a step of comparing the expression level of the protein or the gene with a control group; and (S3) a method of providing information necessary for diagnosing or predicting the prognosis of a cranial nervous system disease.

[0073] In one embodiment of the present invention, the method of the present invention may further include, but is not limited to, (S3) a step of determining that the patient is a patient with a cranial nervous system disease if the expression level of the protein or the gene in step (S2) is higher in the patient than in a control group.

[0074] In another embodiment of the present invention, the method of the present invention may further include, but is not limited to, (S3) a step of determining that the patient has a poor prognosis for the cranial nervous system disease if the expression level of the protein or the gene in the patient is higher than that in the control group in step (S2).

[0075] In yet another embodiment of the present invention, the method of the present invention may further comprise, but is not limited to, (S3) a step of determining that the patient is a patient with a cranial nervous system disease if the expression level of the protein or the gene in the patient is higher than that in a control group in step (S2).

[0076] In yet another embodiment of the present invention, the method of the present invention may further comprise, but is not limited to, (S3) a step of determining that the patient has a poor prognosis for the cranial nervous system disease if the expression level of the protein or the gene in the patient is higher than that in the control group in step (S2).

[0077] In yet another embodiment of the present invention, the expression level of the protein of the present invention may be measured by one or more methods selected from the group consisting of, but not limited to, Western blot, enzyme-linked immune sorbent assay (ELISA), radioimmunoassay (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunohistochemistry (IHC), immunoprecipitation analysis, complement fixation analysis, flow cytometry (Fluorescence Activated Cell Sorter, FACS), and protein chip.

[0078] In yet another embodiment of the present invention, the mRNA level of the gene of the present invention may be measured by any one or more methods selected from the group consisting of reverse transcription polymerase chain reaction (RT-PCR), competitive reverse transcription polymerase chain reaction (competitive RT-PCR), real-time quantitative RT-PCR, multiplex reverse transcription polymerase chain reaction (multiplex PCR), real-time RT-PCR (qRT-PCR), RNase protection method, Northern blotting, DNA chip analysis, methylated DNA binding domain sequencing (MBD-seq) analysis, and reduced representation bisulfite sequencing (RRBS) analysis, but is not limited thereto.

[0079] In the present invention, the mRNA level of a gene can be used interchangeably with the expression level of a gene, but is not limited thereto.

[0080] In yet another embodiment of the present invention, the biological sample of the present invention may be any one or more selected from the group consisting of cerebrospinal fluid, exosomes, nerve cells, immune cells, blood, serum, whole blood, plasma, urine, saliva, tissue, cells, organs, bone marrow, fine needle aspiration specimen, core needle biopsy specimen, and vacuum aspiration biopsy specimen, but is not limited thereto.

[0081] In the present invention, the term "individual" refers to a subject in need of treatment for a disease, and specifically refers to mammals such as humans or non-human primates, mice, rats, dogs, cats, horses, and cows, but is not limited thereto. Furthermore, in the present invention, the term "individual" can be used interchangeably with the term "patient," but is not limited thereto. In one embodiment of the present invention, the term "individual" refers to a subject in need of treatment for a cranial nervous system disease, and specifically refers to a subject in need of treatment for subarachnoid hemorrhage, but is not limited thereto.

[0082] In the present invention, the term "control group" refers to subjects who do not require treatment for a disease, and specifically refers to mammals such as humans or non-human primates, mice, rats, dogs, cats, horses, and cows, but is not limited thereto. Furthermore, in the present invention, the term "control group" can be used interchangeably with "normal individuals," but is not limited thereto. In one embodiment of the present invention, the term "control group" refers to subjects who do not require treatment for a cranial nervous system disease, and specifically refers to subjects who do not require treatment for subarachnoid hemorrhage, but is not limited thereto.

[0083] In one embodiment of the present invention, the control group (normal individuals) may have a mean cerebral blood flow test (TCD velocity) of 20 to 40, 20 to 38, 20 to 36, 20 to 34, 20 to 32, 22 to 40, 22 to 38, 22 to 36, 22 to 34, 22 to 32, 24 to 40, 24 to 38, 24 to 36, 24 to 34, 24 to 32, 26 to 40, 26 to 38, 26 to 36, 26 to 34, 26 to 32, 28 to 40, 28 to 38, 28 to 36, 28 to 34, 28 to 32, 30 to 40, 30 to 38, 30 to 36, 30 to 34, 30 to 32, 32 to 40, 32 to 38, 32 to 36, 32 to 34, or 32, but is not limited thereto.

