WO2015174544A1 - Mental illness determination marker - Google Patents

Mental illness determination marker Download PDF

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WO2015174544A1
WO2015174544A1 PCT/JP2015/064233 JP2015064233W WO2015174544A1 WO 2015174544 A1 WO2015174544 A1 WO 2015174544A1 JP 2015064233 W JP2015064233 W JP 2015064233W WO 2015174544 A1 WO2015174544 A1 WO 2015174544A1
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group
schizophrenia
depression
determination marker
seq
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PCT/JP2015/064233
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French (fr)
Japanese (ja)
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功太郎 服部
浩 功刀
後藤　雄一
新一 ▲高▼坂
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国立研究開発法人国立精神・神経医療研究センター
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Priority to JP2016519420A priority Critical patent/JPWO2015174544A1/en
Publication of WO2015174544A1 publication Critical patent/WO2015174544A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/115—Aptamers, i.e. nucleic acids binding a target molecule specifically and with high affinity without hybridising therewith ; Nucleic acids binding to non-nucleic acids, e.g. aptamers

Definitions

  • the present invention relates to a biomarker for determining the presence / absence of psychiatric schizophrenia, bipolar disorder or depression using a sample derived from a subject, and a mental disease determination method using the biomarker.
  • diagnosis of mental illness is performed based on information such as symptoms, signs, and progress obtained through interviews with the person and family.
  • DSM Diagnostic and Statistical Manual of Mental Disorders
  • This diagnostic criterion does not use an objective test, it often does not agree with the doctor who makes the diagnosis.
  • the structured diagnosis interview is a manual interview method in which the format of the question content, the re-question, etc.
  • Non-Patent Document 2 Non-Patent Document 2
  • the structured diagnostic interview requires a high degree of skill of the diagnostician and an interview time of 1 to 2 hours.
  • DSM diagnosis is determined for a disorder, it cannot be said that it necessarily reflects biological pathology. In fact, even if the pathology is clearly related to a cause other than the brain, there are often cases in which a DSM diagnosis is mistakenly diagnosed as a mental illness. For example, at least 10% of patients diagnosed with depression are said to have side effects of hypothyroidism and anti-inflammatory drugs (Non-patent Document 3). In addition, it has been reported that patients diagnosed with schizophrenia include anti-NMDA receptor antibody encephalitis (Non-patent Document 4).
  • Non-patent Document 5 the current antidepressant has not been sufficiently effective except in some severe cases.
  • the effects of schizophrenia drugs are limited to some symptoms such as hallucinations and delusions.
  • the classification of mental illness is not well established. For example, there are multiple pathologies that can be subdivided into mental illnesses diagnosed with depression, and only some of the currently prescribed antidepressants and depression have therapeutic effects. is there. Therefore, the development of an essential therapy for mental illness requires the development of a new classification based on biological evidence and an appropriate diagnostic technique.
  • a biomarker is a substance such as a nucleic acid or protein produced by a cell in response to the onset of a specific disease, and the presence, severity, or prognosis of an abundance or variation in body fluids or tissues. Therefore, it can be a discriminator for disease determination and treatment guideline determination.
  • Non-Patent Documents 6 and 7 In the field of mental illness, biomarker searches for schizophrenia, depression, or bipolar disorder have been conducted so far, and various candidate molecules such as cytokines and neurotrophic factors have been proposed. However, most of them do not have a sufficient detection effect, and practical biomarkers for mental illness have not been developed yet (Non-Patent Documents 6 and 7).
  • An object of the present invention is to develop and provide a method for accurately detecting the presence or absence of each mental illness in a subject using the mental illness determination marker.
  • a problem of diagnostic criteria is that, when searching for a marker, candidate molecules are selected based only on the significant difference between the average values of the patient group and the healthy group.
  • Mental illness is a syndrome, and it is likely that there are multiple conditions in the syndrome. Even if it can be a useful biomarker for some disease states, if the proportion of the disease state in all cases is small, it may be buried in the significant difference test and excluded.
  • Another problem with the sample is that blood or postmortem brain was used as a marker search sample. Blood directly contacts various organs such as the brain, intestines, liver, and muscles.
  • the postmortem brain is superior in that it uses the target organ itself, which is thought to be the cause of mental illness, and maintains the three-dimensional structure of the brain, but unlike animal experiments that can be removed immediately after death, the human brain is the cause of death. And changes due to postmortem changes are likely to make it difficult to detect changes in important candidate molecules such as neurotransmitters and signal molecules. Furthermore, in neuropathology and brain imaging research, no foci specific to each mental disorder have been found so far, and unlike Parkinson's disease, the superiority of maintaining the three-dimensional structure of the brain is poor.
  • CSF cerebrospinal fluid
  • a psychiatric disease determination marker comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 1 to 249 contained in cerebrospinal fluid, a mutant thereof, or a fragment thereof. (2) selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 1, 3, 5, 6, 7, 8-13, 15-33, 35-54, 56-60, 141-144, 147-162, and 249
  • the psychiatric disorder determination marker according to (2) comprising: (4) selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 9, 13, 26-29, 31-33, 35-54, 56-60, and 159-162, which identify the severity of schizophrenia
  • the psychiatric disease determination marker according to (2) comprising a protein, a mutant thereof, or a fragment thereof.
  • the psychiatric disorder determination marker according to (1) comprising a fragment, wherein the psychiatric disorder is depression.
  • the psychiatric disease determination marker according to (8) comprising: (10) The protein according to (8), comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 44, 66, and 88 to 93, a variant thereof, or a fragment thereof, which identifies the severity of depression A marker for determining mental illness.
  • the protein according to (1) comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 94 to 98, a variant thereof, or a fragment thereof, wherein the mental illness is schizophrenia / bipolar disorder Psychiatric disease determination marker.
  • a mental disorder comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 2, 4, 55, 99 to 103, 145, 146, 179, and 233 to 240, a variant thereof, or a fragment thereof.
  • the psychiatric disorder determination marker according to (1) wherein is schizophrenia / depression.
  • a severity determination step of measuring the amount of the psychiatric disorder determination marker according to (7) in the measurement step and determining the severity of bipolar disorder based on the measurement value obtained in the measurement step The method according to (21) or (22). (31) It further includes a severity determination step of measuring the amount of the psychiatric disorder determination marker according to (10) in the measurement step and determining the severity of depression based on the measurement value obtained in the measurement step. , (23) or (24).
  • the amount of the psychiatric disease determination marker according to (15) is measured, and based on the measurement value obtained in the measurement step in the comparison determination step, the schizophrenia or bipolar disorder
  • the amount of the psychiatric disease determination marker according to (16) is measured, and whether the depression or bipolar disorder is based on the measurement value obtained in the measurement step in the comparison determination step
  • a biomarker capable of easily and accurately determining a psychiatric disease such as schizophrenia, depression, or bipolar disorder in a subject.
  • This psychiatric disorder determination marker can be used to identify morbidity such as schizophrenia, depression, or bipolar disorder in a subject, identification of subtypes, and / or identification of severity.
  • the mental illness determination method of the present invention there is provided a method for accurately determining morbidity, subtype identification, and / or severity identification of each mental illness in a subject using the mental illness determination marker. can do.
  • FIG. 1 It is a conceptual diagram which shows the classification group of the mental disease determination marker of this invention.
  • the distribution chart of the measured value of the depression psychiatric disorder determination marker SCFsR in healthy individuals and patients with each psychiatric disorder is shown.
  • a horizontal straight line in each plot group indicates an average value in the plot group (the same applies to other drawings).
  • N indicates the total number of patients with the mental illness
  • DF indicates the number of drug-free patients in N (the same applies to other figures).
  • the distribution chart of the measured values in healthy individuals and patients with each mental illness of the mental illness determination marker UNC5H3 that subdivides schizophrenia into subtypes is shown.
  • the broken line (95%) is a cutoff value corresponding to the 95th percentile in the healthy group.
  • the relationship between the measured value of ST4S6, which is a psychiatric disorder determination marker for determining the severity of schizophrenia, and the evaluation value based on the schizophrenia evaluation scale PANSS is shown.
  • the distribution chart of the measured value of the LYN B marker for mental disorders for depression / bipolar disorder in healthy individuals and patients with various mental disorders is shown. It is the figure which entered 95th percentile in a healthy subject group as a cut-off value in the measured value distribution map of LYN
  • PIGF was remarkably enhanced in the schizophrenia group (Sz) and a part of depression. Therefore, by using PIGF as a psychiatric disorder determination marker, it is possible to determine the possibility and severity of schizophrenia.
  • It is a figure which shows the measured value distribution of (alpha) 1-Antitrypsin which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of ⁇ 1-Antitrypsin of the first sample cohort shown in Table 12 and
  • (b) shows the second sample cohort shown in Table 13.
  • C) is the figure which combined the data of (a) and (b) by Z conversion.
  • Cont indicates a healthy person
  • Sz indicates schizophrenia
  • MDD indicates depression
  • BD indicates bipolar disorder.
  • Adiponectin which is a schizophrenia / depression mental disease determination marker.
  • A shows the measured value distribution of Adiponectin in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Adiponectin in the second sample cohort shown in Table 13.
  • C is the figure which combined the data of (a) and (b) by Z conversion.
  • A shows the measured value distribution of Antithrombin III which is a mental disease determination marker for depression.
  • A) shows the measured value distribution of Antithrombin III in the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Apo B which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the measured value distribution of Apo B of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of ATS13 which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of ATS13 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of ATS13 of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of b-ECGF which is a bipolar disorder / depression mental disease determination marker.
  • (A) shows the measured value distribution of b-ECGF in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of b-ECGF in the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of C1-Esterase (TM) Inhibitor which is a mental disease determination marker for depression.
  • TM C1-Esterase
  • (A) shows the measured value distribution of the C1-Esterase Inhibitor of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of C3a which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the measured value distribution of C3a of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of C3a of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of C5 which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the measured value distribution of C5 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of C5 of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. In both figures, there was a marked increase in some depression (MDD).
  • (D) is a figure which shows the correlation of the measured value of C5 by ELISA, and the measured value by SOMAscan. It is a figure which shows the measured value distribution of CD5L which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of CD5L of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of CD5L of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Contactin-4 which is a mental disorder determination marker for schizophrenia / depression.
  • (A) shows the measured value distribution of Contactin-4 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Contactin-4 of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of CRP which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the CRP measurement value distribution of the first sample cohort shown in Table 12, and (b) shows the CRP measurement value distribution of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. In both figures, there was a marked increase in some depression (MDD).
  • D is a figure which shows the correlation of the measured value of CRP by ELISA, and the measured value by SOMAscan. It is a figure which shows the measured value distribution of Cystatin (C) which is a schizophrenia / depression mental disease determination marker.
  • A shows the measured value distribution of Cystatin C of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Ephrin-A5 which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of Ephrin-A5 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of ESAM which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the ESAM measurement distribution of the first sample cohort shown in Table 12, and (b) shows the ESAM measurement value distribution of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of FGF-12 which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of FGF-12 in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of FGF-12 in the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of FSTL3 which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of FSTL3 of the first sample cohort shown in Table 12, and (b) shows the FSTL3 measured value distribution of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of GDF-9 which is a mental disease determination marker for schizophrenia.
  • (A) shows the measured value distribution of GDF-9 in the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of GP1BA which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the measured value distribution of GP1BA of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of GP1BA of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of IL-13 (R) Ra1 which is a psychiatric disorder determination marker for schizophrenia.
  • (A) shows the measured value distribution of IL-13 Ra1 of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of JAM-B which is a psychiatric disorder determination marker for schizophrenia / depression / bipolar disorder.
  • (A) shows the measured value distribution of JAM-B of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of MATN2 which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the measured value distribution of MATN2 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MATN2 of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of MEK1 which is a psychiatric disease determination marker for bipolar disorders.
  • (A) shows the measured value distribution of MEK1 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MEK1 of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Midkine which is a mental disease determination marker for bipolar disorders.
  • (A) shows the measured value distribution of the Midkine of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of MIF which is a psychiatric disorder determination marker for bipolar disorders.
  • (A) shows the MIF measurement value distribution of the first sample cohort shown in Table 12, and (b) shows the MIF measurement value distribution of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of MIS which is a mental disease determination marker for depression.
  • (A) shows the MIS measurement value distribution of the first sample cohort shown in Table 12, and (b) shows the MIS measurement value distribution of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of MMP-2 which is a mental disease determination marker for schizophrenia.
  • (A) shows the measured value distribution of MMP-2 in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MMP-2 in the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of MSP which is a psychiatric disease determination marker for bipolar disorders.
  • (A) shows the measured value distribution of MSP of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MSP of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Nectin-like protein 1 which is a mental illness determination marker for depression.
  • (A) shows the measured value distribution of Nectin-like protein 1 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Notch * 1 which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the measured value distribution of Notch 1 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Properdin which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of Properdin of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Properdin of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of TFPI which is a psychiatric disorder determination marker for bipolar disorders.
  • (A) shows the measured value distribution of TFPI of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of TFPI of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of TIG2 which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of TIG2 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of TIG2 of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of Transketolase which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of Transketolase in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Transketolase in the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of TSG-6 which is a schizophrenia / depression mental disease determination marker.
  • (A) shows the measured value distribution of TSG-6 in the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of TWEAK which is a mental disease determination marker for depression.
  • (A) shows the measured value distribution of TWEAK of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of TWEAK of the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion. It is a figure which shows the measured value distribution of UNC5H3 which is a schizophrenia mental disease determination marker.
  • (A) shows the measured value distribution of UNC5H3 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13.
  • (C) is the figure which combined the data of (a) and (b) by Z conversion.
  • Vasoactive®Intestinal®Peptide which is a psychiatric disease determination marker for schizophrenia / depression / bipolar disorder.
  • A shows the measured value distribution of Vasoactive Intestinal Peptide of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13.
  • C is the figure which combined the data of (a) and (b) by Z conversion.
  • Psychiatric disease determination marker 1-1 Outline
  • the psychiatric disorder determination marker of the present invention is composed of a protein or a fragment thereof, and can be used as a biomarker for determining a psychiatric disorder in a subject by measuring the amount contained in a subject-derived sample.
  • Psychiatric disorder broadly refers to a generic term for exogenous, psychogenic, or intrinsic brain functional or organic disorders. However, unless otherwise specified, the term “psychiatric disorder” in the present specification refers to FIG. 1 belonging to three syndromes classified as schizophrenia, depression, or bipolar disorder in the diagnostic category of conventional psychopathology. Means a narrowly defined mental illness.
  • determining (determining) a mental illness refers to identifying a morbidity of a psychiatric disorder in a subject, identifying a subtype of the psychiatric disorder, and / or identifying the severity of the afflicted mental disorder. .
  • the “mental disease determination marker” is a protein selected from the group consisting of the amino acid sequences shown in each SEQ ID NO. In Table 1, a variant thereof, or a fragment thereof for determining a mental disease. It is a biomarker.
  • All of the proteins consisting of the amino acid sequences shown in each SEQ ID No. in Table 1 are proteins derived from human cerebrospinal fluid (CSF), and in the present invention, any one of schizophrenia, depression or bipolar disorder in a subject. It can be a biomarker for determining the above mental disorders.
  • “UniProt No.” shown in Table 1 is the accession number of each protein disclosed in “http://www.uniprot.org/”.
  • variant thereof refers to a variant of each protein comprising the amino acid sequence shown in each SEQ ID NO in Table 1. Specifically, in the amino acid sequence of each protein shown in Table 1, a polypeptide containing addition, deletion, or substitution of one or more amino acids, or 80% or more of the amino acid sequence of each protein shown in Table 1, A polypeptide having 85% or more, preferably 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more amino acid identity.
  • “plurality” refers to 2 to 10, 2 to 9, 2 to 8, 2 to 7, 2 to 6, 2 to 5, 2 to 4, or 2 or 3 integers.
  • amino acid identity refers to each of the amino acid sequences shown in Table 1 when the two amino acid sequences are aligned (aligned), and a gap is introduced as necessary so that the degree of amino acid coincidence between the two is maximized. The ratio (%) of the same amino acid residue of the total number of amino acid residues of the variant with respect to the total number of amino acid residues of the protein.
  • Amino acid identity can be determined using protein search systems with BLAST and FASTA (Karlin, S. et al., 1993, Proc. Natl. Acad. Sci. USA, 90: 5873-5877; Altschul, S.F. et al., 1990, J. Mol.
  • the “fragment” is a peptide fragment consisting of a protein selected from the group consisting of the amino acid sequences shown in each SEQ ID No. in Table 1 or a part of a variant thereof, and the amino acid sequence constituting the protein Among them, 7 or more, less than 100, 10 or more, less than 15, 15 or more, less than 100, preferably 20 or more, less than 100, 25 or more, less than 35, 35 or more, less than 40, or more than 40, or 50 It has less than the total number of consecutive amino acid residues.
  • it is a polypeptide fragment carrying one or more epitopes within its amino acid sequence.
  • the “biomarker composed of a protein selected from the group consisting of the amino acid sequence shown in SEQ ID NO: X, a variant thereof, or a fragment thereof” in Table 1 is referred to as “amino acid sequence shown in SEQ ID NO: X”.
  • X is an arbitrary integer from 1 to 140.
  • the “subject” refers to a human individual that is subjected to the method of the second aspect described later and is a target for which a mental illness is to be determined.
  • the mental disease determination marker of the present invention can determine one or more mental diseases of schizophrenia, depression, or bipolar disorder in a subject.
  • the mental disease determination markers shown in Tables 2 to 5, 7, 9, and 11 below can be roughly classified into groups 1 to 7 shown in FIG. 1 based on their detection attributes.
  • Group to determine schizophrenia Table 2 shows markers for determining mental disorders belonging to Group 1.
  • the mental disease determination marker belonging to this group is a marker for determining schizophrenia. Specifically, it is a marker belonging to the section shown in group 1 in FIG. 1, and SEQ ID NOs: 1, 3, 5, 6, 7, 8 to 13, 15 to 33, 35 to 54, 56 to 60, 141 to A protein selected from the group consisting of the amino acid sequences represented by 144, 147 to 162, and 249.
  • a group consisting of the amino acid sequences shown in SEQ ID NOs: 1, 3, 5, 6, 7, 8 to 13, 15 to 25, 30, 46, 141 to 144, 147 to 158, and 249 in the psychiatric disorder determination marker belonging to this group Or the like can be used to distinguish the morbidity of schizophrenia in a subject.
  • “differentiating (performing)” means determining the high possibility of the subject having a specific mental illness. Therefore, a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 1, 3, 5, 6, 7, 8 to 13, 15 to 25, 30, 46, 141 to 144, 147 to 158, and 249, etc. It is a schizophrenia disease differentiation marker, and by using one or more of these markers, it can be distinguished that a subject is highly likely to suffer from schizophrenia.
  • Proteins selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 1, 3, 5, 6, 11, 12, 30, 46, 141, 142, 149 to 158, and 249 in the psychiatric disorder determination marker belonging to this group Is a biomarker isolated based on a significant difference in the abundance of the protein or the like contained in a cerebrospinal fluid sample between a schizophrenic patient and a healthy subject.
  • the term “healthy state” refers to a state that is not affected by any mental illness in the narrow sense herein, preferably a healthy mental state that is not affected by any mental illness in the broad sense.
  • the risk rate means statistically significant. Specifically, when the difference between the measured values of the subject and the healthy person is statistically processed, it means that there is a significant difference between the two.
  • the risk rate may be less than 5%, 1%, 0.3%, 0.2%, or 0.1%.
  • the test method for statistical processing is not particularly limited as long as a known test method capable of determining the presence or absence of significance is appropriately used. For example, Student's t test or multiple comparison test can be used.
  • Proteins selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 8 to 13, 15 to 25, 141 to 144, 147, 148, and 249 in the psychiatric disorder determination marker belonging to this group are used to identify morbidity of schizophrenia. In addition, it is a determination marker that can subdivide schizophrenia into subtypes.
  • the “subtype” is a subordinate classification group of each mental illness classified based on a biological basis using the mental illness determination marker of the present invention.
  • schizophrenia subtypes belonging to group 1 bipolar disorder subtypes belonging to group 2, subgroups belonging to group 3 and depression.
  • Other examples include schizophrenia / bipolar subtypes belonging to group 4, subtypes of schizophrenia / depression belonging to group 5, bipolar disorder / depression subtypes belonging to group 6 .
  • Such a subtype is classified, for example, as “(mental marker molecule name) subtype of (psychiatric disorder)”.
  • the subject determined as schizophrenia can be identified as “UNC5H3 subtype of schizophrenia”. Details of schizophrenia / bipolar disorder belonging to group 4 and schizophrenia / depression belonging to group 5, and bipolar disorder / depression belonging to group 6 will be described later.
  • Mental illness determination marker capable of identifying a subtype of each psychiatric disorder in the present specification (specifically, SEQ ID NOs: 7 to 13, 15 to 25, 141 to 144, 147, 148 for schizophrenia subtypes, and A protein selected from the group consisting of amino acid sequences shown by H.249, a protein selected from the group consisting of amino acid sequences shown by SEQ ID NOs: 63, 64, 114, and 163 to 169 for bipolar disorder subtypes described later, A protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 66, 76-87, and 174-178, and 180-192 for the depression subtype, SEQ ID NO: 95 for the schizophrenia / bipolar disorder subtype SEQ ID NOs: 2, 4, 7, 100 to 103, 145, 146 for schizophrenia / depression subtype, such as a protein selected from the group consisting of the amino acid sequences represented by ⁇ 98 179 and a protein selected from the group consisting of amino acid sequences represented
  • the “cut-off value” refers to a boundary value for classifying a quantitative result into positive and negative.
  • positive means that there is a high possibility of suffering from a mental illness
  • negative means that there is a high possibility that the person is not affected.
  • the method for setting the cutoff value is not particularly limited.
  • measurement values obtained from a group of healthy subjects can be classified by percentile, and the percentile value used for the classification can be used as a cutoff value.
  • the 95th percentile is a cut-off value.
  • it can be set based on the calculation method described in Akobeng AKActa, 2007, Paediatr, 96: 644-647.
  • the percentile value when using the percentile value as a cut-off value in this specification, use either the 5th percentile or the 95th percentile, positive if less than 5th percentile or greater than 95th percentile, greater than 5th percentile, or less than 95th percentile Negative.
  • the percentile value when using the percentile value as a cut-off value in this specification, use either the 5th percentile or the 95th percentile, positive if less than 5th percentile or greater than 95th percentile, greater than 5th percentile, or less than 95th percentile Negative.
  • a UNC5H3 marker measurement in a subject shows a value greater than the 95th percentile of a healthy group, the subject is likely to have schizophrenia and schizophrenia Can be subdivided into UNC5H3 subtypes.
  • the mental illness determination marker of the present invention which can subdivide each mental illness into subtypes, not only follows the conventional hypothetical mental illness diagnostic criteria but also reflects the molecular pathology of each psychiatric illness. Classification with higher objectivity based on the basis is possible.
  • the use of the psychiatric disorder determination marker of the present invention enables optimal treatment according to each subtype that may exist in the same psychiatric disorder.
  • the above psychiatric disease markers that can be subdivided into subtypes have high psychiatric disease specificity and can accurately determine schizophrenia, bipolar disorder, or depression, and are not newly classified by conventional methods.
  • Psychiatric disorders such as schizophrenia / bipolar disorder, schizophrenia / depression, and depression / bipolar disorder can also be determined with high accuracy.
  • the 95th percentile and the 5th percentile of healthy subjects are cut off values, and the psychiatric disease determination marker that can subdivide proteins that are out of that value and show abnormal values greater than 95th percentile or less than 5th percentile into subtypes.
  • a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 7 to 13, 15 to 20, 141 to 144, 147, 148, and 249 has the 95th percentile of a healthy person as a cutoff value, and the value If it is larger than that, it is possible to subdivide schizophrenia into subtypes in addition to the symptom diagnosis of schizophrenia.
  • the protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 21 to 25 has a cut-off value of the 5th percentile of healthy individuals, and if it is less than that value, it is added to the symptom diagnosis of schizophrenia Schizophrenia can be subdivided into subtypes.
  • psychiatric disorder determination marker belonging to this group, measured values of proteins and the like selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 9, 13, 26 to 34, 35 to 54, 56 to 60, and 159 to 162 are , Showing a correlation with the schizophrenia rating scale. Therefore, the severity of schizophrenia can be identified.
  • severity refers to the degree of symptoms based on an evaluation scale for each mental disorder.
  • PANSS Positive and Negative Syndrome Scale
  • BACS Brief Assessment for Cognition in Schizophrenia
  • Evaluation scale showing correlation with each psychiatric disease determination marker selected from the group consisting of amino acid sequences shown in SEQ ID NOs: 9, 13, 26-29, 31-33, 35-54, 56-60, and 159-162 Is a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 26 to 33, which is correlated with p ⁇ 0.01 in Pearson correlation analysis using PANSS as an evaluation scale, and severe schizophrenia
  • a protein or the like selected from the group is a psychiatric disease determination marker that can identify the severity of schizophrenia that correlates with P ⁇ 0.01 in the Pearson correlation function using BACS as an evaluation scale.
  • the protein or the like selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 9 and 13 is not only the severity of schizophrenia but also schizophrenia It is a psychiatric disease determination marker that can also be used for differentiation of diseases and identification of subtypes.
  • the mental disease determination marker belonging to this group is a marker that can determine bipolar disorder. Specifically, it refers to a protein or the like belonging to the compartments shown in Group 2 in FIG. 1 and selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 47, 61 to 69, 114, and 163 to 173. Using this group of psychiatric disease determination markers, it is possible to identify the presence of bipolar disorder in a subject, identify the subtype of bipolar disorder, and / or identify the severity of bipolar disorder.
  • Proteins selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 61 to 64, 114, and 163 to 172 in the psychiatric disease determination marker belonging to this group can distinguish the presence of bipolar disorder in the subject.
  • the protein comprising the amino acid sequences represented by SEQ ID NOs: 61, 62, and 170 to 172 is a significant difference in the abundance of the protein or the like contained in a cerebrospinal fluid-derived sample between a bipolar disorder patient and a healthy subject. Is a biomarker isolated on the basis of
  • proteins comprising the amino acid sequences represented by SEQ ID NOs: 63, 64, 114, and 163 to 169 are biomarkers that can subdivide bipolar disorder into subtypes.
  • Table 3 the 95th percentile and the 5th percentile of healthy individuals are shown as cut-off values, and proteins showing abnormal values greater than or less than the 95th percentile are shown as psychiatric disease determination markers that can be subdivided into subtypes.
  • the protein consisting of the amino acid sequences shown in SEQ ID NOs: 63, 64, 164, 166, and 168 has a cutoff value of the 5th percentile of healthy subjects, and if it is less than that value, it is classified as affected by bipolar disorder.
  • bipolar disorder can be subdivided into subtypes.
  • the protein consisting of the amino acid sequences shown in SEQ ID NOs: 114, 163, 165, 167, and 169 has a 95th percentile of a healthy person as a cut-off value, and if it is larger than that value, it is classified as a disease disorder of bipolar disorder.
  • bipolar disorder can be subdivided into subtypes.
  • the measured values of proteins and the like selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 47 and 65 to 69 in the psychiatric disorder determination marker belonging to this group are HAM-, which is an evaluation scale for depressive symptoms in bipolar disorder Since it is correlated with D (Hamilton Depression Scale), one aspect of its severity can be identified.
  • a protein or the like selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 47, 65 to 69, and 173 is a bipolar protein that correlates with p ⁇ 0.01 in the Pearson correlation function using HAM-D as an evaluation measure. It is a psychiatric disorder determination marker that can identify the severity of a disorder.
  • the psychiatric disorder determination marker Activin beta A represented by SEQ ID NO: 47 can identify the severity of schizophrenia as described above when the evaluation scale is BACS.
  • Group for Determining Depression Table 4 shows psychiatric disease determination markers belonging to Group 3.
  • the mental disease determination marker belonging to this group is a marker that can determine depression. Specifically, it is a marker belonging to the section shown in group 3 in FIG. 1 and represented by SEQ ID NOs: 13, 14, 17, 44, 56, 66, 70 to 93, 123, 174 to 178, and 180 to 232 A protein or the like selected from the group consisting of amino acid sequences. Using this group of psychiatric disease determination markers, it is possible to identify the morbidity of depression, identify the subtype of depression, and / or identify the severity of depression in the subject.
  • Proteins selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 13, 14, 17, 56, 66, 70 to 87, 123, and 174 to 178, and 180 to 232 in the mental disease determination marker belonging to this group It is possible to differentiate the morbidity of depression in a subject.
  • proteins selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 13, 14, 17, 56, 70 to 75, 82, 83, 123, 174 to 178, 180, and 193 to 232 are depression, etc. It is a biomarker isolated based on a significant difference in the abundance of the protein or the like contained in a cerebrospinal fluid-derived sample between a patient and a healthy person.
  • the psychiatric disorder determination marker represented by SEQ ID NO: 2 is also a psychiatric disorder determination marker for morbidity discrimination based on a significant difference in schizophrenia
  • the psychiatric disorder determination marker represented by SEQ ID NOs: 13 and 17 is a schizophrenia marker.
  • It is also a mental disease determination marker for subtype identification
  • the mental disease determination marker represented by SEQ ID NO: 56 is also a mental disease determination marker for identifying the severity of schizophrenia.
  • proteins selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 66, 76 to 87, 174 to 178, and 180 to 192 are biomarkers that can subdivide depression into subtypes.
  • Table 4 the 95th percentile and the 5th percentile of healthy subjects are shown as cut-off values, and proteins showing abnormal values larger than the 95th percentile or less than the 5th percentile are shown as psychiatric disease determination markers that can be subdivided into subtypes.
  • a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 66, 76 to 81, 176 to 178, 181 to 185, and 189 to 192 has the 95th percentile of a healthy person as a cutoff value
  • depression can be subdivided into subtypes in addition to identifying the presence of depression.
  • a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 82 to 87, 174, 175, 180, and 186 to 188 has a 5th percentile of a healthy person as a cutoff value, and is less than that value Can subdivide depression into subtypes in addition to identifying depression.
  • the psychiatric disease determination marker Fibrinogen represented by SEQ ID NO: 66 is a psychiatric disease determination marker that can identify the severity of bipolar disorder when the evaluation scale HAM-D is used. When identified based on the cut-off value determined based on the measured value of fibrinogen contained in the spinal fluid-derived sample, it can be a marker for identifying the morbidity and severity of depression.
  • the measured values of proteins and the like selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 44, 66, and 88 to 93 in the psychiatric disease determination marker belonging to this group are correlated with the depression evaluation scale HAM-D. From this, the severity of depression can be determined.
  • the protein or the like is a psychiatric disease determination marker that can identify the severity of depression that correlates with p ⁇ 0.01 in the Pearson correlation function using HAM-D as an evaluation scale.
  • Table 5 shows the markers for determining mental disorders belonging to Group 4.
  • the psychiatric disorder determination marker belonging to this group is a marker that can determine schizophrenia / bipolar disorder.
  • “Schizophrenia / bipolar disorder” is a psychiatric disorder that has been classified as one of the psychiatric disorders by the conventional classification method, and is integrated by the determination method of the present invention based on the biological basis using biomarkers. It refers to a new group of mental illnesses that have the characteristics of both schizophrenia and bipolar disorder and cannot be clearly classified into either one. Specifically, it refers to a protein or the like belonging to the compartments shown in Group 4 in FIG. 1 and selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 94 to 98. Use of this group of psychiatric disease determination markers can identify that a subject is likely to have schizophrenia / bipolar disorder.
  • a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 95 to 98 in the psychiatric disorder determination marker belonging to this group can be identified as having a high possibility of suffering from schizophrenia / bipolar disorder, It is a determination marker that can subdivide schizophrenia / bipolar disorder into subtypes.
  • the mental disease determination marker belonging to this group may be classified based on the cut-off value set in the same manner as the subtype identification of the mental disease in the first to third groups.
  • the examiner is likely to have schizophrenia / bipolar disorder and can be identified as “BMP-1 subtype of schizophrenia / bipolar disorder”.
  • Table 5 shows the 95th percentile and 5th percentile of healthy subjects as cut-off values, and proteins showing abnormal values greater than 95th percentile or less than 5th percentile as psychiatric disease determination markers that can be subdivided into subtypes.
  • the BMP-1 protein consisting of the amino acid sequence represented by SEQ ID NO: 95 is determined to be schizophrenia or bipolar disorder when the 95th percentile of a healthy person is set as a cutoff value, and is larger than that value. Can do.
  • a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 96 to 98 has schizophrenia / bipolar disorder when the 5th percentile of healthy subjects has a cut-off value, and is less than that value Can be determined.
  • a supplementary psychiatric disorder determination marker capable of determining whether a subject suffers from schizophrenia or bipolar disorder, from the group consisting of amino acid sequences represented by SEQ ID NOs: 127 to 131 and 242-246 You may use the protein etc. which are selected. Such supplemental markers are shown in Table 6.
  • the markers shown in Table 6 cannot be used alone to distinguish between patients with schizophrenia or patients with bipolar disorder because the differences between healthy individuals and each disease are small. However, if the patient is suffering from a psychiatric disorder, either schizophrenia or bipolar disorder, the two can be differentiated.
  • a protein comprising the amino acid sequence represented by SEQ ID NO: 127, 128, or 130 is schizophrenia when the concentration is lower than the cut-off value, and bipolar disorder when the concentration is high. When the concentration is higher than the cut-off value, schizophrenia can be diagnosed, and bipolar disorder can be diagnosed when the concentration is lower than the cutoff value.
  • Table 7 shows the markers for determining mental disorders belonging to Group 5.
  • the psychiatric disorder determination marker belonging to this group is a marker that can determine schizophrenia / depression.
  • “Schizophrenia / depression” is a mental disorder that has been classified as one of the mental disorders according to the conventional classification method. It refers to a new group of mental illnesses that have the characteristics of both mental disorders and depression and cannot be clearly classified into either one. Specifically, from the group consisting of the amino acid sequences shown in SEQ ID NOs: 2, 4, 55, 99 to 103, 145, 146, 179, and 233 to 240, which belong to the compartment shown by group 5 in FIG. It refers to the selected protein. Use of this group of mental illness determination markers can distinguish that a subject is likely to have schizophrenia / depression.
  • Proteins selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 2, 4, 101 to 103, 145, 146, 179, and 233 to 239 in the psychiatric disorder determination marker belonging to this group are schizophrenia / depression. It is a determination marker that can distinguish a high possibility of suffering from schizophrenia and subdivide schizophrenia / depression into subtypes.
  • the mental disease determination marker belonging to this group may be classified based on the cut-off value set in the same manner as the subtype identification of the mental disease in the first to third groups.
  • the subject in the determination of mental illness using the NADPH-P450 Oxidoreductase marker represented by SEQ ID NO: 101, when the measured value of the NADPH-P450 Oxidoreductase marker in the subject shows a value greater than the 95th percentile of the measured values of the healthy group, The subject is likely to have schizophrenia / depression and can be identified as “NADPH-P450 Oxidoreductase subtype of schizophrenia / depression”.
  • the 95th percentile and the 5th percentile of healthy subjects are cut off values, and the psychiatric disease determination marker that can subdivide proteins that are out of that value and show abnormal values greater than 95th percentile or less than 5th percentile into subtypes As shown.
  • NADPH-P450 Oxidoreductase consisting of the amino acid sequences represented by SEQ ID NOs: 101, 145, 146, 179, and 233 to 237 has a cut-off value of the 95th percentile of a healthy person, and if greater than that value, schizophrenia Can be determined to be symptom / depression.
  • the protein consisting of the amino acid sequences represented by SEQ ID NOs: 102, 103, 138, and 139 has a cutoff value of the 5th percentile of healthy individuals, and if it is less than that value, it is schizophrenia / depression Can be determined.
  • a protein or the like selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 124 to 126 is used as a supplementary mental disease determination marker that can distinguish whether a subject suffers from schizophrenia or depression. May be.
  • Such supplemental markers are shown in Table 8.
  • the markers shown in Table 8 cannot be used alone to distinguish between patients with schizophrenia or depression because the difference between healthy individuals and each disease is small. However, if the patient suffers from either schizophrenia or depression, both can be differentiated.
  • the concentration 4320RFU of the Triosephosphate isomerase protein consisting of the amino acid sequence represented by SEQ ID NO: 124 is used as a cut-off value, and when it is higher than that value, schizophrenia is low Diagnosis of depression. Therefore, it is useful as a marker for auxiliary determination for patients suffering from schizophrenia or depression, which is a psychiatric disorder determination marker belonging to this group.
  • Table 9 shows the markers for determining mental disorders belonging to group 6.
  • the mental disease determination marker belonging to this group is a marker that can determine depression / bipolar disorder.
  • “Depression / bipolar disorder” is a mental illness that has been classified as any mental illness by the conventional classification method, and is depressed by the determination method of the present invention based on a biological basis using a biomarker.
  • a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 108 to 122 in the psychiatric disease determination marker belonging to this group can be identified as having a high possibility of suffering from depression / bipolar disorder, It is a determination marker that can subdivide depression / bipolar disorder into subtypes.
  • the mental disease determination marker belonging to this group may be classified based on the cut-off value set in the same manner as the subtype identification of the mental disease in the first to third groups.
  • the subject in the determination of a mental illness using the LYN B marker represented by SEQ ID NO: 108, if the measured value of the LYN B marker in the subject is greater than the 95th percentile of the measured values of the healthy subject group, the subject The examiner is likely to suffer from depression / bipolar disorder and can be identified as “LynB subtype of depression / bipolar disorder”.
  • a supplementary psychiatric disease determination marker capable of distinguishing whether a subject suffers from depression or bipolar disorder, it is selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 131 to 136, 247, and 248. Or other proteins may be used.
  • the markers shown in Table 10 cannot be used alone to distinguish between a depressed patient or a bipolar disorder patient because the difference between a healthy person and each disease is small. However, if the patient is suffering from mental illness, either depression or bipolar disorder, both can be differentiated. For example, when it is desired to discriminate between depression and bipolar disorder, the protein comprising the amino acid sequence represented by SEQ ID NOs: 131, 132, 133, 136, 247, or 248 is depressed when the concentration is higher than the cut-off value, and when it is low If the concentration of the protein consisting of the amino acid sequence shown by SEQ ID NO: 134 or 135 is lower than the cut-off value, it can be diagnosed as bipolar disorder. Therefore, it is useful as a marker for auxiliary determination for patients suffering from depression or bipolar disorder as a mental disease determination marker belonging to this group.
  • Group for determining schizophrenia, depression or bipolar disorder Table 11 shows the markers for determining mental disorders belonging to Group 7.
  • the mental disease determination marker belonging to this group is a marker that can determine a mental disease. Specifically, it refers to a protein or the like belonging to the compartments shown in Group 7 in FIG. 1 and selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 13, 34, and 137-140.
  • the subject is either schizophrenia, depression, bipolar disorder, schizophrenia / bipolar disorder, schizophrenia / depression, or depression / bipolar disorder. It can be determined that there is a high possibility of having a mental illness.
  • the 2nd aspect of this invention is a mental disease determination method.
  • the present invention can biologically determine the presence or absence of a mental illness in a subject without using a doctor using the mental illness determination marker described in the first aspect.
  • Step 2-2-1 Determination method 2-2-1. Step This determination method includes a measurement step and a comparison determination step as essential steps. Moreover, a severity determination process is included as a selection process. Hereinafter, each step will be specifically described.
  • Measurement process is a method for measuring the amount of the psychiatric disorder determination marker according to the first aspect included in a unit amount of a sample collected from a subject and a healthy person. This is a step of obtaining a measured value.
  • healthy person refers to a human individual who does not have a mental illness, preferably a healthy human individual who does not have any disease or abnormality.
  • the healthy person used in this embodiment is not particularly limited in terms of physical conditions such as sex, age, height, weight, etc., but the number is preferably the same as that of the subject, and physical conditions such as age, height, and weight. Are the same or approximate.
  • a group of a plurality of healthy persons is referred to as a “healthy person group”.
  • sample refers to a biological sample that may contain a protein or fragment thereof that is a psychiatric disorder determination marker.
  • a bodily fluid, a cell (including a cell extract), or a protein recovered therefrom is applicable.
  • Body fluid refers to a liquid biological sample collected directly from a subject. For example, cerebrospinal fluid, blood (including serum, plasma and interstitial fluid), or lymph. In the method of this embodiment, a preferable sample is cerebrospinal fluid or a protein recovered therefrom.
  • the sample collected refers to a sample collected from each of the subject and the healthy subject.
  • the collecting method is not particularly limited as long as it is a known method.
  • the sample when the sample is cerebrospinal fluid, it may be collected by lumbar puncture. Lumbar puncture is less invasive because it can reduce pain or less by using a commercially available local anesthetic in advance, and can reduce side effects by using an atraumatic needle, This method is suitable for collecting cerebrospinal fluid.
  • the collection may be performed based on a known method in the field. For example, in the case of blood or lymph, a known blood collection method may be followed. Specifically, in the case of peripheral blood, it may be collected by injection into a peripheral vein or the like.
  • the sample can be used immediately after collection in the determination method of the present invention, but immediately after collection, it can be ice-cooled, and the supernatant obtained by centrifugation can be thawed and used. Good. Further, if necessary, dilution or concentration, or a blood coagulation inhibitor such as heparin can be added.
  • the “unit amount” is a predetermined unit of volume or weight, and examples thereof include microliter, milliliter, microgram, milligram, gram and the like.
  • the “measured value” is a value indicating the amount of the psychiatric disorder determination marker measured in this step.
  • the measured value may be an absolute value such as volume or weight, or a relative value such as concentration, ionic strength, absorbance or fluorescence intensity.
  • the amount of the psychiatric disease determination marker contained in each of the sample derived from the subject (referred to as “subject sample”) and the sample derived from the healthy subject sample (referred to as “normal subject sample”) is measured.
  • the mental disease determination marker to be measured by this method may be selected according to the mental disease to be determined. For example, when it is determined whether or not a subject is not in a normal state and suffers from any mental illness, it is represented by SEQ ID NOS: 13, 34, and 137 to 140 belonging to Group 7 described in the first aspect. Any one or more of the psychiatric disease determination markers may be selected. Further, when determining whether or not the subject suffers from schizophrenia / bipolar disorder, any one of the mental disease determination markers represented by SEQ ID NOs: 94 to 98 belonging to Group 4 according to the first aspect. One or more may be selected.
  • any one or more of the psychiatric disorder determination markers represented by 233 to 240 may be selected.
  • any one of the mental disease determination markers represented by SEQ ID NOs: 104 to 122 and 241 belonging to group 6 described in the first aspect One or more may be selected.
  • Any one or more of psychiatric disorder determination markers indicated by ⁇ 178 and 180 ⁇ 232 may be selected.
  • the method for measuring a psychiatric disorder determination marker is not particularly limited as long as it is a known peptide quantification method.
  • immunological detection methods, aptamer analysis methods, or mass spectrometry methods can be mentioned.
  • the immunological detection method is a method for quantifying a psychiatric disease determination marker that is a target molecule of the present method, using an antibody or a fragment thereof that specifically binds to the target molecule.
  • the antibody used in the immunological detection method may be any of a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a synthetic antibody, and an antibody fragment.
  • the immunoglobulin molecule can be of any class (eg, IgG, IgE, IgM, IgA, IgD and IgY), or any subclass (eg, IgG1, IgG2, IgG3, IgG4, IgA1). IgA2).
  • the antibody can be derived from any animal including mammals and birds. Examples include mice, rats, guinea pigs, rabbits, goats, donkeys, sheep, camels, horses, chickens or humans.
  • Recombinant antibodies include chimeric antibodies, humanized antibodies, or multispecific antibodies.
  • a chimeric antibody is an antibody prepared by combining amino acid sequences of antibodies from different animals, and is an antibody in which the variable region (V region) of one antibody is replaced with the V region of another antibody.
  • V region variable region
  • the V region of a mouse monoclonal antibody that specifically binds to any of the psychiatric disease determination markers described in the first embodiment is replaced with the V region of a human antibody, the variable region (V region) is derived from a mouse, and the C region is human.
  • Applicable antibodies are derived.
  • a humanized antibody is a complementarity determining region (CDR; CDR1) in a variable region (V region) of a non-human mammal, for example, a mouse antibody that specifically binds to any of the psychiatric disease determination markers described in the first aspect.
  • CDR2 and CDR3 are graft antibodies in which the CDR of the human monoclonal antibody is substituted.
  • Multispecific antibody refers to a multivalent antibody, that is, an antibody that binds to different epitopes in each antigen-binding site in an antibody having a plurality of antigen-binding sites in one molecule. For example, in the case of an antibody having two antigen-binding sites such as IgG, each of the antigen-binding sites specifically binds to the same or different psychiatric disease determination marker described in the first embodiment ( Bispecific antibody).
  • Synthetic antibody refers to an antibody synthesized by using a chemical method or a recombinant DNA method.
  • a monomeric polypeptide molecule in which one or more VL and one or more VH of a specific antibody are artificially linked through a linker peptide having an appropriate length and sequence, or a multimeric polypeptide thereof Applicable.
  • Specific examples of such polypeptides include single chain Fv (scFv: single chain Fragment of variable region) (Pierce Catalog and Handbook, 1994-1995, Pierce Chemical Co., Rockford, IL), diabody ), Triabody or tetrabody.
  • VL and VH are usually located on separate polypeptide chains (light chain and heavy chain).
  • Single-chain Fv is a synthetic antibody fragment having a structure encompassed by one polypeptide chain, in which V regions on these two polypeptide chains are connected by a flexible linker having a sufficient length. Within a single chain Fv, both V regions can self-assemble with each other to form one functional antigen binding site. Single-stranded Fv can be obtained by integrating and expressing a recombinant DNA encoding it in the phage genome using known techniques.
  • a diabody is a molecule having a structure based on a dimeric structure of a single chain Fv (Holliger et al., 1993, Proc. Natl. Acad. Sci. USA 90: 6444-6448).
  • the two variable sites within the single chain Fv cannot self-assemble, but by forming a diabody, i.e., two single chain Fv ,
  • the VL of one Fv chain can be assembled with the VH of the other Fv chain to form two functional antigen binding sites (Marvin et al., 2005, Acta Pharmacol. Sin. 26: 649-658).
  • a cysteine residue to the C-terminus of the single chain Fv, it becomes possible to form a disulfide bond between the two Fv chains and to form a stable diabody (Olafsen et al., 2004). , Prot. Engr. Des. Sel.
  • a diabody is a bivalent antibody fragment, but each antigen-binding site need not bind to the same epitope, and each has a bispecificity that recognizes and specifically binds to a different epitope. You may do it.
  • Triabodies and tetrabodies have their trimer and tetramer structures based on a single-chain Fv structure as in diabodies. These are trivalent and tetravalent antibody fragments, respectively, and may be multispecific antibodies.
  • Antibody fragments include, for example, Fab, F (ab ′ 2 ), Fv and the like.
  • immunological detection methods include enzyme immunoassay (including ELISA and EIA methods), fluorescence immunoassay, radioimmunoassay (RIA), luminescence immunoassay, surface plasmon resonance (SPR method) , Quartz crystal microbalance (QCM) method, immunoturbidimetric method, latex agglutination immunoassay method, latex turbidimetric method, erythrocyte agglutination reaction, particle agglutination reaction method, colloidal gold method, capillary electrophoresis, Western blotting or immunization Histochemical method (immunostaining method) can be mentioned.
  • the aptamer analysis method is a method for quantifying a psychiatric disorder determination marker, which is a target molecule of the present method, using an aptamer that binds firmly and specifically to a target molecule.
  • Aptamers are ligand molecules that have the ability to bind firmly and specifically to a target substance through a three-dimensional structure and specifically inhibit the function of the target substance. Aptamers can be roughly classified into nucleic acid aptamers and peptide aptamers depending on the type of the molecule, and any aptamer may be used.
  • Nucleic acid aptamer refers to an aptamer composed of nucleic acids.
  • the nucleic acid constituting the nucleic acid aptamer may be any of DNA, RNA, or a combination thereof. If necessary, a chemically modified nucleic acid such as PNA, LNA / BNA, methylphosphonate DNA, phosphorothioate DNA, 2′-O-methyl RNA may be included.
  • the nucleic acid aptamer can be prepared by a method known in the art using any of the psychiatric disorder determination markers described in the first embodiment as a target molecule.
  • an RNA aptamer can be prepared by in vitro selection using the SELEX (systematic “evolution” of “ligands” by “exponential” enrichment) method.
  • the SELEX method selects an RNA molecule bound to a psychiatric disorder marker as a target molecule from an RNA pool composed of a large number of RNA molecules having a random sequence region and primer binding regions at both ends.
  • RNA having a strong binding force is selected RNA having a strong binding force.
  • the base sequence lengths of the random sequence region and the primer binding region are not particularly limited. Generally, the random sequence region is 20 to 80 bases, and the primer binding region is 15 to 40 bases.
  • the RNA molecule finally obtained by the above method is used as a mental disease determination marker-binding RNA aptamer.
  • the SELEX method is a known method, and a specific method may be performed according to, for example, Pan et al. (Proc. Natl. Acad. Sci. 1995, U.S.A.92: 11509-11513).
  • a peptide aptamer is an aptamer composed of amino acids, and is a 1-6 kD peptide molecule that recognizes the surface structure of a specific target molecule and binds specifically like an antibody. It can be produced using a phage display method or a cell surface display method. What is necessary is just to produce the manufacturing method of a peptide aptamer based on a well-known method in the said field
  • the antibody or aptamer may be labeled as necessary.
  • a labeling substance known in the art may be used.
  • fluorescent dyes fluorescein, FITC, rhodamine, Texas red, Cy3, Cy5
  • fluorescent proteins eg, PE, APC, GFP
  • enzymes eg, horseradish peroxidase, alkaline phosphatase, glucose) Oxidase
  • radioisotopes eg 3 H, 14 C, 35 S
  • biotin or (strept) avidin eg 3 H, 14 C, 35 S
  • nucleic acid aptamers for example, radioisotopes (eg, 32 P, 3 H, 14 C), DIG, biotin, fluorescent dyes (eg, FITC, Texas, cy3, cy5, cy7, FAM, HEX, VIC, JOE, Rox, TET, Bodipy493, NBD, TAMRA), or a luminescent substance (for example, acridinium ester).
  • An antibody or aptamer labeled with a labeling substance can be a useful tool for detecting an aptamer bound to a target protein.
  • Mass spectrometry includes high performance liquid chromatograph mass spectrometry (LC-MS), high performance liquid chromatograph tandem mass spectrometry (LC-MS / MS), gas chromatograph mass spectrometry (GC-MS), and gas chromatograph tandem mass. Analytical methods (GC-MS / MS), capillary electrophoresis mass spectrometry (CE-MS) and ICP mass spectrometry (ICP-MS) can be mentioned.
  • the measurement value of the healthy person in the measurement step may be determined by measuring the amount of each psychiatric disease determination marker in a sample obtained from a healthy person or a group of healthy persons in advance, and using it as a database. Good.
  • a known protein expected to have no quantitative difference in the sample per unit amount may be further measured as an endogenous control.
  • endogenous control protein include albumin.
  • Comparison determination process is a process of determining the morbidity of a mental illness in a subject based on the measurement value obtained in the measurement process.
  • the comparison of the measured values is a cut-off obtained between the measured values of the subject obtained in the measuring step and the measured values of the healthy person or the healthy person group, or based on the measured values of the subject and the healthy person group. Do by value. At this time, the measured values of the subject and healthy subjects can be corrected using the measured values of the endogenous control protein.
  • Judgment is made when the measured value of the subject is significantly higher or lower than the measured value of the healthy subject or the group of healthy subjects, or when the subject is classified as positive by the cut-off value, the subject is determined to have the mental disease selected in the measuring step. It is determined that the marker is likely to have a psychiatric disorder that can be determined. For example, when any one of SEQ ID NOs: 1 to 6 belonging to one group described in the first aspect is selected as a mental illness determination marker in the measurement step, the measurement value of the subject is significantly higher than the measurement value of the healthy subject When it is high, it can be determined that the subject is likely to have schizophrenia.
  • the measurement value is determined in advance based on the measurement value of the healthy subject group. If the subject is classified as positive according to the determined cut-off value, it can be determined that the subject is likely to have schizophrenia. On the other hand, if the measured value is not significantly high, or if the measured value is classified as negative by the cut-off value, it is determined that the subject is likely not suffering from schizophrenia.
  • Judgment is made by the psychiatric disease determination marker selected and used in the measurement process. For example, when using the psychiatric disease determination marker belonging to Group 7 described in the first aspect, when the measurement value of the subject is classified as positive by a predetermined cutoff value based on the measurement value of the healthy group, It is determined that the subject is not in a healthy state and is likely to have any mental illness.
  • the PSA-ACT protein represented by SEQ ID NO: 94 belonging to Group 4 described in the first embodiment is used as a psychiatric disease determination marker, if the measured value of the subject is significantly higher than the measured value of the healthy subject, the subject is Determine that they are likely to have schizophrenia / bipolar disorder.
  • the psychiatric disease determination marker represented by SEQ ID NOs: 95 to 98 belonging to Group 4 when the subject is positively classified by a predetermined cutoff value based on the measurement value of the healthy group, the subject is Determine that they are likely to have schizophrenia / bipolar disorder.
  • the mental disease determination marker represented by SEQ ID NOs: 99 and 240 belonging to Group 5 described in the first aspect if the measurement value of the subject is significantly higher than the measurement value of the healthy subject, the subject is schizophrenia. Determined to have a high probability of suffering from symptom / depression.
  • the psychiatric disease determination markers represented by SEQ ID NOs: 2, 4, 7, 100 to 103, 145, 146, 179 and 233 to 239 belonging to Group 5 are used, the markers are determined in advance based on the measured values of the healthy group. If the subject is classified as positive by the cut-off value, it is determined that the subject is likely to have schizophrenia / depression.