[0084] In one embodiment of the present invention, the individual (patient) with subarachnoid hemorrhage has a mean value of cerebral blood flow test of 160 to 180, 160 to 178, 160 to 176, 160 to 174, 160 to 172, 162 to 180, 162 to 178, 162 to 176, 162 to 174, 162 to 172, 164 to 180, 164 to 178, 164 to 176, 164 to 174, 164 to 172, The range may be, but is not limited to, 172, 166-180, 166-178, 166-176, 166-174, 166-172, 168-180, 168-178, 168-176, 168-174, 168-172, 170-180, 170-178, 170-176, 170-174, 170-172, or 171.5.

[0085] In one embodiment of the present invention, the biological sample may be, but is not limited to, exosomes, and more specifically, the biological sample may be, but is not limited to, exosomes from which most of the blood has been removed.

[0086] In the present invention, the size of exosomes is 10 to 200, 10 to 190, 10 to 180, 10 to 170, 10 to 160, 10 to 150, 10 to 140, 10 to 130, 10 to 120, 10 to 110, 10 to 100, 10 to 90, 10 to 80, 10 to 70, 10 to 60, 10 to 50, 20 to 200, 20 to 190, 20 to 180, 20 to 170, 20 to 160, 20 to 150, 20 to 140, 20 to 130, 20 to 120, 20 to 110, 20 to 100, 20 to 90, 20 to 80, 20 to 70, 20 to 60, 20 to 50, 30 to 200, 30 to 190, 30 to 180, 30 to 170, 30 to 160, 30 to 150, 30 to 140, 30 to 1 30, 30 to 120, 30 to 110, 30 to 100, 30 to 90, 30 to 80, 30 to 70, 30 to 60, 30 to 50, 40 to 200, 40 to 190, 40 to to 180, 40 to 170, 40 to 160, 40 to 150, 40 to 140, 40 to 130, 40 to 120, 40 to 110, 40 to 100, 40 to 90, 40 to 80 , 40 to 70, 40 to 60, 40 to 50, 50 to 200, 50 to 190, 50 to 180, 50 to 170, 50 to 160, 50 to 150, 50 to 140, 50 to 130, 50 to 120, 50 to 110, 50 to 100, 50 to 90, 50 to 80, 50 to 70, 50 to 60, or 50.

[0087] In one embodiment of the present invention, the biological sample isolated from an individual in step (S1) of the method for providing information necessary for the diagnosis or prognosis of a cranial nervous system disease of the present invention may be extracted by, but is not limited to, the following steps: (S0-1) collecting cerebrospinal fluid drained from an individual; (S0-2) Separating the collected cerebrospinal fluid into blood and cerebrospinal fluid using a centrifuge; (S0-3) collecting and filtering the cerebrospinal fluid of step (S0-2); (S0-4) concentrating the filtered cerebrospinal fluid; (S0-5) filtering the concentrated cerebrospinal fluid; and (S0-6) A step of extracting exosomes from the filtered cerebrospinal fluid.

[0088] In one embodiment of the present invention, after the step (S0-6) of extracting exosomes from a biological sample isolated from an individual, the biological sample may further include, but is not limited to, one or more steps selected from the group consisting of: (S0-7) examining the extracted exosomes using one or more techniques selected from the group consisting of electron microscopy, nanoparticle tracking analysis (NTA), and NanoFC; and (S0-8) The step of verifying with exosome-specific markers.

[0089] In the present invention, the exosome-specific marker may be, but is not limited to, any one or more selected from the group consisting of CD9, CD63, CD81, and CD90. In one embodiment of the present invention, the exosome-specific marker may be, but is not limited to, any one or more selected from the group consisting of CD9 and CD63.

[0090] In the present invention, step (S3) of the method for providing information necessary for the diagnosis or prognosis of a cranial nervous system disease comprises: increasing the expression level of the NRXN2 (Neurexin-2) protein of the present invention or a gene encoding said protein by 10 to 30 times, 10 to 28 times, 10 to 26 times, 10 to 24 times, 10 to 23 times, 12 to 30 times, 12 to 28 times, 12 to 26 times, 12 to 24 times, 12 to 23 times, 14 to 30 times, 14 to 28 times, 14 to 26 times, 14 to 24 times, 14 ... If the serum albumin level is 23 times, 16 to 30 times, 16 to 28 times, 16 to 26 times, 16 to 24 times, 16 to 23 times, 18 to 30 times, 18 to 28 times, 18 to 26 times, 18 to 24 times, 18 to 23 times, 20 to 30 times, 20 to 28 times, 20 to 26 times, 20 to 24 times, 20 to 23 times, 22 to 30 times, 22 to 28 times, 22 to 26 times, 22 to 24 times, 22 to 23 times, or 22.7 times higher, the patient may be determined to be a patient with a cranial nervous system disease, but is not limited thereto.

[0091] In the present invention, the term "method for providing information" may be used interchangeably with "method for providing information." In the present invention, the term "method for providing information" refers to a method for providing information regarding the diagnosis of a disease, which involves analyzing an individual's biological sample or determining whether the level of a biomarker according to the present invention increases or decreases, thereby obtaining information regarding the onset or likelihood (risk) of a disease. For example, the method is a method for measuring the level of a biomarker according to the present invention and comparing it with a control group to provide information regarding whether an individual is diagnosed with a cranial nervous system disease or whether the prognosis is poor, thereby allowing for the prescription of a necessary treatment and the prediction of the therapeutic effect of the treatment.

[0092] The present invention provides a method for treating a patient group of a cranial nervous system disease with a test substance, comprising: (S1) treating the patient group with an animal model of a cranial nervous system disease with a test substance; and (S2) A method for screening a therapeutic agent for a central nervous system disease is provided, which comprises measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein in an animal model treated with the test substance.

[0093] In one embodiment of the present invention, the method may further include, but is not limited to, (S3) a step of selecting the test substance as a candidate therapeutic substance if the expression level of the protein or the gene is decreased.

[0094] In one embodiment of the present invention, the method may further include, but is not limited to, (S3) a step of selecting the test substance as a candidate therapeutic substance if the expression level of the protein or the gene is reduced when compared to a control animal model not treated with the test substance.

[0095] In the present invention, in step (S3) of the method for screening a therapeutic agent for a cranial nervous system disease, the expression level of the protein or the gene is 10 to 30 times, 10 to 28 times, 10 to 26 times, 10 to 24 times, 10 to 23 times, 12 to 30 times, 12 to 28 times, 12 to 26 times, 12 to 24 times, 12 to 23 times, 14 to 30 times, 14 to 28 times, 14 to 26 times, 14 to 24 times, 14 to 23 times, 16 to 28 times, 16 to 24 times, 16 to 23 times, 16 to 24 ... If the test substance exhibits a 30-fold, 16- to 28-fold, 16- to 26-fold, 16- to 24-fold, 16- to 23-fold, 18- to 30-fold, 18- to 28-fold, 18- to 26-fold, 18- to 24-fold, 18- to 23-fold, 20- to 30-fold, 20- to 28-fold, 20- to 26-fold, 20- to 24-fold, 20- to 23-fold, 22- to 30-fold, 22- to 28-fold, 22- to 26-fold, 22- to 24-fold, 22- to 23-fold, or 22.7-fold decrease, the test substance can be selected as a candidate therapeutic substance, but is not limited thereto.

[0096] In the present invention, the animal model may include not only the animal itself that exhibits the disease, but also all biological samples isolated from the animal model. For example, the biological sample means all biological samples defined in the present invention. In one embodiment of the present invention, the expression level of NRXN2 (Neurexin-2) protein or the gene encoding it was measured using exosomes isolated from cerebrospinal fluid, but is not limited thereto.

[0097] In the present invention, the test substance refers to an unknown substance used in screening to administer to an animal model of a cranial nervous system disease and measure the increase or decrease in expression of NRXN2 (Neurexin-2) protein or the gene encoding said protein, and may be one or more selected from the group consisting of nucleotides, DNA, RNA, amino acids, aptamers, proteins, stem cells, stem cell culture medium, compounds, microbial culture medium or extract, natural products, and natural extracts, but is not limited thereto.

[0098] In the present invention, screening can mean selecting a substance having any particular property of interest from a candidate group of various substances using a specific operation or evaluation method.

[0099] In the present invention, the treatment means any action that improves or favorably modifies the target disease and the metabolic disorders caused by it, and can be performed using methods such as chemotherapy, surgery, or biological therapy.

[0100] In the present invention, substances or methods generally used in the art for treating nervous system diseases may be applied. In addition, conventional treatment methods may be used, or commonly used drugs for supporting the treatment of nervous system diseases may be additionally administered, and each candidate substance disclosed in the present invention may be administered, but is not limited thereto.

[0101] In the present invention, treatment means any action that improves or favorably alters the target disease and the metabolic disorders caused by it, and can be performed using methods such as chemotherapy, surgery, or biological therapy.