  • the mental disease determination marker represented by SEQ ID NOs: 104 to 107 and 241 belonging to Group 6 described in the first aspect if the measurement value of the subject is significantly higher than the measurement value of the healthy subject, the subject is Determine that the person is most likely suffering from depression / bipolar disorder.
  • the psychiatric disease determination marker represented by SEQ ID NOs: 108 to 122 belonging to Group 6 if the subject is positively classified according to a predetermined cutoff value based on the measurement value of the healthy group, the subject is Determine that the person is most likely suffering from depression / bipolar disorder.
  • the measured value of the subject is measured by a healthy person as described above. If significantly higher than the value, the subject is determined to be more likely to have schizophrenia.
  • the psychiatric disease determination markers represented by SEQ ID NOs: 7 to 25, 141 to 148, and 249 belonging to one group are used, they are classified as positive according to a cut-off value determined in advance based on the measurement values of the healthy group. If so, the subject is determined to be likely to have schizophrenia.
  • the mental disease determination marker represented by SEQ ID NOs: 61, 62, and 170 to 172 belonging to the two groups described in the first aspect if the measured value of the subject is significantly higher than the measured value of a healthy person, The subject is determined to be more likely to have bipolar disorder.
  • the mental disease determination markers represented by SEQ ID NOs: 63, 64, 114, and 163 to 169 belonging to the two groups are used, the markers are positively classified according to a predetermined cutoff value based on the measurement values of the healthy group. If so, the subject is determined to be likely to have bipolar disorder.
  • the measured value of the subject is healthy. If it is significantly higher than the person's measurement, the subject is determined to be more likely to have depression. Further, when the mental disease determination markers represented by SEQ ID NOs: 66, 76 to 87, 174 to 178, and 180 to 192 belonging to the three groups are used, a cutoff value determined in advance based on the measured values of the healthy group If the subject is classified as positive, the subject is determined to have a high probability of suffering from depression.
  • Severity determination step is a step selected when identifying the severity of a mental illness.
  • the severity determination step is a supplementary step of further identifying the severity of the mental illness on the assumption that the subject suffers from schizophrenia, bipolar disorder, or depression. This process is performed after the comparison determination process in principle, but can be performed simultaneously with the comparison determination process.
  • a mental disease determination marker that can identify the severity of a mental illness indicates that the measured value in the sample of the marker correlates with a predetermined evaluation scale of each mental illness. Therefore, a calibration curve indicating the relationship between the measured value of each severity identification marker and the evaluation scale may be created in advance. From the measured value of the severity identification marker in the subject and the calibration curve, the severity of the mental illness in the subject can be easily identified. For example, when the measured value of the mental illness determination marker and the value of the evaluation scale are in a positive proportional relationship, it can be determined that the severity of the mental illness is high if the measured value is high.
  • the severity of schizophrenia is identified in this step, it is selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 9, 13, 26 to 29, 31 to 60, and 159 to 162 as a psychiatric disorder determination marker.
  • Use protein In the case of a psychiatric disorder determination marker consisting of the amino acid sequences represented by SEQ ID NOs: 26 to 33, the measured value of the marker correlates with PANSS. Further, in the case of a psychiatric disease determination marker selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 9, 13, 34 to 60, and 159 to 162, the measured value of the marker correlates with BACS.
  • a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 47, 65 to 69, and 173 is used.
  • the measured values of these psychiatric disease determination markers are correlated with HAM-D.
  • a protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 44, 66, and 88 to 93 is used.
  • the measured values of these psychiatric disease determination markers are correlated with HAM-D.
  • any one of the mental disease determination markers described in the first aspect may be used in the measurement step, but two or more may be combined and measured.
  • the accuracy of determining mental illness can be further enhanced.
  • the case where the mental disease determination markers belonging to the first group, the second group, the fourth group, and the seventh group described in the first aspect are combined may be mentioned.
  • Based on the results of 7 groups of mental illness determination markers it was determined that the patient is not in a healthy state and is likely to suffer from psychiatric disorders such as schizophrenia, bipolar disorder, or depression. From the results of the disease determination marker, it can be determined that the subject is more likely to suffer from schizophrenia or bipolar disorder than depression. Subsequently, from the results of the group 1 and group 2 mental illness determination markers, it can be determined that the subject is likely to suffer from schizophrenia or bipolar disorder, respectively.
  • the case where the mental disease determination markers belonging to the first group, the third group, the fifth group, and the seventh group described in the first aspect are combined is exemplified. From the results of the 7 groups of mental illness determination markers, it was determined that the patient is not in a healthy state and is likely to suffer from psychiatric disorders such as schizophrenia, bipolar disorder, or depression. From the result of the disease determination marker, it can be determined that the subject is likely to have schizophrenia or depression. Subsequently, from the results of the group 1 and group 3 mental disease determination markers, it can be determined that the subject has a high possibility of suffering from schizophrenia or depression, respectively.
  • the mental disease determination markers belonging to the second group, the third group, the sixth group and the seventh group described in the first aspect are combined. Based on the results of the 7 groups of mental illness determination markers, it is determined that the patient is not in a healthy state and is likely to suffer from psychiatric disorder such as schizophrenia, bipolar disorder, or depression. From the result of the disease determination marker, it can be determined that the subject is highly likely to suffer from depression or bipolar disorder. Subsequently, it can be determined from the results of the mental disorder determination markers in groups 2 and 3 that the subject is highly likely to suffer from bipolar disorder or depression, respectively.
  • the combination of psychiatric disorder determination markers may be from the same group.
  • Integrin1 ⁇ 1 of SEQ ID NO: 27 and Integrin ⁇ 1 of SEQ ID NO: 28 are selected from one group. Since these form heterodimers with each other in vivo, the measurement accuracy can be increased by measuring the respective amounts in combination.
  • Sample collection and sample collection The sample provider for selecting the psychiatric disorder determination marker of the present invention can be obtained from the National Psychiatric and Neurological Research Center (Tokyo, Japan) website, free paper advertisement, or in the center hospital. recruited with recruitment posters posted on. As a result, more than 300 donors were obtained.
  • Table 12 shows sample cohort information for 30 schizophrenia patients, 30 depression patients, 16 bipolar disorder patients, and 30 healthy individuals as controls.
  • N indicates the total number of each mental illness and healthy person
  • DF indicates the number of patients not taking the drug (drug free).
  • the method of collecting cerebrospinal fluid from the subject was performed according to the conventional method.
  • neurological examinations including fundus examination, local anesthesia is performed between the 3rd and 4th lumbar vertebrae and the 4th and 5th lumbar vertebrae, and lumbar puncture is performed with an atraumatic needle (22G, Unisys, UNIEVER puncture needle)
  • atraumatic needle 22G, Unisys, UNIEVER puncture needle
  • about 10 mL of cerebrospinal fluid was collected.
  • the first approximately 2mL is used for general specimen testing (cell count, total protein, sugar, etc.), and the next approximately 8mL is used for isolating a psychiatric disorder marker (Sumitomo Bakelite, Proteo Save). 15 mL).
  • the cerebrospinal fluid was immediately ice-cooled and centrifuged at 4000 g for 15 minutes at 4 ° C. Thereafter, 0.5 mL of the supernatant was dispensed into a low protein adsorption tube and stored in a -80 degree ultra low temperature tank until use.
  • FIG. 2 is a diagram showing a measured value distribution of one of the mental disease determination markers SCFsR represented by SEQ ID NO: 70 belonging to Group 3.
  • DF drug-free
  • proteins belonging to Group 3 can serve as a psychiatric disease determination marker for determining depression. Similar results were observed for the psychiatric disorder determination marker for schizophrenia belonging to Group 1 and the mental illness determination marker for bipolar disorder belonging to Group 2.
  • FIG. 3 is a diagram showing a measured value distribution of UNC5H3 that can subdivide schizophrenia into subtypes among the psychiatric psychiatric disorder determination markers indicated by SEQ ID NO: 7 belonging to Group 1.
  • the 95th percentile in the healthy group was defined as the cut-off value, and the 95th percentile or greater was defined as positive.
  • Six of 30 patients with schizophrenia who showed an abnormal value of 95th percentile or higher can be subdivided as UNC5H3 subtype of schizophrenia.
  • 4 out of 6 cases were drug-free specimens, and it was shown that the possibility of the influence of the therapeutic agent is low. Similar results were observed for the mental disorder determination marker for bipolar disorder belonging to Group 2 and the mental disorder determination marker for depression belonging to Group 3.
  • FIG. 4 is a diagram showing the relationship between the measured value of schizophrenia by ST4S6 which is a psychiatric psychiatric disorder determination marker indicated by SEQ ID NO: 29 belonging to group 1 and the evaluation value by the evaluation scale PANSS.
  • ST4S6 concentration As shown in this figure, there is a correlation between ST4S6 concentration and PANSS, which is a symptom evaluation scale, and the higher the ST4S6 concentration, the more severe the symptoms. Therefore, the presence and severity of schizophrenia can be determined by using ST4S6 as a psychiatric disorder determination marker.
  • Mental illness determination markers having the same characteristics as above were confirmed for some of the mental illness determination markers for bipolar disorder belonging to Group 2 and some of the mental illness determination markers for depression belonging to Group 3.
  • FIG. 5 is a diagram showing a distribution of measured values of LYN B, one of the psychiatric disorder determination markers indicated by SEQ ID NO: 108 belonging to Group 6.
  • LYN B was significantly increased compared with the healthy group (ANCOVA considering gender and age, p ⁇ 0.01).
  • proteins belonging to Group 6 can be a psychiatric disease determination marker for determining depression / bipolar disorder. Similar results were observed for the psychiatric disease determination marker for schizophrenia / bipolar disorder belonging to Group 4 and the psychiatric disease determination marker for schizophrenia / depression belonging to Group 5.
  • FIG. 6 is a diagram in which cut-off values are entered in the measured value distribution diagram of LYN B shown in FIG.
  • the cutoff value was the 95th percentile in the healthy group.
  • LYN B was more than the cut-off value in 7 of 30 patients with depression and 6 of 16 patients with bipolar disorder.
  • Some psychiatric disease markers belonging to group 6 such as LYN B can be used as psychiatric disease markers for determining depression / bipolar disorder subtypes by setting a cut-off value. It was shown that depression / bipolar disorder can be subdivided by using a psychiatric disorder determination marker. Similar results were also found for schizophrenia / bipolar disorder psychiatric disease markers belonging to group 4 and schizophrenia / depressive psychiatric disease markers belonging to group 5.
  • FIG. 7 is a diagram showing a distribution of measured values of a psychiatric disorder determination marker PTH for determining whether the patient suffers from schizophrenia or depression.
  • cerebrospinal fluid PTH was significantly decreased in the schizophrenic patient group compared to the depression patient group (p ⁇ 0.002 in ANCOVA considering gender and age). Even if the schizophrenia group was limited to drug-free cases, it was significantly reduced compared to the depression group (p ⁇ 0.05).
  • this marker can be an auxiliary psychiatric disease determination marker that can appropriately determine whether it is schizophrenia or depression for a subject determined to be schizophrenia or depression, for example.
  • FIG. 8 is a diagram showing the measured value distribution of VEGF sR3, which is a psychiatric disorder determination marker belonging to Group 7.
  • the cutoff value was the 95th percentile in the healthy group. 10 out of 30 cases of schizophrenia, 8 out of 30 cases of depression, and 3 out of 16 cases of bipolar disorder were positive (above cut-off value). If this marker is used, it can be judged whether a test subject is in a healthy state or a mental illness.
  • Example 1 ⁇ Verification of judgment system by mental disease judgment marker>
  • Each psychiatric disease determination marker obtained in Example 1 and Example 2 was verified to be reproducible even when psychiatric disease determination was performed for different sample cohorts.
  • each of the first sample cohort shown in Table 12 and the second sample cohort shown in Table 13 is the same as in Example 1.
  • the analysis was independently performed by the method described above to verify the measurement distribution. Furthermore, the measurement result of the first sample cohort and the measurement result of the second sample cohort were combined by Z conversion, and the distribution was verified.
  • FIG. 9 verifies Fibrinogen, a mental illness determination marker for depression selected from the first sample cohort.
  • A is a measurement distribution of fibrinogen values in the cerebrospinal fluid of each psychiatric patient in the second specimen cohort.
  • fibrinogen levels were significantly increased in depressed patients (MDD), confirming the reproducibility of fibrinogen as a psychiatric disease marker for depression.
  • MDD depressed patients
  • C is the result of combining the measurement results of the first sample cohort and the measurement results of the second sample cohort by Z conversion. In this figure, fibrinogen was also markedly elevated in depressed patients (MDD). Therefore, depression subtypes can be determined by measuring fibrinogen in cerebrospinal fluid.
  • FIG. 10 is a diagram showing a relationship between a measured value of a schizophrenia sample by PIGF which is a psychiatric disorder determination marker for schizophrenia and an evaluation value of a sign task of a schizophrenia cognitive function simple evaluation scale (BACS). .
  • PIGF a psychiatric disorder determination marker for schizophrenia
  • AVS schizophrenia cognitive function simple evaluation scale
  • FIGS. 11 to 48 are measurement distribution diagrams of each psychiatric disorder determination marker, where (a) shows the measurement results in the first sample cohort, (b) shows the measurement results in the second sample cohort, and (c) shows (a) and ( The result of combining the measurement results of b) by Z conversion is shown.
  • the name of the psychiatric disorder determination marker in each figure was described in the above “Simple Description of Drawing”.
  • a significant difference was observed in the patient group as the target disease of the marker, and the reproducibility of detection accuracy as a psychiatric disease determination marker could be confirmed. From this result, it was shown that the mental illness determination marker of the present invention is an effective mental illness marker for assisting the determination of mental illness for any specimen.

Abstract

The purposes of the present invention is to develop and provide a mental illness determination marker that can easily and highly accurately determine the presence/absence of schizophrenia, depression or bipolar disease in subjects. Provided is a mental illness determination marker comprising: proteins made of a group consisting of amino acid sequences represented by sequence numbers 1-249 and included in cerebrospinal fluid; and a variant of the proteins or a segment thereof.

Description

精神疾患判定マーカーPsychiatric disorder marker
　本発明は、被験者由来の試料を用いて精神疾患である統合失調症、双極性障害又はうつ病の有無を判定するためのバイオマーカー及びそれを用いた精神疾患判定方法に関する。 The present invention relates to a biomarker for determining the presence / absence of psychiatric schizophrenia, bipolar disorder or depression using a sample derived from a subject, and a mental disease determination method using the biomarker.
　現代社会において、統合失調症、うつ病、及び双極性障害等の精神疾患（Mental Disorder)は、大きな社会問題となっている。例えば、日本の平成23年（2011年）厚生労働省患者調査によると、日本国内の前記精神疾患の入院患者数は、全疾患を合計した国内総入院患者数の約23%にものぼる。通院患者数や潜在的患者数を加えると、精神疾患患者数は、国内で膨大な数に達すると予想される。これらの精神疾患は生産年齢層に多く見られることから、医療費の負担増のみならず、経済的損失や自殺による社会的損失等、社会に与える影響は計り知れない。 In modern society, mental disorders such as schizophrenia, depression, and bipolar disorder (Mental Disorder) have become a major social problem. For example, according to the 2011 survey by the Ministry of Health, Labor and Welfare in Japan, the number of inpatients with mental illness in Japan is about 23% of the total number of inpatients in Japan. When the number of outpatients and the number of potential patients are added, the number of patients with mental illness is expected to reach an enormous number in Japan. Since these mental illnesses are common in the working age group, not only the cost of medical expenses is increased, but also the social impact such as economic loss and social loss due to suicide is immeasurable.
　一般に、精神疾患は、疾患間で近似する症状が多く、また各疾患において症状が多岐にわたることから各疾患の定義が明確でない。そのため専門家であっても正確な診断は、しばしば困難を極める。 In general, mental diseases have many symptoms that are close to each other, and each disease has a wide variety of symptoms, so the definition of each disease is not clear. For this reason, accurate diagnosis is often extremely difficult even for specialists.
　現在、精神疾患の診断は本人や家族等への問診で聴取した症状、兆候及び経過等の情報に基づいて行われている。通常、米国精神医学会により規定されたDiagnostic and Statistical Manual of Mental Disorders(DSM)が精神疾患の診断基準に使用されている。ところが、この診断基準は、客観的な検査を用いるわけではないため、診断する医師によって一致しないことも多い。米国の大規模調査によると、一般診療でDSM診断に基づいて、うつ病と診断された5639人のうち、構造化診断面接でうつ病性障害と再診断された患者は38.4%に過ぎないことが報告されている（非特許文献１）。構造化診断面接とは、マニュアル化された面接法で、質問内容や再質問等の形式を予め設定することで、面接者による診断の違いを小さくすることができる。構造化診断面接を用いれば、比較的高い一致度は得られる（非特許文献２）が、構造化診断面接には診断者の高い熟練度と1～2時間に及ぶ面接時間が必要な上、後述するように、一致度の高い診断を行うことによる診療上の有益性がそれらに見合わないため、研究以外で行われることはほとんどない。 At present, diagnosis of mental illness is performed based on information such as symptoms, signs, and progress obtained through interviews with the person and family. In general, Diagnostic and Statistical Manual of Mental Disorders (DSM) defined by the American Psychiatric Association are used as diagnostic criteria for mental disorders. However, since this diagnostic criterion does not use an objective test, it often does not agree with the doctor who makes the diagnosis. According to a large U.S. survey, only 38.4% of 5639 people diagnosed with depression based on DSM diagnosis in general practice were rediagnosed as depressive disorder in a structured diagnostic interview Has been reported (Non-patent Document 1). The structured diagnosis interview is a manual interview method in which the format of the question content, the re-question, etc. is set in advance, and the difference in diagnosis by the interviewer can be reduced. If a structured diagnostic interview is used, a relatively high degree of coincidence can be obtained (Non-Patent Document 2), but the structured diagnostic interview requires a high degree of skill of the diagnostician and an interview time of 1 to 2 hours. As will be described later, since the medical benefits of making a diagnosis with a high degree of coincidence are not commensurate with them, it is rarely done outside of research.
　また、DSM診断は障害に対して判定されるため、生物学的な病態を必ずしも反映しているとは言えない。実際、脳以外に原因があることが明確な病態であっても、DSM診断では誤って精神疾患と診断されるケースがしばしばみられる。例えば、うつ病と診断された患者の少なくとも1割以上は、甲状腺機能低下症や抗炎症薬の副作用と言われている（非特許文献３）。また、統合失調症と診断された患者の中に、抗NMDA受容体抗体脳炎が含まれることが報告されている（非特許文献４）。 Also, since DSM diagnosis is determined for a disorder, it cannot be said that it necessarily reflects biological pathology. In fact, even if the pathology is clearly related to a cause other than the brain, there are often cases in which a DSM diagnosis is mistakenly diagnosed as a mental illness. For example, at least 10% of patients diagnosed with depression are said to have side effects of hypothyroidism and anti-inflammatory drugs (Non-patent Document 3). In addition, it has been reported that patients diagnosed with schizophrenia include anti-NMDA receptor antibody encephalitis (Non-patent Document 4).
　ところで、現行の抗うつ薬は、一部の重症症例を除いて十分な効果が認められていない（非特許文献５）。また、統合失調症治療薬の効果も幻聴・妄想等の一部の症状に対する効果に留まっている。このように、精神疾患に対する治療薬の効果が限定される原因の一つとして、精神疾患の分類が十分に確立していない点が挙げられる。例えば、うつ病と診断された精神疾患にも細分類が可能な複数の病態が存在し、現在処方されている抗うつ薬、うつ病における一部の病態にしか治療効果がない、というものである。したがって、精神疾患の本質的な治療法の開発には、生物学的な根拠に基づく新たな分類とそれに応じた適切な診断技術の開発が求められている。 By the way, the current antidepressant has not been sufficiently effective except in some severe cases (Non-patent Document 5). In addition, the effects of schizophrenia drugs are limited to some symptoms such as hallucinations and delusions. Thus, one of the reasons for the limited effect of therapeutic agents for mental illness is that the classification of mental illness is not well established. For example, there are multiple pathologies that can be subdivided into mental illnesses diagnosed with depression, and only some of the currently prescribed antidepressants and depression have therapeutic effects. is there. Therefore, the development of an essential therapy for mental illness requires the development of a new classification based on biological evidence and an appropriate diagnostic technique.
　上記背景から、生物学的根拠に基づく新たな診断方法として、近年バイオマーカーが注目されている。バイオマーカーとは、特定の疾患の発症に応答して細胞が産生する核酸やタンパク質等の物質であって、体液や組織中における存在量又はその変動が対応する疾患の有無、重症度、又は予後を反映することから、疾患判定及び治療指針決定等の鑑別子となり得る。 From the above background, biomarkers have recently attracted attention as a new diagnostic method based on biological evidence. A biomarker is a substance such as a nucleic acid or protein produced by a cell in response to the onset of a specific disease, and the presence, severity, or prognosis of an abundance or variation in body fluids or tissues. Therefore, it can be a discriminator for disease determination and treatment guideline determination.
　精神疾患分野においても、これまで統合失調症、うつ病、又は双極性障害のバイオマーカー探索は行われており、サイトカインや神経栄養因子等、様々な候補分子が提案されてきた。ところが、その多くは十分な検出効果が得られておらず、精神疾患の実用的なバイオマーカーは、未だに開発されていないのが現状である（非特許文献６及び７）。 In the field of mental illness, biomarker searches for schizophrenia, depression, or bipolar disorder have been conducted so far, and various candidate molecules such as cytokines and neurotrophic factors have been proposed. However, most of them do not have a sufficient detection effect, and practical biomarkers for mental illness have not been developed yet (Non-Patent Documents 6 and 7).
　本発明は、被験者における統合失調症、うつ病、又は双極性障害等の精神疾患を簡便かつ高精度に判定することができる実用性の高い精神疾患判定マーカーを開発し、提供することを課題とする。また、本発明は、被験者における統合失調症、うつ病、又は双極性障害等の亜型、及び／又は重症度をさらに判定できる精神疾患判定マーカーを開発し、提供することを課題とする。 It is an object of the present invention to develop and provide a highly practical psychiatric disease determination marker capable of easily and accurately determining a psychiatric disease such as schizophrenia, depression, or bipolar disorder in a subject. To do. Another object of the present invention is to develop and provide a psychiatric disease determination marker that can further determine the subtype and / or severity of schizophrenia, depression, or bipolar disorder in a subject.
　本発明は、前記精神疾患判定マーカーを用いて被験者における各精神疾患の罹患の有無を高精度に検出する方法を開発し、提供することを課題とする。 An object of the present invention is to develop and provide a method for accurately detecting the presence or absence of each mental illness in a subject using the mental illness determination marker.
　従来の精神疾患の診断には、前述した診断基準そのものの問題や、探索に用いた試料の問題がある。診断基準の問題として、マーカー探索に際して、候補分子を患者群及び健常者群の平均値の有意差のみに基づいて選択している点が挙げられる。精神疾患は症候群であり、症候群には複数の病態が存在する可能性が高い。一部の病態については有用なバイオマーカーとなり得ても、全症例におけるその病態の割合が小さい場合には、有意差検定では埋没し、除外されてしまう可能性がある。また、試料の問題として、マーカー探索用の試料に血液や死後脳を用いていた点が挙げられる。血液は、脳をはじめ、腸、肝臓、筋肉等の様々な臓器と直接に接する。一方で、脳と他臓器の間には血液脳関門が存在し、血液中の物質の交換が制限されている。したがって、血液、特に末梢血を用いたマーカー探索では、脳に特徴的な物質の変化の検出が他臓器の影響によって困難となる可能性が高い。つまり、血液は、ノイズが大きく、脳の状態を反映しにくいという問題がある。また死後脳は、精神疾患の原因と考えられる対象臓器そのものを用いる点や脳の立体構造が保たれている点では優れているものの、死後直ちに摘出可能な動物実験と異なり、ヒトの脳は死因や死後経過による変化等の影響を受けるため、神経伝達物質やシグナル分子等の重要候補分子の変化が検出困難となる可能性が高い。さらに、これまで神経病理学や脳画像研究では、各精神疾患に特異的な病巣は見出されておらず、パーキンソン病等と異なり、脳の立体構造が保たれることの優位性は乏しい。 Conventional diagnosis of mental illnesses has the above-mentioned problems of the diagnostic criteria itself and the sample used for searching. A problem of diagnostic criteria is that, when searching for a marker, candidate molecules are selected based only on the significant difference between the average values of the patient group and the healthy group. Mental illness is a syndrome, and it is likely that there are multiple conditions in the syndrome. Even if it can be a useful biomarker for some disease states, if the proportion of the disease state in all cases is small, it may be buried in the significant difference test and excluded. Another problem with the sample is that blood or postmortem brain was used as a marker search sample. Blood directly contacts various organs such as the brain, intestines, liver, and muscles. On the other hand, a blood-brain barrier exists between the brain and other organs, and exchange of substances in the blood is restricted. Therefore, in the marker search using blood, particularly peripheral blood, it is highly likely that it is difficult to detect changes in substances characteristic of the brain due to the influence of other organs. That is, there is a problem that blood is noisy and hardly reflects the state of the brain. The postmortem brain is superior in that it uses the target organ itself, which is thought to be the cause of mental illness, and maintains the three-dimensional structure of the brain, but unlike animal experiments that can be removed immediately after death, the human brain is the cause of death. And changes due to postmortem changes are likely to make it difficult to detect changes in important candidate molecules such as neurotransmitters and signal molecules. Furthermore, in neuropathology and brain imaging research, no foci specific to each mental disorder have been found so far, and unlike Parkinson's disease, the superiority of maintaining the three-dimensional structure of the brain is poor.
　上記問題点を鑑み、本発明者らはマーカー探索に用いる試料として脳脊髄液(cerebrospinal fluid：以下しばしば「CSF」と表記する)を選択した。CSFは、脳と脊髄の周囲にのみ存在する無色透明の液体で、他の臓器とは硬膜、血液とは血液脳関門や血液脳脊髄液関門で隔てられている。CSFは、その大部分が脳の実質より滲出することや、脳細胞とCSFとの間には障壁が事実上存在しないことが近年の研究から明らかとなっている（Wang C, et al., 2012, Cerebrospinal Fluid: Physiology, biomarker and methodology. In: V. S, Dolezal, T., editor. Cerebrospinal Fluid: Functions, Composition and Disorders. New York: Nova Science Publishers; pp.1-37.）。また、診断基準の問題を解決するためにマーカーを選択する統計手法として、従来、頻用されていた健常群と疾患群との平均値の有意差に基づく方法のみならず、カットオフ値を定めて各疾患における異常値を示す症例を選択する方法、及び症例疾患群における重症度を診断する方法を新たに追加した。その結果、249種のタンパク質を精神疾患判定マーカーとして選択することができた。本発明は、上記結果に基づくものであって、以下を提供する。 In view of the above problems, the present inventors selected cerebrospinal fluid (hereinafter often referred to as “CSF”) as a sample used for marker search. CSF is a colorless and transparent fluid that exists only around the brain and spinal cord. It is separated from the other organs by the dura mater and blood by the blood-brain barrier and the blood-cerebrospinal fluid barrier. Recent studies have shown that most CSF exudes from the brain parenchyma and that there is virtually no barrier between brain cells and CSF (Wang C, et al., 2012, Cerebrospinal Fluid: Physiology, biomarker and methodology. In: V. S, Dolezal, T., editor. Cerebrospinal Fluid: Functions, Composition and Disorders. New York: Nova Science Publishers; pp. In addition, as a statistical method for selecting markers in order to solve the problem of diagnostic criteria, not only a method based on a significant difference between the average values of the healthy group and the disease group, which has been frequently used, but also a cutoff value is determined. A method for selecting a case showing an abnormal value in each disease and a method for diagnosing the severity in a case disease group were newly added. As a result, 249 proteins could be selected as markers for determining mental illness. The present invention is based on the above results and provides the following.