[0102] In the present invention, substances or methods generally used in the art for treating nervous system diseases may be applied. In addition, conventional treatment methods may be used, or commonly used drugs for supporting the treatment of nervous system diseases may be additionally administered, and the candidate substances disclosed in the present invention may be administered, but are not limited thereto.

[0103] In the present invention, when the term "comprise" is used, it means that other elements may be further included, rather than excluding other elements, unless otherwise specified. The terms "(a) step of" or "a step of" as used throughout the present invention do not mean "a step for".

[0104] Preferred examples are presented below to facilitate understanding of the present invention. However, the following examples are provided merely to facilitate understanding of the present invention, and the content of the present invention is not limited by the following examples.

[0105] [Examples] Materials and Methods Example 1. Criteria for dividing recruitment groups Prior to analysis, cerebrospinal fluid samples were collected from patients with and without subarachnoid hemorrhage. The criteria for separating the two groups were the diagnosis and the extent of cerebral blood flow testing. Clinical information for the patients is listed in Table 1.

[0106] [Table 1]

[0107] Example 2. Method for extracting exosomes from cerebrospinal fluid Without using any chemical mediators, the blood-mixed cerebrospinal fluid was separated into layers using a centrifuge, and then filtered using a 50 nm filter tube.

[0108] The filtered sample was subjected to size exclusion chromatography to extract exosomes, and the exosome particles were collected by centrifuging at 100,000g for 2 hours. The exosomes were then immersed in RIPA buffer to extract the proteins within the exosomes.

[0109] The specific method for extracting exosomes from cerebrospinal fluid (CSF) was as follows. CSF, which is essential for regulating intracranial pressure in patients with subarachnoid hemorrhage, was collected. The blood in the collected CSF was separated into layers by repeated centrifugation without the use of chemical agents. The separated blood was then discarded, and the CSF alone was collected and filtered. The filtered CSF was concentrated using a concentration pump. The concentrated CSF was then filtered through a 50-100 kDa filter. Exosomes were extracted using size exclusion chromatography and an ultracentrifuge. The extracted exosomes were subjected to proteomic analysis to screen for all expressed proteins. The extracted exosomes were identified using techniques such as electron microscopy, nanoparticle tracking analysis (NTA), or NanoFC, and the extracted nanoparticles were further confirmed to be exosomes using exosome-specific markers. Significant protein differences were then confirmed between the control and experimental groups.

[0110] Exosomes are secreted by various cell types, including stem cells, in the form of membrane-bound vesicles measuring 50nm to 200nm in size, reflecting the state of the cells from which they were derived. They are present in easily accessible bodily fluids such as blood, urine, and cerebrospinal fluid, and can be examined for various pathological functions after isolation. To define exosomes, nanoparticle tracking analysis was used to confirm their expression within the size spectrum, and exosome-specific markers (CD63, CD9, etc.) were identified using NanoFC.

[0111] Example 3. Electron microscope photography process The extracted exosomes were fixed and then photographed using an electron microscope (TEM). Specifically, each nanoparticle was identified using a transmission electron microscope (TEM).

[0112] [Experimental Example] Results Transmission electron microscopy of exosomes in the patient's cerebrospinal fluid confirmed that they were almost completely removed from the blood without the use of chemical mediators (Figure 1). This suggests that the exosomes extracted in this invention could be used as ideal biomarkers.

[0113] Through proteomics, the expression levels of approximately 1,500 proteins were confirmed. Among them, the top 10 proteins were classified into groups, and the protein with the most significant difference among the classified groups was the NRXN2 (Neurexin 2) gene. This suggests the possibility of the NRXN2 protein as a diagnostic marker.

[0114] To analyze the proteomic results, quantification was performed using LC-MS / MS analysis on a mass spectrometer (Orbitrap Tribrid) (Figures 2a and 2b). Specifically, using the normalized data from proteomic analysis, we visualized the expression level of NRXN2 by dividing patients into non-SAH and SAH groups. As a result, we confirmed that the expression level of NRXN2 was approximately 22-fold higher (22.7-fold) in SAH patients compared to non-SAH patients. This suggests that NRXN2 can be used as a biomarker for SAH.

[0115] The above description of the present invention is for illustrative purposes only, and those skilled in the art will understand that the present invention can be easily modified into other specific forms without changing the technical concept or essential features of the present invention. Therefore, it should be understood that the above-described embodiments are illustrative in all respects and are not limiting.