（１）脳脊髄液中に含まれる配列番号１～２４９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる精神疾患判定マーカー。
（２）配列番号１、３、５、６、７、８～１３、１５～３３、３５～５４、５６～６０、１４１～１４４、１４７～１６２、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症である、（１）に記載の精神疾患判定マーカー。
（３）統合失調症を亜型に細分類する、配列番号７～１３、１５～２５、１４１～１４４、１４７、１４８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、（２）に記載の精神疾患判定マーカー。
（４）統合失調症の重症度を同定する、配列番号９、１３、２６～２９、３１～３３、３５～５４、５６～６０、及び１５９～１６２で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、（２）に記載の精神疾患判定マーカー。
（５）配列番号４７、６１～６９、１１４、及び１６３～１７３で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が双極性障害である、（１）に記載の精神疾患判定マーカー。
（６）双極性障害を亜型に細分類する、配列番号６３、６４、１１４、及び１６３～１６９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、（５）に記載の精神疾患判定マーカー。
（７）双極性障害の重症度を同定する、配列番号４７、６５～６９、及び１７３で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、（５）に記載の精神疾患判定マーカー。
（８）配列番号１３、１４、１７、４４、５６、６６、７０～９３、１２３、１７４～１７８、及び１８０～２３２で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患がうつ病である、（１）に記載の精神疾患判定マーカー。
（９）うつ病を亜型に細分類する、配列番号６６、７６～８７、及び１７４～１７８、及び１８０～１９２で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、（８）に記載の精神疾患判定マーカー。
（１０）うつ病の重症度を同定する、配列番号４４、６６、及び８８～９３で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、（８）に記載の精神疾患判定マーカー。
（１１）配列番号９４～９８で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症／双極性障害である、（１）に記載の精神疾患判定マーカー。
（１２）配列番号２、４、５５、９９～１０３、１４５、１４６、１７９、及び２３３～２４０で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症／うつ病である、（１）に記載の精神疾患判定マーカー。
（１３）配列番号１０４～１２２、及び２４１で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患がうつ病／双極性障害である、（１）に記載の精神疾患判定マーカー。
（１４）配列番号１２４～１２６で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症及びうつ病のいずれであるかを判定する、（１）に記載の精神疾患判定マーカー。
（１５）配列番号１２７～１３１、及び２４２～２４６で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症及び双極性障害のいずれであるかを判定する、（１）に記載の精神疾患判定マーカー。
（１６）配列番１３１～１３６、２４７、及び２４８で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患がうつ病及び双極性障害のいずれであるかを判定する、（１）に記載の精神疾患判定マーカー。
（１７）配列番号１３、３４、及び１３７～１４０で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症、うつ病又は双極性障害である、（１）に記載の精神疾患判定マーカー。
（１８）精神疾患判定方法であって、被験者及び健常者から採取された試料の単位量あたりに含まれる（２）、（３）、（５）、（６）、（８）、（９）、（１１）～（１３）及び（１７）に記載のいずれか一以上の精神疾患判定マーカーの量を測定してその測定値を得る測定工程、前記測定工程で得られた被験者及び健常者の測定値を比較して、その結果に基づいて被験者における精神疾患を判定する比較判定工程を含む前記方法。
（１９）前記測定工程において（２）又は（３）に記載の精神疾患判定マーカーから統合失調症を判定する、（１８）に記載の方法。
（２０）前記（３）に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて統合失調症の亜型をさらに判定する、（１９）に記載の方法。
（２１）前記測定工程において（５）又は（６）に記載の精神疾患判定マーカーから双極性障害を判定する、（１８）に記載の方法。
（２２）前記（６）に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて双極性障害の亜型をさらに判定する、（２１）に記載の方法。
（２３）前記測定工程において（８）又は（９）に記載の精神疾患判定マーカーからうつ病を判定する、（１８）に記載の方法。
（２４）前記（８）に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいてうつ病の亜型をさらに判定する、（２３）に記載の方法。
（２５）前記測定工程において（１１）に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて統合失調症／双極性障害を判定する、（１８）に記載の方法。
（２６）前記測定工程において（１２）に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて統合失調症／うつ病を判定する、（１８）に記載の方法。
（２７）前記測定工程において（１３）に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいてうつ病／双極性障害を判定する、（１８）に記載の方法。
（２８）前記測定工程において（１７）に記載の精神疾患判定マーカーから統合失調症、うつ病又は双極性障害を判定する、（１８）に記載の方法。
（２９）前記測定工程において（４）に記載の精神疾患判定マーカーの量を測定し、該測定工程で得られた測定値に基づいて統合失調症の重症度を判定する重症度判定工程をさらに含む、（１９）又は（２０）に記載の方法。
（３０）前記測定工程において（７）に記載の精神疾患判定マーカーの量を測定し、該測定工程で得られた測定値に基づいて双極性障害の重症度を判定する重症度判定工程をさらに含む、（２１）又は（２２）に記載の方法。
（３１）前記測定工程において（１０）に記載の精神疾患判定マーカーの量を測定し、該測定工程で得られた測定値に基づいてうつ病の重症度を判定する重症度判定工程をさらに含む、（２３）又は（２４）に記載の方法。
（３２）前記測定工程において（１５）に記載の精神疾患判定マーカーの量を測定し、前記比較判定工程において測定工程で得られた測定値に基づいて統合失調症及び双極性障害のいずれであるかを判定する、（２５）に記載の方法。
（３３）前記測定工程において（１４）に記載の精神疾患判定マーカーの量を測定し、前記比較判定工程において測定工程で得られた測定値に基づいて統合失調症及びうつ病のいずれであるかを判定する、（２６）に記載の方法。
（３４）前記測定工程において（１６）に記載の精神疾患判定マーカーの量を測定し、前記比較判定工程において測定工程で得られた測定値に基づいてうつ病及び双極性障害のいずれであるかを判定する、（２７）に記載の方法。
（３５）前記測定工程において二以上の異なる精神疾患判定マーカーを測定する、（１８）～（３４）のいずれかに記載の方法。
（３６）前記試料が脳脊髄液である、（１８）～（３５）のいずれかに記載の方法。 (1) A psychiatric disease determination marker comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 1 to 249 contained in cerebrospinal fluid, a mutant thereof, or a fragment thereof.
(2) selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 1, 3, 5, 6, 7, 8-13, 15-33, 35-54, 56-60, 141-144, 147-162, and 249 The psychiatric disorder determination marker according to (1), wherein the psychiatric disorder is schizophrenia.
(3) A protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 7 to 13, 15 to 25, 141 to 144, 147, 148, and 249, and variants thereof, which subdivide schizophrenia into subtypes Or the psychiatric disorder determination marker according to (2), comprising:
(4) selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 9, 13, 26-29, 31-33, 35-54, 56-60, and 159-162, which identify the severity of schizophrenia The psychiatric disease determination marker according to (2), comprising a protein, a mutant thereof, or a fragment thereof.
(5) consisting of a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 47, 61 to 69, 114, and 163 to 173, a variant thereof, or a fragment thereof, wherein the mental illness is bipolar disorder ( The psychiatric disorder determination marker according to 1).
(6) A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 63, 64, 114, and 163 to 169, a variant thereof, or a fragment thereof, which subdivides bipolar disorder into subtypes. The marker for determining mental illness according to 5).
(7) consisting of a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 47, 65 to 69, and 173, a variant thereof, or a fragment thereof, which identifies the severity of bipolar disorder; The psychiatric disorder determination marker described.
(8) a protein selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 13, 14, 17, 44, 56, 66, 70 to 93, 123, 174 to 178, and 180 to 232, a variant thereof, or a The psychiatric disorder determination marker according to (1), comprising a fragment, wherein the psychiatric disorder is depression.
(9) A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 66, 76 to 87, and 174 to 178, and 180 to 192, or a variant or fragment thereof, which subdivides depression into subtypes The psychiatric disease determination marker according to (8), comprising:
(10) The protein according to (8), comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 44, 66, and 88 to 93, a variant thereof, or a fragment thereof, which identifies the severity of depression A marker for determining mental illness.
(11) The protein according to (1), comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 94 to 98, a variant thereof, or a fragment thereof, wherein the mental illness is schizophrenia / bipolar disorder Psychiatric disease determination marker.
(12) A mental disorder comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 2, 4, 55, 99 to 103, 145, 146, 179, and 233 to 240, a variant thereof, or a fragment thereof. The psychiatric disorder determination marker according to (1), wherein is schizophrenia / depression.
(13) A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 104 to 122 and 241; a variant thereof; or a fragment thereof; wherein the mental illness is depression / bipolar disorder; The psychiatric disorder determination marker described.
(14) A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 124 to 126, a variant thereof, or a fragment thereof, and determining whether the mental illness is schizophrenia or depression ( The psychiatric disorder determination marker according to 1).
(15) A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 127 to 131 and 242 to 246, a variant thereof, or a fragment thereof, and the psychiatric disorder is schizophrenia or bipolar disorder The psychiatric disorder determination marker according to (1), wherein
(16) A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 131 to 136, 247, and 248, a variant thereof, or a fragment thereof, and whether the mental illness is depression or bipolar disorder The psychiatric disorder determination marker according to (1), wherein:
(17) A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 13, 34, and 137 to 140, a variant thereof, or a fragment thereof, wherein the mental disorder is schizophrenia, depression, or bipolar disorder The psychiatric disorder determination marker according to (1), wherein:
(18) A method for determining a mental illness, which is included per unit amount of a sample collected from a subject and a healthy person (2), (3), (5), (6), (8), (9) (11) to (13) and (17) a measurement step of measuring the amount of any one or more psychiatric disease determination markers to obtain a measurement value thereof, a subject obtained in the measurement step and healthy subjects The said method including the comparison determination process which compares a measured value and determines the mental disease in a test subject based on the result.
(19) The method according to (18), wherein schizophrenia is determined from the mental disease determination marker according to (2) or (3) in the measurement step.
(20) The method according to (19), wherein a subtype of schizophrenia is further determined based on the measured value of the psychiatric disorder determination marker according to (3) and a predetermined cutoff value.
(21) The method according to (18), wherein bipolar disorder is determined from the mental disease determination marker according to (5) or (6) in the measurement step.
(22) The method according to (21), wherein a subtype of bipolar disorder is further determined based on a measured value of the psychiatric disorder determination marker according to (6) and a predetermined cutoff value.
(23) The method according to (18), wherein depression is determined from the mental disease determination marker according to (8) or (9) in the measurement step.
(24) The method according to (23), wherein a subtype of depression is further determined based on the measured value of the psychiatric disorder determination marker according to (8) and a predetermined cutoff value.
(25) The method according to (18), wherein in the measurement step, schizophrenia / bipolar disorder is determined based on the measured value of the psychiatric disorder determination marker according to (11) and a predetermined cutoff value.
(26) The method according to (18), wherein schizophrenia / depression is determined based on the measured value of the psychiatric disorder determination marker according to (12) and a predetermined cut-off value in the measurement step.
(27) The method according to (18), wherein in the measurement step, depression / bipolar disorder is determined based on the measured value of the psychiatric disorder determination marker according to (13) and a predetermined cutoff value.
(28) The method according to (18), wherein in the measurement step, schizophrenia, depression or bipolar disorder is determined from the psychiatric disorder determination marker according to (17).
(29) A severity determination step of measuring the amount of the psychiatric disorder determination marker according to (4) in the measurement step and determining the severity of schizophrenia based on the measurement value obtained in the measurement step The method according to (19) or (20).
(30) A severity determination step of measuring the amount of the psychiatric disorder determination marker according to (7) in the measurement step and determining the severity of bipolar disorder based on the measurement value obtained in the measurement step The method according to (21) or (22).
(31) It further includes a severity determination step of measuring the amount of the psychiatric disorder determination marker according to (10) in the measurement step and determining the severity of depression based on the measurement value obtained in the measurement step. , (23) or (24).
(32) In the measurement step, the amount of the psychiatric disease determination marker according to (15) is measured, and based on the measurement value obtained in the measurement step in the comparison determination step, the schizophrenia or bipolar disorder The method according to (25), wherein:
(33) In the measurement step, the amount of the psychiatric disease determination marker according to (14) is measured, and whether it is schizophrenia or depression based on the measurement value obtained in the measurement step in the comparison determination step The method according to (26), wherein:
(34) In the measurement step, the amount of the psychiatric disease determination marker according to (16) is measured, and whether the depression or bipolar disorder is based on the measurement value obtained in the measurement step in the comparison determination step The method according to (27), wherein:
(35) The method according to any one of (18) to (34), wherein two or more different psychiatric disease determination markers are measured in the measurement step.
(36) The method according to any one of (18) to (35), wherein the sample is cerebrospinal fluid.
　本明細書は本願の優先権の基礎である日本国特許出願2014-102090号の明細書及び／又は図面に記載される内容を包含する。 This specification includes the contents described in the specification and / or drawings of Japanese Patent Application No. 2014-102090, which is the basis of the priority of the present application.
　本発明の精神疾患判定マーカーによれば、被験者における統合失調症、うつ病、又は双極性障害等の精神疾患を簡便かつ高精度に判定可能なバイオマーカーを提供することができる。この精神疾患判定マーカーは、被験者における統合失調症、うつ病、又は双極性障害等の罹患鑑別、亜型の同定、及び／又は重症度の同定を行うことができる。 According to the psychiatric disease determination marker of the present invention, a biomarker capable of easily and accurately determining a psychiatric disease such as schizophrenia, depression, or bipolar disorder in a subject can be provided. This psychiatric disorder determination marker can be used to identify morbidity such as schizophrenia, depression, or bipolar disorder in a subject, identification of subtypes, and / or identification of severity.
　本発明の精神疾患判定方法によれば、前記精神疾患判定マーカーを用いて被験者における各精神疾患の罹患可能性、亜型の同定、及び／又は重症度の同定を高精度に判定する方法を提供することができる。 According to the mental illness determination method of the present invention, there is provided a method for accurately determining morbidity, subtype identification, and / or severity identification of each mental illness in a subject using the mental illness determination marker. can do.
本発明の精神疾患判定マーカーの分類群を示す概念図である。It is a conceptual diagram which shows the classification group of the mental disease determination marker of this invention. 健常者と各精神疾患患者におけるうつ病用精神疾患判定マーカーSCFsRの測定値分布図を示す。各プロット群における横直線は、そのプロット群における平均値を示す（他の図において、同様とする）。図中、Nはその精神疾患患者の総数を、またDFはN中の薬剤フリー（Drug Free）患者数を示す（他の図において、同様とする）。The distribution chart of the measured value of the depression psychiatric disorder determination marker SCFsR in healthy individuals and patients with each psychiatric disorder is shown. A horizontal straight line in each plot group indicates an average value in the plot group (the same applies to other drawings). In the figure, N indicates the total number of patients with the mental illness, and DF indicates the number of drug-free patients in N (the same applies to other figures). 統合失調症を亜型に細分類する精神疾患判定マーカーUNC5H3の健常者と各精神疾患患者における測定値分布図を示す。破線（95％）は、健常者群における95パーセンタイルに相当するカットオフ値である。The distribution chart of the measured values in healthy individuals and patients with each mental illness of the mental illness determination marker UNC5H3 that subdivides schizophrenia into subtypes is shown. The broken line (95%) is a cutoff value corresponding to the 95th percentile in the healthy group. 統合失調症重症度判定用の精神疾患判定マーカーであるST4S6の測定値と統合失調症の評価尺度PANSSによる評価値との関係を示す。The relationship between the measured value of ST4S6, which is a psychiatric disorder determination marker for determining the severity of schizophrenia, and the evaluation value based on the schizophrenia evaluation scale PANSS is shown. 健常者と各精神疾患患者におけるうつ病／双極性障害用精神疾患判定マーカーLYN Bの測定値分布図を示す。The distribution chart of the measured value of the LYN B marker for mental disorders for depression / bipolar disorder in healthy individuals and patients with various mental disorders is shown. 図５に示したLYN Bの測定値分布図に、健常者群における95パーセンタイルをカットオフ値として記入した図である。It is the figure which entered 95th percentile in a healthy subject group as a cut-off value in the measured value distribution map of LYN | B shown in FIG. 統合失調症とうつ病のいずれに罹患しているかを判定する精神疾患判定マーカーPTHの測定値分布を示す図である。It is a figure which shows the measured value distribution of the psychiatric disorder determination marker PTH which determines whether it is suffering from schizophrenia or depression. 健常者か精神疾患患者かを判定することができる精神疾患判定マーカーVEGF sR3の測定値分布を示す図である。カットオフ値は、健常者群における95パーセンタイルとした。It is a figure which shows the measured value distribution of the psychiatric disorder determination marker VEGF | sR3 which can determine whether it is a healthy person or a mental illness patient. The cutoff value was the 95th percentile in the healthy group. うつ病用精神疾患判定マーカーであるFibrinogenの測定値分布を示す図である。（ａ）に示すようにFibrinogenは、うつ病患者（MDD）の一部で顕著な亢進がみられた。また、（ｂ）に示すようにSOMAscanによるFibrinogen測定値は、ELISAによる測定値とよく相関していた。さらに、異なる検体コホートで、Fibrinogenを用いてELISAにより解析したところ、（ｃ）に示すようにうつ病の一群で顕著な亢進が認められ、Fibrinogenがうつ病用の精神疾患判定マーカーとして機能し得る再現性が得られた。It is a figure which shows the measured value distribution of Fibrinogen which is a mental disease determination marker for depression. As shown in (a), fibrinogen was markedly enhanced in a part of depressed patients (MDD). Further, as shown in (b), the fibrinogen measured value by SOMAscan correlated well with the measured value by ELISA. Furthermore, when analyzed by ELISA using fibrinogen in different specimen cohorts, a marked increase was observed in a group of depression as shown in (c), and fibrinogen could function as a mental disease determination marker for depression Reproducibility was obtained. 統合失調症用精神疾患判定マーカーであるPIGFによる統合失調症検体の測定値と統合失調症認知機能簡易評価尺度(BACS)の符号課題の評価値との関係を示す図である。（ａ）に示すようにPIGF濃度とBACS符号課題の評価値との間には相関があり、PIGF濃度が高いほど、符号課題の評価値が低く、症状が重かった。また、（ｂ）に示すようにPIGFは統合失調症群（Sz）及びうつ病の一部で顕著な亢進がみられた。したがって、PIGFを精神疾患判定マーカーに用いることで、統合失調症に罹患している可能性とその重症度を判定することができる。It is a figure which shows the relationship between the measured value of the schizophrenia sample by PIGF which is the mental disease determination marker for schizophrenia, and the evaluation value of the code task of a schizophrenia cognitive function simple evaluation scale (BACS). As shown in (a), there is a correlation between the PIGF concentration and the evaluation value of the BACS code task. The higher the PIGF concentration, the lower the evaluation value of the code task and the more severe the symptoms. In addition, as shown in (b), PIGF was remarkably enhanced in the schizophrenia group (Sz) and a part of depression. Therefore, by using PIGF as a psychiatric disorder determination marker, it is possible to determine the possibility and severity of schizophrenia. うつ病用精神疾患判定マーカーであるα1-Antitrypsinの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのα1-Antitrypsinの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。図中、Contは健常者を、Szは統合失調症を、MDDはうつ病を、そしてBDは双極性障害を示す。以下の図で同様とする。It is a figure which shows the measured value distribution of (alpha) 1-Antitrypsin which is a mental disease determination marker for depression. (A) shows the measured value distribution of α1-Antitrypsin of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. In the figure, Cont indicates a healthy person, Sz indicates schizophrenia, MDD indicates depression, and BD indicates bipolar disorder. The same applies to the following figures. 統合失調症／うつ病用精神疾患判定マーカーであるAdiponectinの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのAdiponectinの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Adiponectin which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of Adiponectin in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Adiponectin in the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるAntithrombin IIIの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのAntithrombin IIIの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Antithrombin III which is a mental disease determination marker for depression. (A) shows the measured value distribution of Antithrombin III in the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるApo Bの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのApo Bの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Apo B which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of Apo B of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるATS13の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのATS13の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of ATS13 which is a mental disease determination marker for depression. (A) shows the measured value distribution of ATS13 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of ATS13 of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 双極性障害／うつ病用精神疾患判定マーカーであるb-ECGFの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのb-ECGFの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of b-ECGF which is a bipolar disorder / depression mental disease determination marker. (A) shows the measured value distribution of b-ECGF in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of b-ECGF in the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるC1-Esterase Inhibitorの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのC1-Esterase Inhibitorの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of C1-Esterase (TM) Inhibitor which is a mental disease determination marker for depression. (A) shows the measured value distribution of the C1-Esterase Inhibitor of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるC3aの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのC3aの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of C3a which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of C3a of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of C3a of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるC5の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのC5の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。いずれの図でも、うつ病（MDD）の一部で顕著な亢進がみられた。（Ｄ）は、ELISAによるC5の測定値とSOMAscanによる測定値の相関を示す図である。It is a figure which shows the measured value distribution of C5 which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of C5 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of C5 of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. In both figures, there was a marked increase in some depression (MDD). (D) is a figure which shows the correlation of the measured value of C5 by ELISA, and the measured value by SOMAscan. うつ病用精神疾患判定マーカーであるCD5Lの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのCD5Lの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of CD5L which is a mental disease determination marker for depression. (A) shows the measured value distribution of CD5L of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of CD5L of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるContactin-4の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのContactin-4の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Contactin-4 which is a mental disorder determination marker for schizophrenia / depression. (A) shows the measured value distribution of Contactin-4 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Contactin-4 of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるCRPの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのCRPの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。いずれの図でも、うつ病（MDD）の一部で顕著な亢進がみられた。（Ｄ）は、ELISAによるCRPの測定値とSOMAscanによる測定値の相関を示す図である。It is a figure which shows the measured value distribution of CRP which is a schizophrenia / depression mental disease determination marker. (A) shows the CRP measurement value distribution of the first sample cohort shown in Table 12, and (b) shows the CRP measurement value distribution of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. In both figures, there was a marked increase in some depression (MDD). (D) is a figure which shows the correlation of the measured value of CRP by ELISA, and the measured value by SOMAscan. 統合失調症／うつ病用精神疾患判定マーカーであるCystatin Cの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのCystatin Cの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Cystatin (C) which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of Cystatin C of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるEphrin-A5の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのEphrin-A5の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Ephrin-A5 which is a mental disease determination marker for depression. (A) shows the measured value distribution of Ephrin-A5 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるESAMの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのESAMの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of ESAM which is a schizophrenia / depression mental disease determination marker. (A) shows the ESAM measurement distribution of the first sample cohort shown in Table 12, and (b) shows the ESAM measurement value distribution of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるFGF-12の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのFGF-12の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of FGF-12 which is a mental disease determination marker for depression. (A) shows the measured value distribution of FGF-12 in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of FGF-12 in the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるFSTL3の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのFSTL3の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of FSTL3 which is a mental disease determination marker for depression. (A) shows the measured value distribution of FSTL3 of the first sample cohort shown in Table 12, and (b) shows the FSTL3 measured value distribution of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症用精神疾患判定マーカーであるGDF-9の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのGDF-9の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of GDF-9 which is a mental disease determination marker for schizophrenia. (A) shows the measured value distribution of GDF-9 in the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるGP1BAの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのGP1BAの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of GP1BA which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of GP1BA of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of GP1BA of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症用精神疾患判定マーカーであるIL-13 Ra1の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのIL-13 Ra1の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of IL-13 (R) Ra1 which is a psychiatric disorder determination marker for schizophrenia. (A) shows the measured value distribution of IL-13 Ra1 of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病／双極性障害用精神疾患判定マーカーであるJAM-Bの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのJAM-Bの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of JAM-B which is a psychiatric disorder determination marker for schizophrenia / depression / bipolar disorder. (A) shows the measured value distribution of JAM-B of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるMATN2の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのMATN2の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of MATN2 which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of MATN2 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MATN2 of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 双極性障害用精神疾患判定マーカーであるMEK1の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのMEK1の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of MEK1 which is a psychiatric disease determination marker for bipolar disorders. (A) shows the measured value distribution of MEK1 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MEK1 of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 双極性障害用精神疾患判定マーカーであるMidkineの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのMidkineの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Midkine which is a mental disease determination marker for bipolar disorders. (A) shows the measured value distribution of the Midkine of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 双極性障害用精神疾患判定マーカーであるMIFの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのMIFの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of MIF which is a psychiatric disorder determination marker for bipolar disorders. (A) shows the MIF measurement value distribution of the first sample cohort shown in Table 12, and (b) shows the MIF measurement value distribution of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるMISの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのMISの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of MIS which is a mental disease determination marker for depression. (A) shows the MIS measurement value distribution of the first sample cohort shown in Table 12, and (b) shows the MIS measurement value distribution of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症用精神疾患判定マーカーであるMMP-2の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのMMP-2の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of MMP-2 which is a mental disease determination marker for schizophrenia. (A) shows the measured value distribution of MMP-2 in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MMP-2 in the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 双極性障害用精神疾患判定マーカーであるMSPの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのMSPの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of MSP which is a psychiatric disease determination marker for bipolar disorders. (A) shows the measured value distribution of MSP of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of MSP of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるNectin-like protein 1の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのNectin-like protein 1の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Nectin-like protein 1 which is a mental illness determination marker for depression. (A) shows the measured value distribution of Nectin-like protein 1 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるNotch 1の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのNotch 1の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Notch * 1 which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of Notch 1 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるProperdinの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのProperdinの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Properdin which is a mental disease determination marker for depression. (A) shows the measured value distribution of Properdin of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Properdin of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 双極性障害用精神疾患判定マーカーであるTFPIの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのTFPIの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of TFPI which is a psychiatric disorder determination marker for bipolar disorders. (A) shows the measured value distribution of TFPI of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of TFPI of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるTIG2の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのTIG2の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of TIG2 which is a mental disease determination marker for depression. (A) shows the measured value distribution of TIG2 of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of TIG2 of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるTransketolaseの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのTransketolaseの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Transketolase which is a mental disease determination marker for depression. (A) shows the measured value distribution of Transketolase in the first sample cohort shown in Table 12, and (b) shows the measured value distribution of Transketolase in the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病用精神疾患判定マーカーであるTSG-6の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのTSG-6の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of TSG-6 which is a schizophrenia / depression mental disease determination marker. (A) shows the measured value distribution of TSG-6 in the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. うつ病用精神疾患判定マーカーであるTWEAKの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのTWEAKの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of TWEAK which is a mental disease determination marker for depression. (A) shows the measured value distribution of TWEAK of the first sample cohort shown in Table 12, and (b) shows the measured value distribution of TWEAK of the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症精神疾患判定マーカーであるUNC5H3の測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのUNC5H3の測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of UNC5H3 which is a schizophrenia mental disease determination marker. (A) shows the measured value distribution of UNC5H3 of the first sample cohort shown in Table 12 and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion. 統合失調症／うつ病／双極性障害用精神疾患判定マーカーであるVasoactive Intestinal Peptideの測定値分布を示す図である。（ａ）は表１２に示す第１検体コホートの、また（ｂ）は表１３に示す第２検体コホートのVasoactive Intestinal Peptideの測定値分布を示す。（ｃ）は（ａ）及び（ｂ）のデータをZ変換により結合した図である。It is a figure which shows the measured value distribution of Vasoactive®Intestinal®Peptide which is a psychiatric disease determination marker for schizophrenia / depression / bipolar disorder. (A) shows the measured value distribution of Vasoactive Intestinal Peptide of the first sample cohort shown in Table 12, and (b) shows the second sample cohort shown in Table 13. (C) is the figure which combined the data of (a) and (b) by Z conversion.
１．精神疾患判定マーカー
１－１．概要
　本発明の第１の態様は精神疾患判定マーカーである。本発明の精神疾患判定マーカーは、タンパク質又はその断片から構成され、被験者由来の試料に含まれる量を測定することで、該被験者における精神疾患を判定するバイオマーカーとして利用することができる。 1. Psychiatric disease determination marker 1-1. Outline | summary The 1st aspect of this invention is a psychiatric disorder determination marker. The psychiatric disorder determination marker of the present invention is composed of a protein or a fragment thereof, and can be used as a biomarker for determining a psychiatric disorder in a subject by measuring the amount contained in a subject-derived sample.
１－２．構成
　本明細書において使用する用語について、以下で定義する。
　精神疾患とは、広義には外因性、心因性、又は内因性の脳の機能的又は器質的障害の総称をいう。しかしながら、本明細書において「精神疾患」とは、特に断りのない限り、従来の精神病理学の診断カテゴリーにおいて統合失調症、うつ病、又は双極性障害に分類される3つの症候群に属する図１に示す狭義の精神疾患を意味する。 1-2. Configuration Terms used in this specification are defined below.
Psychiatric disorder broadly refers to a generic term for exogenous, psychogenic, or intrinsic brain functional or organic disorders. However, unless otherwise specified, the term “psychiatric disorder” in the present specification refers to FIG. 1 belonging to three syndromes classified as schizophrenia, depression, or bipolar disorder in the diagnostic category of conventional psychopathology. Means a narrowly defined mental illness.
　本明細書において「精神疾患（を）判定（する）」とは、被験者における精神疾患の罹患鑑別、精神疾患の亜型の同定、及び／又は罹患している精神疾患の重症度の同定をいう。 As used herein, “determining (determining) a mental illness” refers to identifying a morbidity of a psychiatric disorder in a subject, identifying a subtype of the psychiatric disorder, and / or identifying the severity of the afflicted mental disorder. .
　本明細書において「精神疾患判定マーカー」とは、表１の各配列番号に示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片から構成される、精神疾患判定のためのバイオマーカーである。 In the present specification, the “mental disease determination marker” is a protein selected from the group consisting of the amino acid sequences shown in each SEQ ID NO. In Table 1, a variant thereof, or a fragment thereof for determining a mental disease. It is a biomarker.
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-I000002
Figure JPOXMLDOC01-appb-I000003
Figure JPOXMLDOC01-appb-I000004
Figure JPOXMLDOC01-appb-I000005
Figure JPOXMLDOC01-appb-I000006
Figure JPOXMLDOC01-appb-I000007
Figure JPOXMLDOC01-appb-T000001
Figure JPOXMLDOC01-appb-I000002
Figure JPOXMLDOC01-appb-I000003
Figure JPOXMLDOC01-appb-I000004
Figure JPOXMLDOC01-appb-I000005
Figure JPOXMLDOC01-appb-I000006
Figure JPOXMLDOC01-appb-I000007
　表１の各配列番号に示すアミノ酸配列からなるタンパク質は、いずれもヒト脳脊髄液（CSF）由来のタンパク質であって、本発明において被験者における統合失調症、うつ病又は双極性障害のいずれか一以上の精神疾患を判定するバイオマーカーとなり得る。なお、表１に記載の「UniProt No.」は、「http://www.uniprot.org/」に開示された各タンパク質のアクセッションナンバーである。 All of the proteins consisting of the amino acid sequences shown in each SEQ ID No. in Table 1 are proteins derived from human cerebrospinal fluid (CSF), and in the present invention, any one of schizophrenia, depression or bipolar disorder in a subject. It can be a biomarker for determining the above mental disorders. “UniProt No.” shown in Table 1 is the accession number of each protein disclosed in “http://www.uniprot.org/”.
　本明細書において「その変異体」とは、表１の各配列番号に示すアミノ酸配列からなる各タンパク質の変異体をいう。具体的には、表１に示す各タンパク質のアミノ酸配列において、1～複数個のアミノ酸の付加、欠失、又は置換を含むポリペプチド、又は表１に示す各タンパク質のアミノ酸配列と80％以上、85％以上、好ましくは90％以上、95％以上、96％以上、97％以上、98％以上、又は99％以上のアミノ酸同一性を有するポリペプチドをいう。ここで、「複数個」とは、2～10、2～9、2～8、2～7、2～6、2～5、2～4、又は2若しくは3個の整数をいう。また、「アミノ酸同一性」とは、二つのアミノ酸配列を整列（アラインメント）し、必要に応じてギャップを導入して、両者のアミノ酸一致度が最も高くなるようにしたときの表１に示す各タンパク質の全アミノ酸残基数に対する変異体の全アミノ酸残基数の同一アミノ酸残基の割合（％）をいう。アミノ酸同一性は、BLASTやFASTAによるタンパク質の検索システムを用いて決定することができる（Karlin，S．et al., 1993, Proc. Natl. Acad. Sci. USA, 90: 5873-5877；Altschul，S．F．et al., 1990, J. Mol. Biol., 215: 403-410；Pearson，W．R．et al., 1988, Proc. Natl. Acad. Sci. USA, 85: 2444-2448）。表１に示す各タンパク質の変異体の具体例には、スプライス変異等が挙げられる。 As used herein, “variant thereof” refers to a variant of each protein comprising the amino acid sequence shown in each SEQ ID NO in Table 1. Specifically, in the amino acid sequence of each protein shown in Table 1, a polypeptide containing addition, deletion, or substitution of one or more amino acids, or 80% or more of the amino acid sequence of each protein shown in Table 1, A polypeptide having 85% or more, preferably 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more amino acid identity. Here, “plurality” refers to 2 to 10, 2 to 9, 2 to 8, 2 to 7, 2 to 6, 2 to 5, 2 to 4, or 2 or 3 integers. In addition, “amino acid identity” refers to each of the amino acid sequences shown in Table 1 when the two amino acid sequences are aligned (aligned), and a gap is introduced as necessary so that the degree of amino acid coincidence between the two is maximized. The ratio (%) of the same amino acid residue of the total number of amino acid residues of the variant with respect to the total number of amino acid residues of the protein. Amino acid identity can be determined using protein search systems with BLAST and FASTA (Karlin, S. et al., 1993, Proc. Natl. Acad. Sci. USA, 90: 5873-5877; Altschul, S.F. et al., 1990, J. Mol. Biol., 215: 403-410; Pearson, WR et al., 1988, Proc. Natl. Acad. Sci. USA, 85: 2444-2448 ). Specific examples of the mutant of each protein shown in Table 1 include splice mutation.
　本明細書において「断片」とは、表１の各配列番号に示すアミノ酸配列からなる群から選択されるタンパク質又はその変異体の一部からなるペプチド断片であって、当該タンパク質を構成するアミノ酸配列のうち、7個以上全数未満、10個以上全数未満、15個以上全数未満、好ましくは20個以上全数未満、25個以上全数未満、35個以上全数未満、40個以上全数未満、又は50個以上全数未満の連続するアミノ酸残基を有する。好ましくは1又は複数のエピトープをそのアミノ酸配列内に保有するポリペプチド断片である。 In the present specification, the “fragment” is a peptide fragment consisting of a protein selected from the group consisting of the amino acid sequences shown in each SEQ ID No. in Table 1 or a part of a variant thereof, and the amino acid sequence constituting the protein Among them, 7 or more, less than 100, 10 or more, less than 15, 15 or more, less than 100, preferably 20 or more, less than 100, 25 or more, less than 35, 35 or more, less than 40, or more than 40, or 50 It has less than the total number of consecutive amino acid residues. Preferably, it is a polypeptide fragment carrying one or more epitopes within its amino acid sequence.
　本明細書では、以下、表１の「配列番号Xに示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片から構成されるバイオマーカー」を「配列番号Xに示すアミノ酸配列からなる群から選択されるタンパク質等」と表記する。ここで、Xは1～140の任意の整数である。 In the present specification, hereinafter, the “biomarker composed of a protein selected from the group consisting of the amino acid sequence shown in SEQ ID NO: X, a variant thereof, or a fragment thereof” in Table 1 is referred to as “amino acid sequence shown in SEQ ID NO: X”. A protein selected from the group consisting of. Here, X is an arbitrary integer from 1 to 140.
　本明細書において「被験者」とは、後述する第２態様の方法に供され、精神疾患を判定すべき対象となるヒト個体をいう。 In the present specification, the “subject” refers to a human individual that is subjected to the method of the second aspect described later and is a target for which a mental illness is to be determined.
　本発明の精神疾患判定マーカーは、被験者における統合失調症、うつ病又は双極性障害のいずれか一以上の精神疾患を判定することができる。以下の表２～５、７、９及び１１に示す精神疾患判定マーカーは、その検出属性に基づいて図１に示す１～７群に大別することができる。 The mental disease determination marker of the present invention can determine one or more mental diseases of schizophrenia, depression, or bipolar disorder in a subject. The mental disease determination markers shown in Tables 2 to 5, 7, 9, and 11 below can be roughly classified into groups 1 to 7 shown in FIG. 1 based on their detection attributes.
　以下、各群に属する精神疾患判定マーカーについて、具体的に説明をする。
（１）統合失調症を判定する群
　１群に属する精神疾患判定マーカーを表２に示す。 Hereinafter, the mental disease determination marker belonging to each group will be described in detail.
(1) Group to determine schizophrenia Table 2 shows markers for determining mental disorders belonging to Group 1.
Figure JPOXMLDOC01-appb-T000008
Figure JPOXMLDOC01-appb-I000009
Figure JPOXMLDOC01-appb-I000010
Figure JPOXMLDOC01-appb-T000008
Figure JPOXMLDOC01-appb-I000009
Figure JPOXMLDOC01-appb-I000010
　この群に属する精神疾患判定マーカーは、統合失調症を判定するマーカーである。具体的には、図１において1群で示す区画に属するマーカーであって、配列番号１、３、５、６、７、８～１３、１５～３３、３５～５４、５６～６０、１４１～１４４、１４７～１６２、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質等をいう。本群の精神疾患判定マーカーを用いれば、被験者における統合失調症の罹患鑑別、統合失調症の亜型の同定、及び／又は統合失調症の重症度の同定ができる。 The mental disease determination marker belonging to this group is a marker for determining schizophrenia. Specifically, it is a marker belonging to the section shown in group 1 in FIG. 1, and SEQ ID NOs: 1, 3, 5, 6, 7, 8 to 13, 15 to 33, 35 to 54, 56 to 60, 141 to A protein selected from the group consisting of the amino acid sequences represented by 144, 147 to 162, and 249. By using this group of psychiatric disease determination markers, it is possible to identify schizophrenia morbidity, identify schizophrenia subtypes, and / or identify the severity of schizophrenia in a subject.
　本群に属する精神疾患判定マーカーにおいて配列番号１、３、５、６、７、８～１３、１５～２５、３０、４６、１４１～１４４、１４７～１５８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質等は、被験者における統合失調症の罹患を鑑別することができる。 A group consisting of the amino acid sequences shown in SEQ ID NOs: 1, 3, 5, 6, 7, 8 to 13, 15 to 25, 30, 46, 141 to 144, 147 to 158, and 249 in the psychiatric disorder determination marker belonging to this group Or the like can be used to distinguish the morbidity of schizophrenia in a subject.
　本明細書において、「罹患（を）鑑別（する）」とは、被験者における特定の精神疾患の罹患可能性の高さを判断することをいう。したがって、配列番号１、３、５、６、７、８～１３、１５～２５、３０、４６、１４１～１４４、１４７～１５８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質等は、統合失調症罹患鑑別マーカーであり、このマーカーを1以上用いることで、被験体が統合失調症に罹患している可能性が高いことを鑑別できる。 In this specification, “differentiating (performing)” means determining the high possibility of the subject having a specific mental illness. Therefore, a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 1, 3, 5, 6, 7, 8 to 13, 15 to 25, 30, 46, 141 to 144, 147 to 158, and 249, etc. It is a schizophrenia disease differentiation marker, and by using one or more of these markers, it can be distinguished that a subject is highly likely to suffer from schizophrenia.
　本群に属する精神疾患判定マーカーにおいて配列番号１、３、５、６、１１、１２、３０、４６、１４１、１４２、１４９～１５８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質等は、統合失調症患者と健常者間の脳脊髄液試料に含まれる当該タンパク質等の存在量の有意差に基づいて単離されたバイオマーカーである。 Proteins selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 1, 3, 5, 6, 11, 12, 30, 46, 141, 142, 149 to 158, and 249 in the psychiatric disorder determination marker belonging to this group Is a biomarker isolated based on a significant difference in the abundance of the protein or the like contained in a cerebrospinal fluid sample between a schizophrenic patient and a healthy subject.
　本明細書において「健常状態」とは、本明細書における狭義のいずれの精神疾患にも罹患していない状態、好ましくは広義のいずれの精神疾患にも罹患していない健全な精神状態であることをいう。 As used herein, the term “healthy state” refers to a state that is not affected by any mental illness in the narrow sense herein, preferably a healthy mental state that is not affected by any mental illness in the broad sense. Say.
　本明細書において「有意」とは、統計学的に有意であることをいう。具体的には、被験者と健常者の測定値の差異を統計学的に処理したときに、両者間に有意差があることをいう。例えば、危険率（有意水準）が5％、1％、0.3％、0.2％又は0.1％より小さい場合が挙げられる。統計学的処理の検定方法は、有意性の有無を判断可能な公知の検定方法を適宜使用すればよく、特に限定しない。例えば、スチューデントt検定法、多重比較検定法を用いることができる。 In this specification, “significant” means statistically significant. Specifically, when the difference between the measured values of the subject and the healthy person is statistically processed, it means that there is a significant difference between the two. For example, the risk rate (significance level) may be less than 5%, 1%, 0.3%, 0.2%, or 0.1%. The test method for statistical processing is not particularly limited as long as a known test method capable of determining the presence or absence of significance is appropriately used. For example, Student's t test or multiple comparison test can be used.
　表２では、年齢・性を共変数とした共分散分析で危険率0.2％より小（p<0.002）を示すタンパク質を、健常との差を有する精神疾患判定マーカーとして「○」印を付けて示している。 In Table 2, a protein with a risk factor of less than 0.2% (p <0.002) in the covariance analysis with age and sex as covariates is marked with “○” as a psychiatric disorder determination marker with a difference from normal. Show.
　本群に属する精神疾患判定マーカーにおいて配列番号８～１３、１５～２５、１４１～１４４、１４７、１４８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質等は、統合失調症の罹患鑑別に加えて、統合失調症を亜型に細分類することができる判定マーカーである。 Proteins selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 8 to 13, 15 to 25, 141 to 144, 147, 148, and 249 in the psychiatric disorder determination marker belonging to this group are used to identify morbidity of schizophrenia. In addition, it is a determination marker that can subdivide schizophrenia into subtypes.
　本明細書において「亜型」とは、本発明の精神疾患判定マーカーを用いた生物学的根拠に基づいて分類される、各精神疾患の下位の分類群である。例えば、１群に属する統合失調症の亜型、２群に属する双極性障害の亜型、３群に属する及びうつ病の亜型が挙げられる。この他にも、４群に属する統合失調症／双極性障害の亜型、５群に属する統合失調症／うつ病の亜型、６群に属する双極性障害／うつ病の亜型が挙げられる。このような亜型は、例えば、「（精神疾患）の（判定マーカー分子名）亜型」として分類される。具体的には、配列番号７で示されるUNC5H3マーカーを用いた精神疾患の判定において、統合失調症と判定された被験者は、「統合失調症のUNC5H3亜型」と同定することができる。なお、４群に属する統合失調症／双極性障害、５群に属する統合失調症／うつ病、及び６群に属する双極性障害／うつ病についての詳細は、後述する。 In the present specification, the “subtype” is a subordinate classification group of each mental illness classified based on a biological basis using the mental illness determination marker of the present invention. For example, schizophrenia subtypes belonging to group 1, bipolar disorder subtypes belonging to group 2, subgroups belonging to group 3 and depression. Other examples include schizophrenia / bipolar subtypes belonging to group 4, subtypes of schizophrenia / depression belonging to group 5, bipolar disorder / depression subtypes belonging to group 6 . Such a subtype is classified, for example, as “(mental marker molecule name) subtype of (psychiatric disorder)”. Specifically, in the determination of mental illness using the UNC5H3 marker represented by SEQ ID NO: 7, the subject determined as schizophrenia can be identified as “UNC5H3 subtype of schizophrenia”. Details of schizophrenia / bipolar disorder belonging to group 4 and schizophrenia / depression belonging to group 5, and bipolar disorder / depression belonging to group 6 will be described later.
　本明細書において各精神疾患の亜型を同定可能な精神疾患判定マーカー（具体的には、統合失調症亜型用の配列番号７～１３、１５～２５、１４１～１４４、１４７、１４８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質等、後述する双極性障害亜型用の配列番号６３、６４、１１４、及び１６３～１６９で示すアミノ酸配列からなる群から選択されるタンパク質等、うつ病亜型用の配列番号６６、７６～８７、及び１７４～１７８、及び１８０～１９２で示すアミノ酸配列からなる群から選択されるタンパク質、統合失調症／双極性障害亜型用の配列番号９５～９８で示すアミノ酸配列からなる群から選択されるタンパク質等、統合失調症／うつ病亜型用の配列番号２、４、７、１００～１０３、１４５、１４６、１７９、及び２３３～２３９で示すアミノ酸配列からなる群から選択されるタンパク質等、及びうつ病／双極性障害亜型用の配列番号１０８～１２２で示すアミノ酸配列からなる群から選択されるタンパク質等）は、健常者群の脳脊髄液由来の試料に含まれる当該精神疾患判定マーカーの測定値に基づいて設定されるカットオフ値により単離されたバイオマーカーである。 Mental illness determination marker capable of identifying a subtype of each psychiatric disorder in the present specification (specifically, SEQ ID NOs: 7 to 13, 15 to 25, 141 to 144, 147, 148 for schizophrenia subtypes, and A protein selected from the group consisting of amino acid sequences shown by H.249, a protein selected from the group consisting of amino acid sequences shown by SEQ ID NOs: 63, 64, 114, and 163 to 169 for bipolar disorder subtypes described later, A protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 66, 76-87, and 174-178, and 180-192 for the depression subtype, SEQ ID NO: 95 for the schizophrenia / bipolar disorder subtype SEQ ID NOs: 2, 4, 7, 100 to 103, 145, 146 for schizophrenia / depression subtype, such as a protein selected from the group consisting of the amino acid sequences represented by ~ 98 179 and a protein selected from the group consisting of amino acid sequences represented by 233 to 239, and a protein selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 108 to 122 for depression / bipolar disorder subtypes) Is a biomarker isolated by a cut-off value set based on the measured value of the psychiatric disorder determination marker contained in the cerebrospinal fluid-derived sample of the healthy group.
　本明細書において「カットオフ値」とは、定量結果を陽性、陰性に分類するための境界値をいう。ここでいう陽性とは、精神疾患に罹患している可能性が高いことを、また陰性とは罹患していない可能性が高いことを、それぞれ示す。カットオフ値の設定法は特に限定しない。例えば、健常者群から得られた測定値をパーセンタイルで分類し、その分類に用いたパーセンタイル値をカットオフ値とすることができる。具体的には、健常者群から得られた測定値の95パーセンタイルで被験者の測定値の陽性又は陰性を分類した場合、95パーセンタイルがカットオフ値となる。他にも、Akobeng AKActa, 2007, Paediatr, 96:644-647に記載の算出方法に基づいて設定することができる。 In this specification, the “cut-off value” refers to a boundary value for classifying a quantitative result into positive and negative. Here, positive means that there is a high possibility of suffering from a mental illness, and negative means that there is a high possibility that the person is not affected. The method for setting the cutoff value is not particularly limited. For example, measurement values obtained from a group of healthy subjects can be classified by percentile, and the percentile value used for the classification can be used as a cutoff value. Specifically, when classifying a subject's measurement value positive or negative by the 95th percentile of the measurement value obtained from a group of healthy subjects, the 95th percentile is a cut-off value. In addition, it can be set based on the calculation method described in Akobeng AKActa, 2007, Paediatr, 96: 644-647.
　本明細書でパーセンタイル値をカットオフ値として用いる場合、5パーセンタイル又は95パーセンタイルのいずれかを用い、5パーセンタイル未満、又は95パーセンタイルより大の場合を陽性、5パーセンタイル以上、又は95パーセンタイル以下の場合を陰性とする。例えば、被験者におけるUNC5H3マーカーの測定値が健常者群の測定値の95パーセンタイルより大きい値を示した場合、その被検者は、統合失調症に罹患している可能性が高く、かつ統合失調症のUNC5H3亜型として細分類することができる。 When using the percentile value as a cut-off value in this specification, use either the 5th percentile or the 95th percentile, positive if less than 5th percentile or greater than 95th percentile, greater than 5th percentile, or less than 95th percentile Negative. For example, if a UNC5H3 marker measurement in a subject shows a value greater than the 95th percentile of a healthy group, the subject is likely to have schizophrenia and schizophrenia Can be subdivided into UNC5H3 subtypes.
　従来、一般的な精神疾患は、本人や家族等への問診結果や経過状況、構造化診断面接等により診断されていたが、同じ精神疾患内の個体間でも抗精神疾患薬（例えば、抗うつ薬等）による薬理効果に大きな差異が生じるという問題があった。この原因の一つとして、従来法では診断された精神疾患のカテゴリーが大き過ぎ、同一精神疾患内に複数の病態（本明細書でいうところの亜型）が存在していたために一部の病態にのみ薬理効果が認められた可能性が考えられる。各精神疾患を亜型に細分類可能な本発明の精神疾患判定マーカーを用いれば、従来の仮説的な精神疾患の診断基準に従うだけではなく、各精神疾患の分子病態を反映する、生物学的根拠に基づいた客観性のより高い分類が可能となる。 Conventionally, general mental illness has been diagnosed based on the results of interviews with the person or family, progress status, structured diagnostic interviews, etc., but antipsychotic drugs (for example, antidepressant) are also used among individuals within the same mental illness. There has been a problem that a large difference occurs in the pharmacological effects of drugs and the like. One reason for this is that the category of mental illness diagnosed by the conventional method is too large, and there are several pathological conditions (subtypes referred to in this specification) within the same mental illness. It is possible that only a pharmacological effect was observed. By using the mental illness determination marker of the present invention, which can subdivide each mental illness into subtypes, not only follows the conventional hypothetical mental illness diagnostic criteria but also reflects the molecular pathology of each psychiatric illness. Classification with higher objectivity based on the basis is possible.
　また本発明の精神疾患判定マーカーを用いれば、同一精神疾患内に存在し得る各亜型に応じた至適な治療が可能となる。亜型に細分類できる上記精神疾患判定マーカーは、精神疾患特異性が高く、統合失調症、双極性障害、又はうつ病を高精度に判定することができる他、従来法では分類し得ない新たな精神疾患群である統合失調症／双極性障害、統合失調症／うつ病、及びうつ病／双極性障害も高精度に判定することができる。 In addition, the use of the psychiatric disorder determination marker of the present invention enables optimal treatment according to each subtype that may exist in the same psychiatric disorder. The above psychiatric disease markers that can be subdivided into subtypes have high psychiatric disease specificity and can accurately determine schizophrenia, bipolar disorder, or depression, and are not newly classified by conventional methods. Psychiatric disorders such as schizophrenia / bipolar disorder, schizophrenia / depression, and depression / bipolar disorder can also be determined with high accuracy.
　表２では、健常者の95パーセンタイル及び5パーセンタイルをカットオフ値とし、その値を外れ、95パーセンタイルより大、又は5パーセンタイル未満の異常値を示すタンパク質を亜型に細分類可能な精神疾患判定マーカーとして示している。例えば、配列番号７～１３、１５～２０、１４１～１４４、１４７、１４８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質等は、健常者の95パーセンタイルをカットオフ値とし、その値よりも大きい場合には、統合失調症の罹患鑑別に加えて、統合失調症を亜型に細分類することができる。一方、配列番号２１～２５で示すアミノ酸配列からなる群から選択されるタンパク質等は、健常者の5パーセンタイルをカットオフ値とし、その値未満の場合には、統合失調症の罹患鑑別に加えて、統合失調症を亜型に細分類することができる。 In Table 2, the 95th percentile and the 5th percentile of healthy subjects are cut off values, and the psychiatric disease determination marker that can subdivide proteins that are out of that value and show abnormal values greater than 95th percentile or less than 5th percentile into subtypes. As shown. For example, a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 7 to 13, 15 to 20, 141 to 144, 147, 148, and 249 has the 95th percentile of a healthy person as a cutoff value, and the value If it is larger than that, it is possible to subdivide schizophrenia into subtypes in addition to the symptom diagnosis of schizophrenia. On the other hand, the protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 21 to 25 has a cut-off value of the 5th percentile of healthy individuals, and if it is less than that value, it is added to the symptom diagnosis of schizophrenia Schizophrenia can be subdivided into subtypes.
　また、本群に属する精神疾患判定マーカーにおいて配列番号９、１３、２６～３４、３５～５４、５６～６０、及び１５９～１６２で示すアミノ酸配列からなる群から選択されるタンパク質等の測定値は、統合失調症の評価尺度と相関性を示す。そのため、統合失調症の重症度を同定することができる。 Further, in the psychiatric disorder determination marker belonging to this group, measured values of proteins and the like selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 9, 13, 26 to 34, 35 to 54, 56 to 60, and 159 to 162 are , Showing a correlation with the schizophrenia rating scale. Therefore, the severity of schizophrenia can be identified.