Claims

1. A biomarker composition for diagnosing or predicting the prognosis of a neurological disease, comprising an NRXN2 (Neurexin-2) protein or a gene encoding said protein as an active ingredient.

2. 2. The biomarker composition for diagnosing or predicting the prognosis of a cranial nervous system disease according to claim 1, wherein the cranial nervous system disease is any one selected from the group consisting of stroke, cerebral infarction, cerebral embolism, cerebral thrombosis, cerebral arteriosclerosis, cerebral hemorrhage, subarachnoid hemorrhage, Alzheimer's disease, and dementia.

3. A composition for diagnosing or predicting the prognosis of a neurological disease, comprising as an active ingredient a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein.

4. the preparation capable of measuring the expression level of the protein is characterized in that it is any one or more preparations selected from the group consisting of an antibody, a peptide, an aptamer, a protein, and a compound that specifically binds to the protein; The composition for diagnosing or predicting the prognosis of a nervous system disease according to claim 3, characterized in that the preparation capable of measuring the expression level of the gene is any one or more preparations selected from the group consisting of primers, antisense oligonucleotides, and probes that specifically bind to the mRNA of the gene.

5. A kit for diagnosing or predicting the prognosis of a neurological disease, comprising a composition for diagnosing or predicting the prognosis of a neurological disease, the composition containing as an active ingredient a preparation capable of measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein, and instructions.

6. The kit for diagnosing or predicting the prognosis of a cranial nervous system disease comprises:

6. The kit for diagnosing or predicting the prognosis of a cranial nervous system disease according to claim 5, characterized in that the kit is at least one selected from the group consisting of a microarray, an aptamer chip kit, an ELISA (Enzyme Linked Immunosorbent Assay) kit, a blotting kit, an immunoprecipitation kit, an immunofluorescence test kit, a protein chip kit, a reverse transcription polymerase chain reaction (RT-PCR) kit, and a real-time polymerase chain reaction (qRT-PCR) kit.

7. The kit for diagnosing or predicting the prognosis of a cranial nervous system disease described in claim 5, wherein the instructions instruct that if the expression level of NRXN2 (Neurexin-2) protein or the gene encoding said protein is higher in a patient than in a control group, the patient is determined to be a patient with a cranial nervous system disease.

8. (S1) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein from a biological sample isolated from an individual; and (S2) comparing the expression level of the protein or the gene with a control group; A method for providing information necessary for diagnosing or predicting the prognosis of a nervous system disease.

9. The method for providing information necessary for the diagnosis or prognosis of a cranial nervous system disease according to claim 8, characterized in that the method further comprises: (S3) determining that the patient is a cranial nervous system disease patient if the expression level of the protein or the gene is higher in the patient than in a control group in step (S2).

10. The expression level of the protein is measured by one or more methods selected from the group consisting of Western blot, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), radial immunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunohistochemistry (IHC), immunoprecipitation analysis, complement fixation analysis, flow cytometry (Fluorescence Activated Cell Sorter, FACS), and protein chip; The mRNA levels of the genes were measured using reverse transcription polymerase chain reaction (RT-PCR), competitive reverse transcription polymerase chain reaction (competitive RT-PCR), real-time quantitative RT-PCR, multiplex reverse transcription polymerase chain reaction (multiplex PCR), real-time polymerase chain reaction (qRT-PCR), RNase protection assay, Northern blotting, DNA chip analysis, methylated DNA binding domain sequencing (MBDseq) assay, and reduced representation bisulfite sequencing (RRBS).

9. The method for providing information necessary for the diagnosis or prognosis of a neurological disease according to claim 8, wherein the measurement is performed by any one or more methods selected from the group consisting of (a) nucleotide sequencing (nucleotide sequencing) analysis methods.

11. The method for providing information necessary for the diagnosis or prognosis of a cranial nervous system disease according to claim 8, wherein the biological sample is at least one selected from the group consisting of cerebrospinal fluid, exosomes, neurons, immune cells, blood, serum, whole blood, plasma, urine, saliva, tissue, cells, organs, bone marrow, fine needle aspiration specimens, core needle biopsy specimens, and vacuum aspiration biopsy specimens.

12. (S1) treating an animal model of a patient group of cranial nervous system diseases with a test substance; and (S2) measuring the expression level of NRXN2 (Neurexin-2) protein or a gene encoding said protein in an animal model treated with the test substance.

13. The method for screening a therapeutic agent for a cranial nervous system disease according to claim 12, further comprising: (S3) selecting the test substance as a candidate therapeutic substance if the expression level of the protein or the gene is decreased.

Images