　本明細書において「重症度」とは、各精神疾患の評価尺度に基づく症状の程度をいう。統合失調症の評価尺度には、PANSS（陽性・陰性症状評価尺度；Positive and Negative Syndrome Scale)、又はBACS(統合失調症認知機能簡易評価尺度；(Brief Assessment for Cognition in Schizophrenia)を利用すればよい。配列番号９、１３、２６～２９、３１～３３、３５～５４、５６～６０、及び１５９～１６２で示すアミノ酸配列からなる群から選択される各精神疾患判定マーカーと相関性を示す評価尺度は、以下に示す通りである。配列番号２６～３３で示すアミノ酸配列からなる群から選択されるタンパク質等は、PANSSを評価尺度としてピアソン相関分析においてp＜0.01で相関する、統合失調症の重症度を同定することができる精神疾患判定マーカーである。一方、配列番号９、１３、３４～５４、５６～６０、及び１５９～１６２で示すアミノ酸配列からなる群から選択されるタンパク質等は、BACSを評価尺度としてピアソン相関関数においてp＜0.01で相関する、統合失調症の重症度を同定することができる精神疾患判定マーカーである。 In this specification, “severity” refers to the degree of symptoms based on an evaluation scale for each mental disorder. For assessment of schizophrenia, PANSS (Positive and Negative Syndrome Scale) or BACS (Brief Assessment for Cognition in Schizophrenia) may be used. Evaluation scale showing correlation with each psychiatric disease determination marker selected from the group consisting of amino acid sequences shown in SEQ ID NOs: 9, 13, 26-29, 31-33, 35-54, 56-60, and 159-162 Is a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 26 to 33, which is correlated with p <0.01 in Pearson correlation analysis using PANSS as an evaluation scale, and severe schizophrenia A marker for determining the degree of psychiatric disorder, from the amino acid sequences represented by SEQ ID NOs: 9, 13, 34 to 54, 56 to 60, and 159 to 162 A protein or the like selected from the group is a psychiatric disease determination marker that can identify the severity of schizophrenia that correlates with P <0.01 in the Pearson correlation function using BACS as an evaluation scale.
　統合失調症の重症度を同定可能な前記精神疾患判定マーカーにおいて、配列番号９及び１３で示すアミノ酸配列からなる群から選択されるタンパク質等は、統合失調症の重症度のみならず、統合失調症の罹患の鑑別及び亜型の同定も可能な精神疾患判定マーカーである。 In the psychiatric disease determination marker capable of identifying the severity of schizophrenia, the protein or the like selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 9 and 13 is not only the severity of schizophrenia but also schizophrenia It is a psychiatric disease determination marker that can also be used for differentiation of diseases and identification of subtypes.
（２）双極性障害を判定する群
　２群に属する精神疾患判定マーカーを表３に示す。 (2) Group for determining bipolar disorder Table 3 shows the markers for determining mental disorders belonging to Group 2.
Figure JPOXMLDOC01-appb-T000011
Figure JPOXMLDOC01-appb-T000011
　この群に属する精神疾患判定マーカーは、双極性障害であることを判定できるマーカーである。具体的には、図１において２群で示す区画に属するマーカーであって、配列番号４７、６１～６９、１１４、及び１６３～１７３で示すアミノ酸配列からなる群から選択されるタンパク質等をいう。本群の精神疾患判定マーカーを用いれば、被験者における双極性障害の罹患鑑別、双極性障害の亜型の同定、及び／又は双極性障害の重症度の同定ができる。 The mental disease determination marker belonging to this group is a marker that can determine bipolar disorder. Specifically, it refers to a protein or the like belonging to the compartments shown in Group 2 in FIG. 1 and selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 47, 61 to 69, 114, and 163 to 173. Using this group of psychiatric disease determination markers, it is possible to identify the presence of bipolar disorder in a subject, identify the subtype of bipolar disorder, and / or identify the severity of bipolar disorder.
　本群に属する精神疾患判定マーカーにおいて配列番号６１～６４、１１４、及び１６３～１７２で示すアミノ酸配列からなる群から選択されるタンパク質等は、被験者における双極性障害の罹患を鑑別することができる。このうち、配列番号６１、６２、及び１７０～１７２で示すアミノ酸配列からなるタンパク質等は、双極性障害患者と健常者間の脳脊髄液由来の試料に含まれる当該タンパク質等の存在量の有意差に基づいて単離されたバイオマーカーである。 Proteins selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 61 to 64, 114, and 163 to 172 in the psychiatric disease determination marker belonging to this group can distinguish the presence of bipolar disorder in the subject. Among these, the protein comprising the amino acid sequences represented by SEQ ID NOs: 61, 62, and 170 to 172 is a significant difference in the abundance of the protein or the like contained in a cerebrospinal fluid-derived sample between a bipolar disorder patient and a healthy subject. Is a biomarker isolated on the basis of
　表３では、年齢・性を共変数とした共分散分析でp<0.002を示すタンパク質を、有意差を有する精神疾患判定マーカーとして「○」印を付けて示している。 In Table 3, a protein showing p <0.002 by covariance analysis with age and sex as covariates is marked with “◯” as a psychiatric disorder determination marker having a significant difference.
　また、配列番号６３、６４、１１４、及び１６３～１６９で示すアミノ酸配列からなるタンパク質等は、双極性障害を亜型に細分類することができるバイオマーカーである。表３では、健常者の95パーセンタイル及び5パーセンタイルをカットオフ値とし、95パーセンタイルより大、又は5パーセンタイル未満の異常値を示すタンパク質を亜型に細分類可能な精神疾患判定マーカーとして示している。例えば、配列番号６３、６４、１６４、１６６、及び１６８で示すアミノ酸配列からなるタンパク質等は、健常者の5パーセンタイルをカットオフ値とし、その値未満の場合には、双極性障害の罹患鑑別に加えて、双極性障害を亜型に細分類することができる。また、配列番号１１４、１６３、１６５、１６７、及び１６９で示すアミノ酸配列からなるタンパク質等は、健常者の95パーセンタイルをカットオフ値とし、その値より大きい場合には、双極性障害の罹患鑑別に加えて、双極性障害を亜型に細分類することができる。 In addition, proteins comprising the amino acid sequences represented by SEQ ID NOs: 63, 64, 114, and 163 to 169 are biomarkers that can subdivide bipolar disorder into subtypes. In Table 3, the 95th percentile and the 5th percentile of healthy individuals are shown as cut-off values, and proteins showing abnormal values greater than or less than the 95th percentile are shown as psychiatric disease determination markers that can be subdivided into subtypes. For example, the protein consisting of the amino acid sequences shown in SEQ ID NOs: 63, 64, 164, 166, and 168 has a cutoff value of the 5th percentile of healthy subjects, and if it is less than that value, it is classified as affected by bipolar disorder. In addition, bipolar disorder can be subdivided into subtypes. In addition, the protein consisting of the amino acid sequences shown in SEQ ID NOs: 114, 163, 165, 167, and 169 has a 95th percentile of a healthy person as a cut-off value, and if it is larger than that value, it is classified as a disease disorder of bipolar disorder. In addition, bipolar disorder can be subdivided into subtypes.
　一方、本群に属する精神疾患判定マーカーにおいて配列番号４７、及び６５～６９で示すアミノ酸配列からなる群から選択されるタンパク質等の測定値は、双極性障害におけるうつ症状の評価尺度であるHAM-D（ハミルトンうつ病評価尺度；Hamilton Depression Scale）と相関性を示すことから、その重症度の一側面を同定することができる。具体的には、配列番号４７、６５～６９、及び１７３で示すアミノ酸配列からなる群から選択されるタンパク質等は、HAM-Dを評価尺度としてピアソン相関関数においてp＜0.01で相関する、双極性障害の重症度を同定することができる精神疾患判定マーカーである。なお、配列番号４７で示す精神疾患判定マーカーActivin beta Aは、評価尺度をBACSとした場合には、前述のように統合失調症の重症度を同定できる。 On the other hand, the measured values of proteins and the like selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 47 and 65 to 69 in the psychiatric disorder determination marker belonging to this group are HAM-, which is an evaluation scale for depressive symptoms in bipolar disorder Since it is correlated with D (Hamilton Depression Scale), one aspect of its severity can be identified. Specifically, a protein or the like selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 47, 65 to 69, and 173 is a bipolar protein that correlates with p <0.01 in the Pearson correlation function using HAM-D as an evaluation measure. It is a psychiatric disorder determination marker that can identify the severity of a disorder. The psychiatric disorder determination marker Activin beta A represented by SEQ ID NO: 47 can identify the severity of schizophrenia as described above when the evaluation scale is BACS.
（３）うつ病を判定する群
　３群に属する精神疾患判定マーカーを表４に示す。 (3) Group for Determining Depression Table 4 shows psychiatric disease determination markers belonging to Group 3.
Figure JPOXMLDOC01-appb-T000012
Figure JPOXMLDOC01-appb-I000013
Figure JPOXMLDOC01-appb-T000012
Figure JPOXMLDOC01-appb-I000013
　この群に属する精神疾患判定マーカーは、うつ病を判定できるマーカーである。具体的には、図１において３群で示す区画に属するマーカーであって、配列番号１３、１４、１７、４４、５６、６６、７０～９３、１２３、１７４～１７８、及び１８０～２３２で示すアミノ酸配列からなる群から選択されるタンパク質等をいう。本群の精神疾患判定マーカーを用いれば、被験者におけるうつ病の罹患鑑別、うつ病の亜型の同定、及び／又はうつ病の重症度の同定ができる。 The mental disease determination marker belonging to this group is a marker that can determine depression. Specifically, it is a marker belonging to the section shown in group 3 in FIG. 1 and represented by SEQ ID NOs: 13, 14, 17, 44, 56, 66, 70 to 93, 123, 174 to 178, and 180 to 232 A protein or the like selected from the group consisting of amino acid sequences. Using this group of psychiatric disease determination markers, it is possible to identify the morbidity of depression, identify the subtype of depression, and / or identify the severity of depression in the subject.
　表４では、年齢・性を共変数とした共分散分析でp<0.001、p<0.003又はp<0.005を示すタンパク質を、有意差を有する精神疾患判定マーカーとして示している。 In Table 4, proteins showing p <0.001, p <0.003, or p <0.005 by covariance analysis with age and sex as covariates are shown as psychiatric disease determination markers having a significant difference.
　本群に属する精神疾患判定マーカーにおいて配列番号１３、１４、１７、５６、６６、７０～８７、１２３、及び１７４～１７８、及び１８０～２３２で示すアミノ酸配列からなる群から選択されるタンパク質等は、被験者におけるうつ病の罹患を鑑別することができる。このうち、配列番号１３、１４、１７、５６、７０～７５、８２、８３、１２３、１７４～１７８、１８０、及び１９３～２３２で示すアミノ酸配列からなる群から選択されるタンパク質等は、うつ病患者と健常者間の脳脊髄液由来の試料に含まれる当該タンパク質等の存在量の有意差に基づいて単離されたバイオマーカーである。ここで、配列番号２で示す精神疾患判定マーカーは、統合失調症の有意差に基づく罹患鑑別用の精神疾患判定マーカーでもあり、配列番号１３及び１７で示す精神疾患判定マーカーは、統合失調症の亜型同定用の精神疾患判定マーカーでもあり、配列番号５６で示す精神疾患判定マーカーは、統合失調症の重症度同定用の精神疾患判定マーカーでもある。 Proteins selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 13, 14, 17, 56, 66, 70 to 87, 123, and 174 to 178, and 180 to 232 in the mental disease determination marker belonging to this group, It is possible to differentiate the morbidity of depression in a subject. Among these, proteins selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 13, 14, 17, 56, 70 to 75, 82, 83, 123, 174 to 178, 180, and 193 to 232 are depression, etc. It is a biomarker isolated based on a significant difference in the abundance of the protein or the like contained in a cerebrospinal fluid-derived sample between a patient and a healthy person. Here, the psychiatric disorder determination marker represented by SEQ ID NO: 2 is also a psychiatric disorder determination marker for morbidity discrimination based on a significant difference in schizophrenia, and the psychiatric disorder determination marker represented by SEQ ID NOs: 13 and 17 is a schizophrenia marker. It is also a mental disease determination marker for subtype identification, and the mental disease determination marker represented by SEQ ID NO: 56 is also a mental disease determination marker for identifying the severity of schizophrenia.
　また、配列番号６６、７６～８７、１７４～１７８、及び１８０～１９２で示すアミノ酸配列からなる群から選択されるタンパク質等は、うつ病を亜型に細分類することができるバイオマーカーである。表４では、健常者の95パーセンタイル及び5パーセンタイルをカットオフ値とし、95パーセンタイルより大、又は5パーセンタイル未満の異常値を示すタンパク質を亜型に細分類可能な精神疾患判定マーカーとして示している。例えば、配列番号６６、７６～８１、１７６～１７８、１８１～１８５、及び１８９～１９２で示すアミノ酸配列からなる群から選択されるタンパク質等は、健常者の95パーセンタイルをカットオフ値とし、その値より大の場合には、うつ病の罹患鑑別に加えて、うつ病を亜型に細分類することができる。一方、配列番号８２～８７、１７４、１７５、１８０、及び１８６～１８８で示すアミノ酸配列からなる群から選択されるタンパク質等は、健常者の5パーセンタイルをカットオフ値とし、その値未満の場合には、うつ病の罹患鑑別に加えて、うつ病を亜型に細分類することができる。ここで、配列番号６６で示す精神疾患判定マーカーFibrinogenは、評価尺度HAM-Dを用いた場合には、双極性障害の重症度を同定可能な精神疾患判定マーカーとなるが、健常者群の脳脊髄液由来の試料に含まれるFibrinogenの測定値に基づいて決定されるカットオフ値に基づいて同定した場合には、うつ病の罹患鑑別及び重症度の同定用マーカーとなり得る。 In addition, proteins selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 66, 76 to 87, 174 to 178, and 180 to 192 are biomarkers that can subdivide depression into subtypes. In Table 4, the 95th percentile and the 5th percentile of healthy subjects are shown as cut-off values, and proteins showing abnormal values larger than the 95th percentile or less than the 5th percentile are shown as psychiatric disease determination markers that can be subdivided into subtypes. For example, a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 66, 76 to 81, 176 to 178, 181 to 185, and 189 to 192 has the 95th percentile of a healthy person as a cutoff value, In larger cases, depression can be subdivided into subtypes in addition to identifying the presence of depression. On the other hand, a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 82 to 87, 174, 175, 180, and 186 to 188 has a 5th percentile of a healthy person as a cutoff value, and is less than that value Can subdivide depression into subtypes in addition to identifying depression. Here, the psychiatric disease determination marker Fibrinogen represented by SEQ ID NO: 66 is a psychiatric disease determination marker that can identify the severity of bipolar disorder when the evaluation scale HAM-D is used. When identified based on the cut-off value determined based on the measured value of fibrinogen contained in the spinal fluid-derived sample, it can be a marker for identifying the morbidity and severity of depression.
　本群に属する精神疾患判定マーカーにおいて配列番号４４、６６、及び８８～９３で示すアミノ酸配列からなる群から選択されるタンパク質等の測定値は、うつ病の評価尺度HAM-Dと相関性を示すことから、うつ病の重症度を判定することができる。具遺体的には、前記タンパク質等は、HAM-Dを評価尺度としてピアソン相関関数においてp＜0.01で相関する、うつ病の重症度を同定することができる精神疾患判定マーカーである。 The measured values of proteins and the like selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 44, 66, and 88 to 93 in the psychiatric disease determination marker belonging to this group are correlated with the depression evaluation scale HAM-D. From this, the severity of depression can be determined. Specifically, the protein or the like is a psychiatric disease determination marker that can identify the severity of depression that correlates with p <0.01 in the Pearson correlation function using HAM-D as an evaluation scale.
（４）統合失調症／双極性障害を判定する群
　４群に属する精神疾患判定マーカーを表５に示す。 (4) Group for determining schizophrenia / bipolar disorder Table 5 shows the markers for determining mental disorders belonging to Group 4.
Figure JPOXMLDOC01-appb-T000015
Figure JPOXMLDOC01-appb-T000015
　この群に属する精神疾患判定マーカーは、統合失調症／双極性障害を判定できるマーカーである。「統合失調症／双極性障害」とは、従来の分類法でいずれかの精神疾患に分類されていた精神疾患で、バイオマーカーを用いた生物学的根拠に基づく本発明の判定方法により、統合失調症と双極性障害の両精神疾患の特徴を有し、いずれか一方には明確に分類し得ない新たな精神疾患群をいう。具体的には、図１において４群で示す区画に属するマーカーであって、配列番号９４～９８で示すアミノ酸配列からなる群から選択されるタンパク質等をいう。本群の精神疾患判定マーカーを用いれば、被験者が統合失調症／双極性障害に罹患している可能性が高いことを鑑別できる。 The psychiatric disorder determination marker belonging to this group is a marker that can determine schizophrenia / bipolar disorder. “Schizophrenia / bipolar disorder” is a psychiatric disorder that has been classified as one of the psychiatric disorders by the conventional classification method, and is integrated by the determination method of the present invention based on the biological basis using biomarkers. It refers to a new group of mental illnesses that have the characteristics of both schizophrenia and bipolar disorder and cannot be clearly classified into either one. Specifically, it refers to a protein or the like belonging to the compartments shown in Group 4 in FIG. 1 and selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 94 to 98. Use of this group of psychiatric disease determination markers can identify that a subject is likely to have schizophrenia / bipolar disorder.
　本群に属する精神疾患判定マーカーにおいて配列番号９５～９８で示すアミノ酸配列からなる群から選択されるタンパク質等は、統合失調症／双極性障害に罹患して可能性が高いことを鑑別できると共に、統合失調症／双極性障害を亜型に細分類することができる判定マーカーである。本群に属する精神疾患判定マーカーも前述の１～３群における精神疾患の亜型同定と同様にカットオフ値を設定し、その値に基づいて分類すればよい。例えば、配列番号９５で示されるBMP-1マーカーを用いた精神疾患の判定において、被験者におけるBMP-1マーカーの測定値が健常者群の測定値の95パーセンタイルより大きい値を示した場合、その被検者は、統合失調症／双極性障害に罹患している可能性が高く、かつ「統合失調症／双極性障害のBMP-1亜型」と同定することができる。 A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 95 to 98 in the psychiatric disorder determination marker belonging to this group can be identified as having a high possibility of suffering from schizophrenia / bipolar disorder, It is a determination marker that can subdivide schizophrenia / bipolar disorder into subtypes. The mental disease determination marker belonging to this group may be classified based on the cut-off value set in the same manner as the subtype identification of the mental disease in the first to third groups. For example, in the determination of psychiatric disorder using the BMP-1 marker represented by SEQ ID NO: 95, when the measured value of the BMP-1 marker in the subject is greater than the 95th percentile of the measured values of the healthy subject group, The examiner is likely to have schizophrenia / bipolar disorder and can be identified as “BMP-1 subtype of schizophrenia / bipolar disorder”.
　表５では、健常者の95パーセンタイル及び5パーセンタイルをカットオフ値とし、95パーセンタイルより大、又は5パーセンタイル未満の異常値を示すタンパク質を亜型に細分類可能な精神疾患判定マーカーとして示している。例えば、配列番号９５で示すアミノ酸配列からなるBMP-1タンパク質は、健常者の95パーセンタイルをカットオフ値とし、その値より大の場合には、統合失調症又は双極性障害であると判定することができる。一方、配列番号９６～９８で示すアミノ酸配列からなる群から選択されるタンパク質等は、健常者の5パーセンタイルをカットオフ値とし、その値未満の場合には、統合失調症／双極性障害であると判定することができる。 Table 5 shows the 95th percentile and 5th percentile of healthy subjects as cut-off values, and proteins showing abnormal values greater than 95th percentile or less than 5th percentile as psychiatric disease determination markers that can be subdivided into subtypes. For example, the BMP-1 protein consisting of the amino acid sequence represented by SEQ ID NO: 95 is determined to be schizophrenia or bipolar disorder when the 95th percentile of a healthy person is set as a cutoff value, and is larger than that value. Can do. On the other hand, a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 96 to 98 has schizophrenia / bipolar disorder when the 5th percentile of healthy subjects has a cut-off value, and is less than that value Can be determined.
　被験者が統合失調症又は双極性障害のいずれに罹患しているかを判定することのできる補足用の精神疾患判定マーカーとして、配列番号１２７～１３１、及び２４２～２４６で示されるアミノ酸配列からなる群から選択されるタンパク質等を用いてもよい。そのような補足用マーカーを表６に示す。 As a supplementary psychiatric disorder determination marker capable of determining whether a subject suffers from schizophrenia or bipolar disorder, from the group consisting of amino acid sequences represented by SEQ ID NOs: 127 to 131 and 242-246 You may use the protein etc. which are selected. Such supplemental markers are shown in Table 6.
Figure JPOXMLDOC01-appb-T000016
Figure JPOXMLDOC01-appb-T000016
　表６に示すマーカーは、健常者と各々の疾患との差は小さいため、単独では統合失調症患者、又は双極性障害患者の罹患鑑別には使用できない。しかし、患者が統合失調症又は双極性障害のどちらの精神疾患に罹患している場合、両者を鑑別することができる。例えば、統合失調症か双極性障害か判別したい場合、配列番号１２７、１２８、又は１３０で示すアミノ酸配列からなるタンパク質等は濃度がカットオフ値より低い場合に統合失調症、高い場合に双極性障害であると診断でき、配列番号１２９、１３１、及び２４２～２４６で示すアミノ酸配列からなるタンパク質等は濃度がカットオフ値より高い場合に統合失調症、低い場合に双極性障害と診断できる。それ故、本群に属する精神疾患判定マーカーで統合失調症又は双極性障害に罹患した患者に対する補助的判定用のマーカーとして有用である。なお、表６のカットオフ値は、Akobeng AKActa, 2007, Paediatr, 96:644-647に記載の算出方法に基づいて、ROC曲線を作成し,(1-感度)2+(1-特異度)2が最小になる値を求めた（表８、表１０についても同様）。 The markers shown in Table 6 cannot be used alone to distinguish between patients with schizophrenia or patients with bipolar disorder because the differences between healthy individuals and each disease are small. However, if the patient is suffering from a psychiatric disorder, either schizophrenia or bipolar disorder, the two can be differentiated. For example, when it is desired to discriminate between schizophrenia and bipolar disorder, a protein comprising the amino acid sequence represented by SEQ ID NO: 127, 128, or 130 is schizophrenia when the concentration is lower than the cut-off value, and bipolar disorder when the concentration is high. When the concentration is higher than the cut-off value, schizophrenia can be diagnosed, and bipolar disorder can be diagnosed when the concentration is lower than the cutoff value. Therefore, it is useful as a marker for auxiliary determination for patients suffering from schizophrenia or bipolar disorder, which is a psychiatric disorder determination marker belonging to this group. The cut-off values in Table 6 are calculated based on the calculation method described in Akobeng AKActa, 2007, Paediatr, 96: 644-647, and (1−sensitivity) 2 + (1−specificity) A value that minimizes 2 was obtained (the same applies to Tables 8 and 10).
（５）統合失調症／うつ病を判定する群
　５群に属する精神疾患判定マーカーを表７に示す。 (5) Group for determining schizophrenia / depression Table 7 shows the markers for determining mental disorders belonging to Group 5.
Figure JPOXMLDOC01-appb-T000017
Figure JPOXMLDOC01-appb-T000017
　この群に属する精神疾患判定マーカーは、統合失調症／うつ病を判定できるマーカーである。「統合失調症／うつ病」とは、従来の分類法でいずれかの精神疾患に分類されていた精神疾患で、バイオマーカーを用いた生物学的根拠に基づく本発明の判定方法により、統合失調症とうつ病の両精神疾患の特徴を有し、いずれか一方には明確に分類し得ない新たな精神疾患群をいう。具体的には、図１において５群で示す区画に属するマーカーであって、配列番号２、４、５５、９９～１０３、１４５、１４６、１７９、及び２３３～２４０で示すアミノ酸配列からなる群から選択されるタンパク質等をいう。本群の精神疾患判定マーカーを用いれば、被験者が統合失調症／うつ病に罹患している可能性が高いことを鑑別できる。 The psychiatric disorder determination marker belonging to this group is a marker that can determine schizophrenia / depression. “Schizophrenia / depression” is a mental disorder that has been classified as one of the mental disorders according to the conventional classification method. It refers to a new group of mental illnesses that have the characteristics of both mental disorders and depression and cannot be clearly classified into either one. Specifically, from the group consisting of the amino acid sequences shown in SEQ ID NOs: 2, 4, 55, 99 to 103, 145, 146, 179, and 233 to 240, which belong to the compartment shown by group 5 in FIG. It refers to the selected protein. Use of this group of mental illness determination markers can distinguish that a subject is likely to have schizophrenia / depression.
　本群に属する精神疾患判定マーカーにおいて配列番号２、４、１０１～１０３、１４５、１４６、１７９、及び２３３～２３９で示すアミノ酸配列からなる群から選択されるタンパク質等は、統合失調症／うつ病に罹患している可能性が高いことを鑑別できると共に、統合失調症／うつ病を亜型に細分類することができる判定マーカーである。本群に属する精神疾患判定マーカーも前述の１～３群における精神疾患の亜型同定と同様にカットオフ値を設定し、その値に基づいて分類すればよい。例えば、配列番号１０１で示されるNADPH-P450 Oxidoreductaseマーカーを用いた精神疾患の判定において、被験者におけるNADPH-P450 Oxidoreductaseマーカーの測定値が健常者群の測定値の95パーセンタイルより大きい値を示した場合、その被検者は、統合失調症／うつ病に罹患している可能性が高く、かつ「統合失調症／うつ病のNADPH-P450 Oxidoreductase亜型」と同定することができる。 Proteins selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 2, 4, 101 to 103, 145, 146, 179, and 233 to 239 in the psychiatric disorder determination marker belonging to this group are schizophrenia / depression. It is a determination marker that can distinguish a high possibility of suffering from schizophrenia and subdivide schizophrenia / depression into subtypes. The mental disease determination marker belonging to this group may be classified based on the cut-off value set in the same manner as the subtype identification of the mental disease in the first to third groups. For example, in the determination of mental illness using the NADPH-P450 Oxidoreductase marker represented by SEQ ID NO: 101, when the measured value of the NADPH-P450 Oxidoreductase marker in the subject shows a value greater than the 95th percentile of the measured values of the healthy group, The subject is likely to have schizophrenia / depression and can be identified as “NADPH-P450 Oxidoreductase subtype of schizophrenia / depression”.
　表７では、健常者の95パーセンタイル及び5パーセンタイルをカットオフ値とし、その値を外れ、95パーセンタイルより大、又は5パーセンタイル未満の異常値を示すタンパク質を亜型に細分類可能な精神疾患判定マーカーとして示している。例えば、配列番号１０１、１４５、１４６、１７９、及び２３３～２３７で示すアミノ酸配列からなるNADPH-P450 Oxidoreductaseは、健常者の95パーセンタイルをカットオフ値とし、その値より大の場合には、統合失調症／うつ病であると判定することができる。一方、配列番号１０２、１０３、１３８、及び１３９で示すアミノ酸配列からなるタンパク質等は、健常者の5パーセンタイルをカットオフ値とし、その値未満の場合には、統合失調症／うつ病であると判定することができる。 In Table 7, the 95th percentile and the 5th percentile of healthy subjects are cut off values, and the psychiatric disease determination marker that can subdivide proteins that are out of that value and show abnormal values greater than 95th percentile or less than 5th percentile into subtypes As shown. For example, NADPH-P450 Oxidoreductase consisting of the amino acid sequences represented by SEQ ID NOs: 101, 145, 146, 179, and 233 to 237 has a cut-off value of the 95th percentile of a healthy person, and if greater than that value, schizophrenia Can be determined to be symptom / depression. On the other hand, the protein consisting of the amino acid sequences represented by SEQ ID NOs: 102, 103, 138, and 139 has a cutoff value of the 5th percentile of healthy individuals, and if it is less than that value, it is schizophrenia / depression Can be determined.
　被験者が統合失調症又はうつ病のいずれに罹患しているかを鑑別することのできる補足用の精神疾患判定マーカーとして、配列番号１２４～１２６で示すアミノ酸配列からなる群から選択されるタンパク質等を用いてもよい。そのような補足用マーカーを表８に示す。 A protein or the like selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 124 to 126 is used as a supplementary mental disease determination marker that can distinguish whether a subject suffers from schizophrenia or depression. May be. Such supplemental markers are shown in Table 8.
Figure JPOXMLDOC01-appb-T000018
Figure JPOXMLDOC01-appb-T000018
　表８に示すマーカーは、健常者と各々の疾患との差が小さいため、単独では統合失調症患者、又はうつ病の罹患鑑別には使用できない。しかし、患者が統合失調症又はうつ病のいずれかに罹患している場合、両者を鑑別することができる。例えば、統合失調症かうつ病かを判別したい場合、配列番号１２４で示すアミノ酸配列からなるTriosephosphate isomeraseタンパク質等の濃度4320RFUをカットオフ値として、その値よりもより高い場合に統合失調症、低い場合にうつ病であると診断できる。それ故、本群に属する精神疾患判定マーカーで統合失調症又はうつ病に罹患した患者に対する補助的判定用のマーカーとして有用である。 The markers shown in Table 8 cannot be used alone to distinguish between patients with schizophrenia or depression because the difference between healthy individuals and each disease is small. However, if the patient suffers from either schizophrenia or depression, both can be differentiated. For example, when it is desired to discriminate between schizophrenia and depression, the concentration 4320RFU of the Triosephosphate isomerase protein consisting of the amino acid sequence represented by SEQ ID NO: 124 is used as a cut-off value, and when it is higher than that value, schizophrenia is low Diagnosis of depression. Therefore, it is useful as a marker for auxiliary determination for patients suffering from schizophrenia or depression, which is a psychiatric disorder determination marker belonging to this group.
（６）うつ病／双極性障害を判定する群
　６群に属する精神疾患判定マーカーを表９に示す。 (6) Group for determining depression / bipolar disorder Table 9 shows the markers for determining mental disorders belonging to group 6.
Figure JPOXMLDOC01-appb-T000019
Figure JPOXMLDOC01-appb-T000019
　この群に属する精神疾患判定マーカーは、うつ病／双極性障害を判定できるマーカーである。「うつ病／双極性障害」とは、従来の分類法でいずれかの精神疾患に分類されていた精神疾患で、バイオマーカーを用いた生物学的根拠に基づく本発明の判定方法により、うつ病と双極性障害の両精神疾患の特徴を有し、いずれか一方には明確に分類し得ない新たな精神疾患群をいう。具体的には、図１において６群で示す区画に属するマーカーであって、配列番号１０４～１２２及び２４１で示すアミノ酸配列からなる群から選択されるタンパク質等をいう。本群の精神疾患判定マーカーを用いれば、被験者がうつ病／双極性障害に罹患している可能性が高いことを鑑別できる。 The mental disease determination marker belonging to this group is a marker that can determine depression / bipolar disorder. “Depression / bipolar disorder” is a mental illness that has been classified as any mental illness by the conventional classification method, and is depressed by the determination method of the present invention based on a biological basis using a biomarker. A new group of psychiatric disorders that have characteristics of both psychiatric disorders of bipolar disorder and cannot be clearly classified into either one. Specifically, it refers to a protein or the like belonging to the compartments shown in group 6 in FIG. 1 and selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 104 to 122 and 241. Use of this group of mental illness determination markers can distinguish that a subject is likely to be suffering from depression / bipolar disorder.
　本群に属する精神疾患判定マーカーにおいて配列番号１０８～１２２で示すアミノ酸配列からなる群から選択されるタンパク質等は、うつ病／双極性障害に罹患している可能性が高いことを鑑別できると共に、うつ病／双極性障害を亜型に細分類することができる判定マーカーである。本群に属する精神疾患判定マーカーも前述の１～３群における精神疾患の亜型同定と同様にカットオフ値を設定し、その値に基づいて分類すればよい。例えば、例えば、配列番号１０８で示されるLYN Bマーカーを用いた精神疾患の判定において、被験者におけるLYN Bマーカーの測定値が健常者群の測定値の95パーセンタイルより大きい値を示した場合、その被検者は、うつ病／双極性障害に罹患している可能性が高く、かつ「うつ病／双極性障害のLynB亜型」と同定することができる。 A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 108 to 122 in the psychiatric disease determination marker belonging to this group can be identified as having a high possibility of suffering from depression / bipolar disorder, It is a determination marker that can subdivide depression / bipolar disorder into subtypes. The mental disease determination marker belonging to this group may be classified based on the cut-off value set in the same manner as the subtype identification of the mental disease in the first to third groups. For example, in the determination of a mental illness using the LYN B marker represented by SEQ ID NO: 108, if the measured value of the LYN B marker in the subject is greater than the 95th percentile of the measured values of the healthy subject group, the subject The examiner is likely to suffer from depression / bipolar disorder and can be identified as “LynB subtype of depression / bipolar disorder”.
　被験者がうつ病又は双極性障害のいずれに罹患しているかを鑑別することのできる補足用の精神疾患判定マーカーとして、配列番号１３１～１３６、２４７、及び２４８で示すアミノ酸配列からなる群から選択されるタンパク質等を用いてもよい。 As a supplementary psychiatric disease determination marker capable of distinguishing whether a subject suffers from depression or bipolar disorder, it is selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 131 to 136, 247, and 248. Or other proteins may be used.
Figure JPOXMLDOC01-appb-T000020
Figure JPOXMLDOC01-appb-T000020
　表１０に示すマーカーは、健常者と各々の疾患との差は小さいため、単独ではうつ病患者、又は双極性障害患者の罹患鑑別には使用できない。しかし、患者がうつ病又は双極性障害のどちらの精神疾患に罹患している場合、両者を鑑別することができる。例えば、うつ病か双極性障害か判別したい場合、配列番号１３１、１３２、１３３、１３６、２４７、又は２４８で示すアミノ酸配列からなるタンパク質等は濃度がカットオフ値より高い場合にうつ病、低い場合に双極性障害であると診断でき、配列番号１３４、又は１３５で示すアミノ酸配列からなるタンパク質等は濃度がカットオフ値より低い場合にうつ病、高い場合に双極性障害と診断できる。それ故、本群に属する精神疾患判定マーカーでうつ病又は双極性障害に罹患した患者に対する補助的判定用のマーカーとして有用である。 The markers shown in Table 10 cannot be used alone to distinguish between a depressed patient or a bipolar disorder patient because the difference between a healthy person and each disease is small. However, if the patient is suffering from mental illness, either depression or bipolar disorder, both can be differentiated. For example, when it is desired to discriminate between depression and bipolar disorder, the protein comprising the amino acid sequence represented by SEQ ID NOs: 131, 132, 133, 136, 247, or 248 is depressed when the concentration is higher than the cut-off value, and when it is low If the concentration of the protein consisting of the amino acid sequence shown by SEQ ID NO: 134 or 135 is lower than the cut-off value, it can be diagnosed as bipolar disorder. Therefore, it is useful as a marker for auxiliary determination for patients suffering from depression or bipolar disorder as a mental disease determination marker belonging to this group.
（７）統合失調症、うつ病又は双極性障害を判定する群
　７群に属する精神疾患判定マーカーを表１１に示す。 (7) Group for determining schizophrenia, depression or bipolar disorder Table 11 shows the markers for determining mental disorders belonging to Group 7.
Figure JPOXMLDOC01-appb-T000021
Figure JPOXMLDOC01-appb-T000021
　この群に属する精神疾患判定マーカーは、精神疾患を判定できるマーカーである。具体的には、図１において７群で示す区画に属するマーカーであって、配列番号１３、３４、及び１３７～１４０で示すアミノ酸配列からなる群から選択されるタンパク質等をいう。本群の精神疾患判定マーカーを用いれば、被験者が統合失調症、うつ病、双極性障害、統合失調症／双極性障害、統合失調症／うつ病、又はうつ病／双極性障害のいずれかの精神疾患に罹患している可能性が高いことを判定できる。 The mental disease determination marker belonging to this group is a marker that can determine a mental disease. Specifically, it refers to a protein or the like belonging to the compartments shown in Group 7 in FIG. 1 and selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 13, 34, and 137-140. Using this group of psychiatric markers, the subject is either schizophrenia, depression, bipolar disorder, schizophrenia / bipolar disorder, schizophrenia / depression, or depression / bipolar disorder. It can be determined that there is a high possibility of having a mental illness.
２．精神疾患判定方法
２－１．概要
　本発明の第２の態様は精神疾患判定方法である。本発明は、第１態様に記載の精神疾患判定マーカーを用いて医師を介することなく、生物学的に被験者における精神疾患の有無を判定することができる。 2. 2. Mental disease determination method 2-1. Outline | summary The 2nd aspect of this invention is a mental disease determination method. The present invention can biologically determine the presence or absence of a mental illness in a subject without using a doctor using the mental illness determination marker described in the first aspect.
２－２．判定方法
２－２－１．工程
　本判定方法は、必須の工程として測定工程及び比較判定工程を含む。また、選択工程として重症度判定工程を含む。以下、各工程について具体的に説明をする。 2-2. Determination method 2-2-1. Step This determination method includes a measurement step and a comparison determination step as essential steps. Moreover, a severity determination process is included as a selection process. Hereinafter, each step will be specifically described.
（１）測定工程
　「測定工程」とは、被験者及び健常者から採取された試料の単位量あたりに含まれる第１態様に記載の精神疾患判定マーカーを定量するため、その量を測定してその測定値を得る工程である。 (1) Measurement process The “measurement process” is a method for measuring the amount of the psychiatric disorder determination marker according to the first aspect included in a unit amount of a sample collected from a subject and a healthy person. This is a step of obtaining a measured value.
　本明細書において「健常者」とは、精神疾患を有さないヒト個体、好ましくはいずれの疾患や異常も有さない健康なヒト個体をいう。本態様で用いる健常者は、性別、年齢、身長、体重等の身体的条件や、人数は特に制限はしないが、好ましくは被験体と同性で、また年齢、身長、及び体重等の身体的条件が同一又は近似である。また、本明細書では複数の健常者からなる集団を「健常者群」とする。 As used herein, “healthy person” refers to a human individual who does not have a mental illness, preferably a healthy human individual who does not have any disease or abnormality. The healthy person used in this embodiment is not particularly limited in terms of physical conditions such as sex, age, height, weight, etc., but the number is preferably the same as that of the subject, and physical conditions such as age, height, and weight. Are the same or approximate. In the present specification, a group of a plurality of healthy persons is referred to as a “healthy person group”.
　本明細書において「試料」とは、精神疾患判定マーカーであるタンパク質又はその断片を含み得る生物学的試料をいう。例えば、体液、細胞（細胞抽出液を含む）、又はそれから回収されたタンパク質等が該当する。「体液」とは、被験者から直接採取される液体状の生体試料をいう。例えば、脳脊髄液、血液（血清、血漿及び間質液を含む）、又はリンパ液が該当する。本態様の方法において、好ましい試料は、脳脊髄液又はそれから回収されたタンパク質等である。 As used herein, “sample” refers to a biological sample that may contain a protein or fragment thereof that is a psychiatric disorder determination marker. For example, a bodily fluid, a cell (including a cell extract), or a protein recovered therefrom is applicable. “Body fluid” refers to a liquid biological sample collected directly from a subject. For example, cerebrospinal fluid, blood (including serum, plasma and interstitial fluid), or lymph. In the method of this embodiment, a preferable sample is cerebrospinal fluid or a protein recovered therefrom.
　「採取された試料」とは、前記被験者及び健常者のそれぞれから採取された試料をいう。採取方法は、既知の方法であればよく、特に限定はしない。例えば、試料が脳脊髄液の場合、腰椎穿刺により採取すればよい。腰椎穿刺は、事前に市販の局所麻酔薬を用いることで、痛みを採血以下にすることが可能であり、また無外傷性針を用いることで、副作用を軽減できることから侵襲性が比較的低く、脳脊髄液を採取する場合には好適な方法である。また、試料が体液の場合、採取は、当該分野の公知の方法に基づいて行なえばよい。例えば、血液やリンパ液であれば、公知の採血方法に従えばよい。具体的には、末梢血であれば末梢部の静脈等に注射をして採取すればよい。試料は、採取後、速やかに本発明の判定方法で使用することもできるが、採取後、直ちに氷冷し、遠心により得られた上清を超低温槽で保存したものを解凍し使用してもよい。また、必要に応じて希釈若しくは濃縮、又はヘパリンのような血液凝固阻止剤を添加することもできる。 “The sample collected” refers to a sample collected from each of the subject and the healthy subject. The collecting method is not particularly limited as long as it is a known method. For example, when the sample is cerebrospinal fluid, it may be collected by lumbar puncture. Lumbar puncture is less invasive because it can reduce pain or less by using a commercially available local anesthetic in advance, and can reduce side effects by using an atraumatic needle, This method is suitable for collecting cerebrospinal fluid. When the sample is a body fluid, the collection may be performed based on a known method in the field. For example, in the case of blood or lymph, a known blood collection method may be followed. Specifically, in the case of peripheral blood, it may be collected by injection into a peripheral vein or the like. The sample can be used immediately after collection in the determination method of the present invention, but immediately after collection, it can be ice-cooled, and the supernatant obtained by centrifugation can be thawed and used. Good. Further, if necessary, dilution or concentration, or a blood coagulation inhibitor such as heparin can be added.
　「単位量」は、容量又は重量の所定の単位であって、例えば、マイクロリットル、ミリリットル、マイクログラム、ミリグラム、グラム等が挙げられる。 The “unit amount” is a predetermined unit of volume or weight, and examples thereof include microliter, milliliter, microgram, milligram, gram and the like.
　本明細書において「測定値」とは、本工程で測定される精神疾患判定マーカーの量を示す値である。測定値は、容量又は重量のような絶対値であってもよく、また濃度、イオン強度、吸光度又は蛍光強度のような相対値であってもよい。 In the present specification, the “measured value” is a value indicating the amount of the psychiatric disorder determination marker measured in this step. The measured value may be an absolute value such as volume or weight, or a relative value such as concentration, ionic strength, absorbance or fluorescence intensity.
　本工程では、被験者由来の試料（「被験者試料」という）と健常者試料由来の試料（「健常者試料」という）のそれぞれに含まれる精神疾患判定マーカーの量を測定する。 In this step, the amount of the psychiatric disease determination marker contained in each of the sample derived from the subject (referred to as “subject sample”) and the sample derived from the healthy subject sample (referred to as “normal subject sample”) is measured.
　本方法で測定すべき精神疾患判定マーカーは、判定すべき精神疾患によって選択すればよい。例えば、被験者が健常状態ではなく、いずれかの精神疾患に罹患しているか否かを判定する場合には、第１態様に記載の７群に属する配列番号１３、３４、及び１３７～１４０で示される精神疾患判定マーカーのいずれか一以上を選択すればよい。また、被験者が統合失調症／双極性障害に罹患しているか否かを判定する場合には、第１態様に記載の４群に属する配列番号９４～９８で示される精神疾患判定マーカーのいずれか一以上を選択すればよい。被験者が統合失調症／うつ病に罹患しているか否かを判定する場合には、第１態様に記載の５群に属する配列番号２、４、７、９９～１０３、１４５、１４６、１７９、及び２３３～２４０で示される精神疾患判定マーカーのいずれか一以上を選択すればよい。被験者がうつ病／双極性障害に罹患しているか否かを判定する場合には、第１態様に記載の６群に属する配列番号１０４～１２２、及び２４１で示される精神疾患判定マーカーのいずれか一以上を選択すればよい。また、被験者が統合失調症に罹患しているか否かを判定する場合には、第１態様に記載の１群に属する配列番号１、３、５、６、８～１３、１５～２５、３０、４６、１４１～１４４、１４７～１５８、及び２４９で示される精神疾患判定マーカーのいずれか一以上を選択すればよい。被験者が双極性障害に罹患しているか否かを判定する場合には、第１態様に記載の２群に属する配列番号６１～６４、１１４、及び１６３～１７２で示される精神疾患判定マーカーのいずれか一以上を選択すればよい。そして、被験者がうつ病に罹患しているか否かを判定する場合には、第１態様に記載の３群に属する配列番号１３、１４、１７、５６、６６、７０～８７、１２３、及び１７４～１７８、１８０～２３２で示される精神疾患判定マーカーのいずれか一以上を選択すればよい。 The mental disease determination marker to be measured by this method may be selected according to the mental disease to be determined. For example, when it is determined whether or not a subject is not in a normal state and suffers from any mental illness, it is represented by SEQ ID NOS: 13, 34, and 137 to 140 belonging to Group 7 described in the first aspect. Any one or more of the psychiatric disease determination markers may be selected. Further, when determining whether or not the subject suffers from schizophrenia / bipolar disorder, any one of the mental disease determination markers represented by SEQ ID NOs: 94 to 98 belonging to Group 4 according to the first aspect. One or more may be selected. When determining whether or not a subject suffers from schizophrenia / depression, SEQ ID NOs: 2, 4, 7, 99 to 103, 145, 146, 179, belonging to Group 5 according to the first aspect, And any one or more of the psychiatric disorder determination markers represented by 233 to 240 may be selected. When determining whether or not the subject is suffering from depression / bipolar disorder, any one of the mental disease determination markers represented by SEQ ID NOs: 104 to 122 and 241 belonging to group 6 described in the first aspect One or more may be selected. Further, when determining whether or not the subject suffers from schizophrenia, SEQ ID NOs: 1, 3, 5, 6, 8 to 13, 15 to 25, 30 belonging to the group 1 described in the first aspect. 46, 141 to 144, 147 to 158, and 249 may be selected from one or more of the mental disease determination markers. When determining whether or not a subject suffers from bipolar disorder, any one of the mental disease determination markers represented by SEQ ID NOs: 61 to 64, 114, and 163 to 172 belonging to Group 2 described in the first aspect Or more than one. When determining whether or not the subject suffers from depression, SEQ ID NOs: 13, 14, 17, 56, 66, 70 to 87, 123, and 174 belonging to the three groups described in the first aspect. Any one or more of psychiatric disorder determination markers indicated by ˜178 and 180˜232 may be selected.
　精神疾患判定マーカーの測定方法は、公知のペプチドの定量方法であればよく、特に限定はしない。例えば、免疫学的検出法、アプタマー解析法、又は質量分析法が挙げられる。 The method for measuring a psychiatric disorder determination marker is not particularly limited as long as it is a known peptide quantification method. For example, immunological detection methods, aptamer analysis methods, or mass spectrometry methods can be mentioned.
　免疫学的検出法は、標的分子と特異的に結合する抗体又はその断片を用いて、本発方法の標的分子である精神疾患判定マーカーを定量する方法である。免疫学的検出法で使用する抗体は、モノクローナル抗体、ポリクローナル抗体、組換え抗体、合成抗体、及び抗体フラグメントのいずれを使用してもよい。 The immunological detection method is a method for quantifying a psychiatric disease determination marker that is a target molecule of the present method, using an antibody or a fragment thereof that specifically binds to the target molecule. The antibody used in the immunological detection method may be any of a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a synthetic antibody, and an antibody fragment.
　抗体がポリクローナル抗体又はモノクローナル抗体の場合、免疫グロブリン分子は、任意のクラス（例えば、IgG、IgE、IgM、IgA、IgD及びIgY）、又は任意のサブクラス（例えば、IgG1、IgG2、IgG3、IgG4、IgA1、IgA2）でよい。抗体は、哺乳動物及び鳥を含めたいずれの動物由来とすることができる。例えば、マウス、ラット、モルモット、ウサギ、ヤギ、ロバ、ヒツジ、ラクダ、ウマ、ニワトリ又はヒト等が挙げられる。 If the antibody is a polyclonal or monoclonal antibody, the immunoglobulin molecule can be of any class (eg, IgG, IgE, IgM, IgA, IgD and IgY), or any subclass (eg, IgG1, IgG2, IgG3, IgG4, IgA1). IgA2). The antibody can be derived from any animal including mammals and birds. Examples include mice, rats, guinea pigs, rabbits, goats, donkeys, sheep, camels, horses, chickens or humans.
　組換え抗体は、キメラ抗体、ヒト化抗体、又は多重特異性抗体を含む。キメラ抗体は、異なる動物由来の抗体のアミノ酸配列を組み合わせて作製される抗体で、ある抗体の可変領域（V領域）を他の抗体のV領域で置換した抗体である。例えば、第１態様に記載のいずれかの精神疾患判定マーカーと特異的に結合するマウスモノクローナル抗体のV領域をヒト抗体のV領域と置き換え、可変領域（V領域）がマウス由来、C領域がヒト由来となった抗体が該当する。ヒト化抗体は、ヒト以外の哺乳動物、例えば、第１態様に記載のいずれかの精神疾患判定マーカーと特異的に結合するマウス抗体の可変領域（V領域）における相補性決定領域（CDR；CDR1、CDR2及びCDR3）をヒトモノクローナル抗体のCDRとを置換したグラフト抗体である。多重特異性抗体は、多価抗体、すなわち抗原結合部位を一分子内に複数有する抗体において、それぞれの抗原結合部位が異なるエピトープと結合する抗体をいう。例えば、IgGのように2つの抗原結合部位を有する抗体であれば、それぞれの抗原結合部位が第１態様に記載の同一の又は異なる精神疾患判定マーカーと特異的に結合する二重特異性抗体（Bispecific抗体）が挙げられる。 Recombinant antibodies include chimeric antibodies, humanized antibodies, or multispecific antibodies. A chimeric antibody is an antibody prepared by combining amino acid sequences of antibodies from different animals, and is an antibody in which the variable region (V region) of one antibody is replaced with the V region of another antibody. For example, the V region of a mouse monoclonal antibody that specifically binds to any of the psychiatric disease determination markers described in the first embodiment is replaced with the V region of a human antibody, the variable region (V region) is derived from a mouse, and the C region is human. Applicable antibodies are derived. A humanized antibody is a complementarity determining region (CDR; CDR1) in a variable region (V region) of a non-human mammal, for example, a mouse antibody that specifically binds to any of the psychiatric disease determination markers described in the first aspect. , CDR2 and CDR3) are graft antibodies in which the CDR of the human monoclonal antibody is substituted. Multispecific antibody refers to a multivalent antibody, that is, an antibody that binds to different epitopes in each antigen-binding site in an antibody having a plurality of antigen-binding sites in one molecule. For example, in the case of an antibody having two antigen-binding sites such as IgG, each of the antigen-binding sites specifically binds to the same or different psychiatric disease determination marker described in the first embodiment ( Bispecific antibody).
　合成抗体は、化学的方法又は組換えDNA法を用いることによって合成した抗体をいう。例えば、適当な長さと配列を有するリンカーペプチド等を介して、特定の抗体の一以上のVL及び一以上のVHを人工的に連結させた一量体ポリペプチド分子、又はその多量体ポリペプチドが該当する。このようなポリペプチドの具体例としては、一本鎖Fv（scFv ：single chain Fragment oｆ variable region）（Pierce Catalog and Handbook, 1994-1995, Pierce Chemical Co., Rockford, IL参照）、ダイアボディ（diabody）、トリアボディ（triabody）又はテトラボディ(tetrabody)等が挙げられる。免疫グロブリン分子において、VL及びVHは、通常別々のポリペプチド鎖（軽鎖と重鎖）上に位置する。一本鎖Fvは、これら2つのポリペプチド鎖上のV領域を十分な長さの柔軟性リンカーによって連結し、1本のポリペプチド鎖に包含した構造を有する合成抗体断片である。一本鎖Fv内において両V領域は、互いに自己集合して1つの機能的な抗原結合部位を形成することができる。一本鎖Fvは、それをコードする組換えDNAを、公知技術を用いてファージゲノムに組み込み、発現させることで得ることができる。ダイアボディは、一本鎖Fvの二量体構造を基礎とした構造を有する分子である(Holliger et al., 1993, Proc. Natl. Acad. Sci. USA 90:6444-6448)。例えば、上記リンカーの長さが約12アミノ酸残基よりも短い場合、一本鎖Fv内の2つの可変部位は自己集合できないが、ダイアボディを形成させることにより、すなわち、2つの一本鎖Fvを相互作用させることにより、一方のFv鎖のVLが他方のFv鎖のVHと集合可能となり、2つの機能的な抗原結合部位を形成することができる（Marvin et al., 2005, Acta Pharmacol. Sin. 26:649-658）。さらに、一本鎖FvのC末端にシステイン残基を付加させることにより、2本のFv鎖同士のジスルフィド結合が可能となり、安定的なダイアボディを形成させることもできる(Olafsen et al., 2004, Prot. Engr. Des. Sel. 17:21-27)。このようにダイアボディは二価の抗体断片であるが、それぞれの抗原結合部位は、同一エピトープと結合する必要はなく、それぞれが異なるエピトープを認識し、特異的に結合する二重特異性を有していてもよい。トリアボディ、及びテトラボディは、ダイアボディと同様に一本鎖Fv構造を基本としたその三量体、及び四量体構造を有する。それぞれ、三価、及び四価の抗体断片であり、多重特異性抗体であってもよい。 Synthetic antibody refers to an antibody synthesized by using a chemical method or a recombinant DNA method. For example, a monomeric polypeptide molecule in which one or more VL and one or more VH of a specific antibody are artificially linked through a linker peptide having an appropriate length and sequence, or a multimeric polypeptide thereof Applicable. Specific examples of such polypeptides include single chain Fv (scFv: single chain Fragment of variable region) (Pierce Catalog and Handbook, 1994-1995, Pierce Chemical Co., Rockford, IL), diabody ), Triabody or tetrabody. In immunoglobulin molecules, VL and VH are usually located on separate polypeptide chains (light chain and heavy chain). Single-chain Fv is a synthetic antibody fragment having a structure encompassed by one polypeptide chain, in which V regions on these two polypeptide chains are connected by a flexible linker having a sufficient length. Within a single chain Fv, both V regions can self-assemble with each other to form one functional antigen binding site. Single-stranded Fv can be obtained by integrating and expressing a recombinant DNA encoding it in the phage genome using known techniques. A diabody is a molecule having a structure based on a dimeric structure of a single chain Fv (Holliger et al., 1993, Proc. Natl. Acad. Sci. USA 90: 6444-6448). For example, if the length of the linker is shorter than about 12 amino acid residues, the two variable sites within the single chain Fv cannot self-assemble, but by forming a diabody, i.e., two single chain Fv , The VL of one Fv chain can be assembled with the VH of the other Fv chain to form two functional antigen binding sites (Marvin et al., 2005, Acta Pharmacol. Sin. 26: 649-658). Furthermore, by adding a cysteine residue to the C-terminus of the single chain Fv, it becomes possible to form a disulfide bond between the two Fv chains and to form a stable diabody (Olafsen et al., 2004). , Prot. Engr. Des. Sel. 17: 21-27). Thus, a diabody is a bivalent antibody fragment, but each antigen-binding site need not bind to the same epitope, and each has a bispecificity that recognizes and specifically binds to a different epitope. You may do it. Triabodies and tetrabodies have their trimer and tetramer structures based on a single-chain Fv structure as in diabodies. These are trivalent and tetravalent antibody fragments, respectively, and may be multispecific antibodies.
　抗体フラグメントとは、例えば、Fab、F(ab’2）、Fv等が該当する。 Antibody fragments include, for example, Fab, F (ab ′ 2 ), Fv and the like.
　免疫学的検出法には、例えば、酵素免疫測定法（ELISA法、EIA法を含む）、蛍光免疫測定法、放射免疫測定法（RIA）、発光免疫測定法、表面プラズモン共鳴法（SPR法）、水晶振動子マイクロバランス（QCM）法、免疫比濁法、ラテックス凝集免疫測定法、ラテックス比濁法、赤血球凝集反応、粒子凝集反応法、金コロイド法、キャピラリー電気泳動法、ウェスタンブロット法又は免疫組織化学法（免疫染色法）が挙げられる。 Examples of immunological detection methods include enzyme immunoassay (including ELISA and EIA methods), fluorescence immunoassay, radioimmunoassay (RIA), luminescence immunoassay, surface plasmon resonance (SPR method) , Quartz crystal microbalance (QCM) method, immunoturbidimetric method, latex agglutination immunoassay method, latex turbidimetric method, erythrocyte agglutination reaction, particle agglutination reaction method, colloidal gold method, capillary electrophoresis, Western blotting or immunization Histochemical method (immunostaining method) can be mentioned.
　アプタマー解析法は、標的分子と強固、かつ特異的に結合するアプタマーを用いて、本発方法の標的分子である精神疾患判定マーカーを定量する方法である。アプタマーは、立体構造によって標的物質と強固、かつ特異的に結合し、標的物質の機能を特異的に抑制する能力を持つリガンド分子である。アプタマーは、その分子の種類により、核酸アプタマーとペプチドアプタマーに大別することができるが、いずれのアプタマーであってもよい。 The aptamer analysis method is a method for quantifying a psychiatric disorder determination marker, which is a target molecule of the present method, using an aptamer that binds firmly and specifically to a target molecule. Aptamers are ligand molecules that have the ability to bind firmly and specifically to a target substance through a three-dimensional structure and specifically inhibit the function of the target substance. Aptamers can be roughly classified into nucleic acid aptamers and peptide aptamers depending on the type of the molecule, and any aptamer may be used.
　核酸アプタマーは、核酸で構成されるアプタマーをいう。核酸アプタマーを構成する核酸は、DNA、RNA又はそれらの組合せのいずれであってもよい。必要に応じて、PNA、LNA／BNA、メチルホスホネート型DNA、ホスホロチオエート型DNA、2'-O-メチル型RNA等の化学修飾核酸を含むこともできる。 Nucleic acid aptamer refers to an aptamer composed of nucleic acids. The nucleic acid constituting the nucleic acid aptamer may be any of DNA, RNA, or a combination thereof. If necessary, a chemically modified nucleic acid such as PNA, LNA / BNA, methylphosphonate DNA, phosphorothioate DNA, 2′-O-methyl RNA may be included.
　核酸アプタマーは、第１態様に記載のいずれかの精神疾患判定マーカーを標的分子として、当該分野で公知の方法により作製することができる。例えば、RNAアプタマーであれば、SELEX(systematic evolution of ligands by exponential enrichment)法を用いて試験管内選別により作製することができる。SELEX法とは、ランダム配列領域とその両端にプライマー結合領域を有する多数のRNA分子によって構成されるRNAプールから標的分子である精神疾患判定マーカーに結合したRNA分子を選択し、回収後にRT-PCR反応によって増幅した後、得られたcDNA分子を鋳型として転写を行い、それを次のラウンドのRNAプールにするという一連のサイクルを数～数十ラウンド繰り返して、精神疾患判定マーカーに対して、より結合力の強いRNAを選択する方法である。ランダム配列領域とプライマー結合領域の塩基配列長は特に限定はしない。一般的にランダム配列領域は、20～80塩基、プライマー結合領域は、それぞれ15～40塩基の範囲である。以上の方法によって最終的に得られたRNA分子を精神疾患判定マーカー結合性RNAアプタマーとして利用する。なお、SELEX法は、公知の方法であり、具体的な方法は、例えば、Panら（Proc. Natl. Acad. Sci. 1995, U.S.A.92: 11509-11513）に準じて行えばよい。 The nucleic acid aptamer can be prepared by a method known in the art using any of the psychiatric disorder determination markers described in the first embodiment as a target molecule. For example, an RNA aptamer can be prepared by in vitro selection using the SELEX (systematic “evolution” of “ligands” by “exponential” enrichment) method. The SELEX method selects an RNA molecule bound to a psychiatric disorder marker as a target molecule from an RNA pool composed of a large number of RNA molecules having a random sequence region and primer binding regions at both ends. After amplification by reaction, transcription is performed using the obtained cDNA molecule as a template, and a series of cycles to make it into the RNA pool of the next round is repeated several to several tens of rounds. This is a method of selecting RNA having a strong binding force. The base sequence lengths of the random sequence region and the primer binding region are not particularly limited. Generally, the random sequence region is 20 to 80 bases, and the primer binding region is 15 to 40 bases. The RNA molecule finally obtained by the above method is used as a mental disease determination marker-binding RNA aptamer. The SELEX method is a known method, and a specific method may be performed according to, for example, Pan et al. (Proc. Natl. Acad. Sci. 1995, U.S.A.92: 11509-11513).
　ペプチドアプタマーとは、アミノ酸で構成されるアプタマーで、抗体と同様に、特定の標的分子の表面構造を認識して、特異的に結合する1～6kDのペプチド分子である。ファージディスプレイ法や細胞表層ディスプレイ法を用いて製造することができる。ペプチドアプタマーの製造法は、当該分野で公知の方法に基づいて作製すればよい。例えば、Whaley, S.R., et al., 2000, Nature, 405, 665-668を参照することができる。 A peptide aptamer is an aptamer composed of amino acids, and is a 1-6 kD peptide molecule that recognizes the surface structure of a specific target molecule and binds specifically like an antibody. It can be produced using a phage display method or a cell surface display method. What is necessary is just to produce the manufacturing method of a peptide aptamer based on a well-known method in the said field | area. For example, reference can be made to Whaley, S.R., et al., 2000, Nature, 405, 665-668.
　上記抗体又はアプタマーは、必要に応じて標識されていてもよい。標識は、当該分野で公知の標識物質を利用すればよい。抗体及びペプチドアプタマーの場合、例えば、蛍光色素（フルオレセイン、FITC、ローダミン、テキサスレッド、Cy3、Cy5）、蛍光タンパク質（例えば、PE、APC、GFP）、酵素（例えば、西洋ワサビペルオキシダーゼ、アルカリフォスファターゼ、グルコースオキシダーゼ）、放射性同位元素（例えば、3H、14C、35S）又はビオチン若しくは（ストレプト）アビジンにより標識することができる。また、核酸アプタマーの場合、例えば、放射性同位元素（例えば、32P、3H、14C）、DIG、ビオチン、蛍光色素（例えば、FITC、Texas、cy3、cy5、cy7、FAM、HEX、VIC、JOE、Rox、TET、Bodipy493、NBD、TAMRA）、又は発光物質（例えば、アクリジニウムエスター）が挙げられる。￥標識物質で標識された抗体やアプタマーは、標的タンパク質と結合したアプタマーを検出する際に有用なツールとなり得る。 The antibody or aptamer may be labeled as necessary. For the labeling, a labeling substance known in the art may be used. For antibodies and peptide aptamers, for example, fluorescent dyes (fluorescein, FITC, rhodamine, Texas red, Cy3, Cy5), fluorescent proteins (eg, PE, APC, GFP), enzymes (eg, horseradish peroxidase, alkaline phosphatase, glucose) Oxidase), radioisotopes (eg 3 H, 14 C, 35 S) or biotin or (strept) avidin. In the case of nucleic acid aptamers, for example, radioisotopes (eg, 32 P, 3 H, 14 C), DIG, biotin, fluorescent dyes (eg, FITC, Texas, cy3, cy5, cy7, FAM, HEX, VIC, JOE, Rox, TET, Bodipy493, NBD, TAMRA), or a luminescent substance (for example, acridinium ester). An antibody or aptamer labeled with a labeling substance can be a useful tool for detecting an aptamer bound to a target protein.
　質量分析法には、高速液体クロマトグラフ質量分析法（LC-MS）、高速液体クロマトグラフタンデム質量分析法（LC-MS/MS）、ガスクロマトグラフ質量分析法（GC-MS）、ガスクロマトグラフタンデム質量分析法（GC-MS/MS）、キャピラリー電気泳動質量分析法（CE-MS）及びICP質量分析法（ICP-MS）が挙げられる。 Mass spectrometry includes high performance liquid chromatograph mass spectrometry (LC-MS), high performance liquid chromatograph tandem mass spectrometry (LC-MS / MS), gas chromatograph mass spectrometry (GC-MS), and gas chromatograph tandem mass. Analytical methods (GC-MS / MS), capillary electrophoresis mass spectrometry (CE-MS) and ICP mass spectrometry (ICP-MS) can be mentioned.
　上記分析法は、いずれも当該分野に公知の技術であって、それらの方法に準じて行えばよい。例えば、Green, M.R. and Sambrook, J., 2012, Molecular Cloning: A Laboratory Manual Fourth Ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York；Christopher J., et al., 2005, Chemical Review,105:1103-1169；Iijima Y. et al., 2008,.The Plant Journal, 54,949-962；Hirai M. et al.,2004, Proc Natl Acad Sci USA, 101(27) 10205-10210；Sato S, et al., 2004,,The Plant Journal, 40(1)151-163； Shimizu M. et al., 2005, Proteomics, 5,3919-3931に記載の方法に準じて行うことができる。また、各メーカーから各種核酸又はペプチド定量キットが市販されており、それらを利用することもできる。 The above analysis methods are all techniques known in the art, and may be performed according to these methods. For example, Green, MR and Sambrook, J., 2012, Molecular Cloning: A Laboratory Manual Fourth Ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York; Christopher J., et alhem, 2005, Review : 1103-1169 ； Iijima Y. et al., 2008, .The Plan Journal, 54,949-962; Hirai M. et al., 2004, Proc Natl Acad Sci USA, 101 (27) 10205-10210; Sato S, et al., 2004, The The Plant Journal, (40 (1) 151-163; Shimizu M. et al., 2005, Proteomics, 5,3919-3931. In addition, various nucleic acid or peptide quantification kits are commercially available from each manufacturer, and they can also be used.
　なお、前記測定工程における健常者の測定値は、予め健常者又は健常者群から得られた試料中の各精神疾患判定マーカー量を測定しておき、それをデータベース化したものを使用してもよい。 In addition, the measurement value of the healthy person in the measurement step may be determined by measuring the amount of each psychiatric disease determination marker in a sample obtained from a healthy person or a group of healthy persons in advance, and using it as a database. Good.
　本工程では、被験者及び健常者の測定値を補正するために、単位量あたりの試料において量的差異のないことが期待される公知のタンパク質を内在性コントロールとして、さらに測定してもよい。このような内在性コントロール用タンパク質には、例えば、アルブミンが挙げられる。 In this step, in order to correct the measured values of the subject and the healthy person, a known protein expected to have no quantitative difference in the sample per unit amount may be further measured as an endogenous control. Examples of such endogenous control protein include albumin.
（２）比較判定工程
　「比較判定工程」とは、前記測定工程で得られた測定値に基づいて被験者における精神疾患の罹患可能性を判定する工程である。 (2) Comparison determination process The “comparison determination process” is a process of determining the morbidity of a mental illness in a subject based on the measurement value obtained in the measurement process.
　測定値の比較は、前記測定工程で得られた被験者の測定値と健常者若しくは健常者群の測定値間で、又は被験者の測定値と健常者群の測定値に基づいて得られたカットオフ値で、行う。この時、前記内在性コントロール用タンパク質の測定値を用いて、被験者及び健常者の測定値を補正することもできる。 The comparison of the measured values is a cut-off obtained between the measured values of the subject obtained in the measuring step and the measured values of the healthy person or the healthy person group, or based on the measured values of the subject and the healthy person group. Do by value. At this time, the measured values of the subject and healthy subjects can be corrected using the measured values of the endogenous control protein.
　判定は、被験者の測定値が健常者又は健常者群の測定値よりも有意に高い又は低い場合、又はカットオフ値により陽性に分類された場合、その被験者は、測定工程において選択した精神疾患判定マーカーが判定可能な精神疾患を有している可能性が高いと判定する。例えば、測定工程で第１態様に記載の１群に属する配列番号１～６で示されるいずれか一を精神疾患判定マーカーとして選択した場合、被験者の測定値が健常者の測定値よりも有意に高いときには、その被験者は統合失調症に罹患している可能性が高いと判定することができる。また、測定工程で第１態様に記載の１群に属する配列番号７～２５で示されるいずれか一を精神疾患判定マーカーとして選択した場合、その測定値が健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類されれば、その被験者は統合失調症に罹患している可能性が高いと判定することができる。一方、測定値が有意に高くない場合、又はカットオフ値により陰性に分類された場合、その被験者は統合失調症に罹患していない可能性が高いと判定する。 Judgment is made when the measured value of the subject is significantly higher or lower than the measured value of the healthy subject or the group of healthy subjects, or when the subject is classified as positive by the cut-off value, the subject is determined to have the mental disease selected in the measuring step. It is determined that the marker is likely to have a psychiatric disorder that can be determined. For example, when any one of SEQ ID NOs: 1 to 6 belonging to one group described in the first aspect is selected as a mental illness determination marker in the measurement step, the measurement value of the subject is significantly higher than the measurement value of the healthy subject When it is high, it can be determined that the subject is likely to have schizophrenia. In addition, when any one of SEQ ID NOs: 7 to 25 belonging to one group described in the first aspect is selected as a mental illness determination marker in the measurement step, the measurement value is determined in advance based on the measurement value of the healthy subject group. If the subject is classified as positive according to the determined cut-off value, it can be determined that the subject is likely to have schizophrenia. On the other hand, if the measured value is not significantly high, or if the measured value is classified as negative by the cut-off value, it is determined that the subject is likely not suffering from schizophrenia.
　判定は、測定工程で選択し、使用した精神疾患判定マーカーによって行う。例えば、第１態様に記載の７群に属する精神疾患判定マーカーを使用した場合、被験者の測定値が健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類された場合、その被験者は、健常状態ではなく、いずれかの精神疾患に罹患している可能性が高いと判定する。 Judgment is made by the psychiatric disease determination marker selected and used in the measurement process. For example, when using the psychiatric disease determination marker belonging to Group 7 described in the first aspect, when the measurement value of the subject is classified as positive by a predetermined cutoff value based on the measurement value of the healthy group, It is determined that the subject is not in a healthy state and is likely to have any mental illness.
　第１態様に記載の４群に属する配列番号９４で示されるPSA-ACTタンパク質を精神疾患判定マーカーとして使用した場合、被験者の測定値が健常者の測定値よりも有意に高ければ、その被験者は、統合失調症／双極性障害に罹患している可能性が高いと判定する。また、４群に属する配列番号９５～９８で示される精神疾患判定マーカーを使用した場合、健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類された場合、その被験者は、統合失調症／双極性障害に罹患している可能性が高いと判定する。 When the PSA-ACT protein represented by SEQ ID NO: 94 belonging to Group 4 described in the first embodiment is used as a psychiatric disease determination marker, if the measured value of the subject is significantly higher than the measured value of the healthy subject, the subject is Determine that they are likely to have schizophrenia / bipolar disorder. In addition, when using the psychiatric disease determination marker represented by SEQ ID NOs: 95 to 98 belonging to Group 4, when the subject is positively classified by a predetermined cutoff value based on the measurement value of the healthy group, the subject is Determine that they are likely to have schizophrenia / bipolar disorder.
　第１態様に記載の５群に属する配列番号９９及び２４０で示される精神疾患判定マーカーを使用した場合、被験者の測定値が健常者の測定値よりも有意に高ければ、その被験者は、統合失調症／うつ病に罹患している可能性が高いと判定する。また、５群に属する配列番号２、４、７、１００～１０３、１４５、１４６、１７９及び２３３～２３９で示される精神疾患判定マーカーを使用した場合、健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類された場合、その被験者は、統合失調症／うつ病に罹患している可能性が高いと判定する。 When the mental disease determination marker represented by SEQ ID NOs: 99 and 240 belonging to Group 5 described in the first aspect is used, if the measurement value of the subject is significantly higher than the measurement value of the healthy subject, the subject is schizophrenia. Determined to have a high probability of suffering from symptom / depression. In addition, when the psychiatric disease determination markers represented by SEQ ID NOs: 2, 4, 7, 100 to 103, 145, 146, 179 and 233 to 239 belonging to Group 5 are used, the markers are determined in advance based on the measured values of the healthy group. If the subject is classified as positive by the cut-off value, it is determined that the subject is likely to have schizophrenia / depression.
　第１態様に記載の６群に属する配列番号１０４～１０７、及び２４１で示される精神疾患判定マーカーを使用した場合、被験者の測定値が健常者の測定値よりも有意に高ければ、その被験者は、うつ病／双極性障害に罹患している可能性が高いと判定する。また、６群に属する配列番号１０８～１２２で示される精神疾患判定マーカーを使用した場合、健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類された場合、その被験者は、うつ病／双極性障害に罹患している可能性が高いと判定する。 When the mental disease determination marker represented by SEQ ID NOs: 104 to 107 and 241 belonging to Group 6 described in the first aspect is used, if the measurement value of the subject is significantly higher than the measurement value of the healthy subject, the subject is Determine that the person is most likely suffering from depression / bipolar disorder. In addition, when the psychiatric disease determination marker represented by SEQ ID NOs: 108 to 122 belonging to Group 6 is used, if the subject is positively classified according to a predetermined cutoff value based on the measurement value of the healthy group, the subject is Determine that the person is most likely suffering from depression / bipolar disorder.
　第１態様に記載の１群に属する配列番号１、３、５、６、３０及び１４９～１５８で示される精神疾患判定マーカーを使用した場合、前述のように被験者の測定値が健常者の測定値よりも有意に高ければ、その被験者は、統合失調症に罹患している可能性が高いと判定する。また、１群に属する配列番号７～２５、１４１～１４８、及び２４９で示される精神疾患判定マーカーを使用した場合、健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類された場合、その被験者は、統合失調症に罹患している可能性が高いと判定する。 When the mental disease determination marker represented by SEQ ID NOs: 1, 3, 5, 6, 30, and 149 to 158 belonging to one group described in the first aspect is used, the measured value of the subject is measured by a healthy person as described above. If significantly higher than the value, the subject is determined to be more likely to have schizophrenia. In addition, when the psychiatric disease determination markers represented by SEQ ID NOs: 7 to 25, 141 to 148, and 249 belonging to one group are used, they are classified as positive according to a cut-off value determined in advance based on the measurement values of the healthy group. If so, the subject is determined to be likely to have schizophrenia.
　第１態様に記載の２群に属する配列番号６１、６２、及び１７０～１７２で示される精神疾患判定マーカーを使用した場合、被験者の測定値が健常者の測定値よりも有意に高ければ、その被験者は、双極性障害に罹患している可能性が高いと判定する。また、２群に属する配列番号６３、６４、１１４、及び１６３～１６９で示される精神疾患判定マーカーを使用した場合、健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類された場合、その被験者は、双極性障害に罹患している可能性が高いと判定する。 When the mental disease determination marker represented by SEQ ID NOs: 61, 62, and 170 to 172 belonging to the two groups described in the first aspect is used, if the measured value of the subject is significantly higher than the measured value of a healthy person, The subject is determined to be more likely to have bipolar disorder. In addition, when the mental disease determination markers represented by SEQ ID NOs: 63, 64, 114, and 163 to 169 belonging to the two groups are used, the markers are positively classified according to a predetermined cutoff value based on the measurement values of the healthy group. If so, the subject is determined to be likely to have bipolar disorder.
　そして、第１態様に記載の３群に属する配列番号１３、１４、１７、５６、７０～７５、１２３、及び１９３～２３２で示される精神疾患判定マーカーを使用した場合、被験者の測定値が健常者の測定値よりも有意に高ければ、その被験者は、うつ病に罹患している可能性が高いと判定する。また、３群に属する配列番号６６、７６～８７、１７４～１７８、及び１８０～１９２で示される精神疾患判定マーカーを使用した場合、健常者群の測定値に基づいてあらかじめ定められたカットオフ値により陽性に分類された場合、その被験者は、うつ病に罹患している可能性が高いと判定する。 When the mental disease determination marker represented by SEQ ID NOs: 13, 14, 17, 56, 70 to 75, 123, and 193 to 232 belonging to the three groups described in the first aspect is used, the measured value of the subject is healthy. If it is significantly higher than the person's measurement, the subject is determined to be more likely to have depression. Further, when the mental disease determination markers represented by SEQ ID NOs: 66, 76 to 87, 174 to 178, and 180 to 192 belonging to the three groups are used, a cutoff value determined in advance based on the measured values of the healthy group If the subject is classified as positive, the subject is determined to have a high probability of suffering from depression.
（３）重症度判定工程
　「重症度判定工程」は、精神疾患の重症度を同定する場合に、選択される工程である。 (3) Severity determination step The “severity determination step” is a step selected when identifying the severity of a mental illness.
　本工程では、比較判定工程において、精神疾患を鑑別し、特定していることを前提とする。つまり、重症度判定工程とは、被験者において統合失調症、双極性障害、又はうつ病の罹患を前提として、その精神疾患の重症度をさらに同定する補足的工程である。本工程は、原則として比較判定工程後に行うが、比較判定工程と同時に行うこともできる。 In this process, it is assumed that the mental illness is identified and specified in the comparison and determination process. That is, the severity determination step is a supplementary step of further identifying the severity of the mental illness on the assumption that the subject suffers from schizophrenia, bipolar disorder, or depression. This process is performed after the comparison determination process in principle, but can be performed simultaneously with the comparison determination process.
　精神疾患の重症度を同定可能な精神疾患判定マーカー（重症度同定マーカー）は、当該マーカーの試料中の測定値が各精神疾患の所定の評価尺度と相関性を示す。したがって、各重症度同定マーカーの測定値と評価尺度との関係を示す検量線を予め作成しておけばよい。被験者における重症度同定マーカーの測定値と検量線から、その被検者における精神疾患の重症度を容易に同定することができる。例えば、精神疾患判定マーカーの測定値と評価尺度の値が正の比例関係にある場合、測定値が高ければ精神疾患の重症度が高いと判定することができる。 A mental disease determination marker (severity identification marker) that can identify the severity of a mental illness indicates that the measured value in the sample of the marker correlates with a predetermined evaluation scale of each mental illness. Therefore, a calibration curve indicating the relationship between the measured value of each severity identification marker and the evaluation scale may be created in advance. From the measured value of the severity identification marker in the subject and the calibration curve, the severity of the mental illness in the subject can be easily identified. For example, when the measured value of the mental illness determination marker and the value of the evaluation scale are in a positive proportional relationship, it can be determined that the severity of the mental illness is high if the measured value is high.
　本工程において統合失調症の重症度を同定する場合には、精神疾患判定マーカーとして配列番号９、１３、２６～２９、３１～６０、及び１５９～１６２で示すアミノ酸配列からなる群から選択されるタンパク質等を用いる。配列番号２６～３３で示すアミノ酸配列からなる精神疾患判定マーカーであれば、当該マーカーの測定値はPANSSと相関性を示す。また、配列番号９、１３、３４～６０、及び１５９～１６２で示すアミノ酸配列からなる群から選択される精神疾患判定マーカーであれば、当該マーカーの測定値はBACSと相関性を示す。また、双極性障害の重症度を同定する場合であれば、配列番号４７、６５～６９、及び１７３で示すアミノ酸配列からなる群から選択されるタンパク質等を用いる。これらの精神疾患判定マーカーの測定値は、HAM-Dと相関性を示す。さらに、うつ病の重症度を同定する場合であれば、配列番号４４、６６、及び８８～９３で示すアミノ酸配列からなる群から選択されるタンパク質等を用いる。これらの精神疾患判定マーカーの測定値は、HAM-Dと相関性を示す。 When the severity of schizophrenia is identified in this step, it is selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 9, 13, 26 to 29, 31 to 60, and 159 to 162 as a psychiatric disorder determination marker. Use protein. In the case of a psychiatric disorder determination marker consisting of the amino acid sequences represented by SEQ ID NOs: 26 to 33, the measured value of the marker correlates with PANSS. Further, in the case of a psychiatric disease determination marker selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 9, 13, 34 to 60, and 159 to 162, the measured value of the marker correlates with BACS. In the case of identifying the severity of bipolar disorder, a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 47, 65 to 69, and 173 is used. The measured values of these psychiatric disease determination markers are correlated with HAM-D. Furthermore, when identifying the severity of depression, a protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 44, 66, and 88 to 93 is used. The measured values of these psychiatric disease determination markers are correlated with HAM-D.
２－２－２．組み合わせ
　本態様の判定方法では、前記測定工程において、第１態様に記載の精神疾患判定マーカーのいずれか1つを使用すればよいが、2つ以上を組み合わせて測定することもできる。複数の精神疾患判定マーカーを組み合わせて測定することによって、精神疾患の判定精度を一層高めることができる。 2-2-2. Combination In the determination method of this aspect, any one of the mental disease determination markers described in the first aspect may be used in the measurement step, but two or more may be combined and measured. By measuring in combination with a plurality of markers for determining mental illness, the accuracy of determining mental illness can be further enhanced.
　例えば、第１態様に記載の１群、２群、４群及び７群に属する精神疾患判定マーカーを組み合わせる場合が挙げられる。７群の精神疾患判定マーカーの結果から、健常状態ではなく、統合失調症、双極性障害又はうつ病のいずれかの精神疾患に罹患している可能性が高いと判定し、また４群の精神疾患判定マーカーの結果から、その被験者は、うつ病ではなく、統合失調症又は双極性障害に罹患している可能性が高いと判定することができる。続いて、１群及び２群の精神疾患判定マーカーの結果から、その被験者は、それぞれ統合失調症、又は双極性障害に罹患している可能性が高いと判定することができる。 For example, the case where the mental disease determination markers belonging to the first group, the second group, the fourth group, and the seventh group described in the first aspect are combined may be mentioned. Based on the results of 7 groups of mental illness determination markers, it was determined that the patient is not in a healthy state and is likely to suffer from psychiatric disorders such as schizophrenia, bipolar disorder, or depression. From the results of the disease determination marker, it can be determined that the subject is more likely to suffer from schizophrenia or bipolar disorder than depression. Subsequently, from the results of the group 1 and group 2 mental illness determination markers, it can be determined that the subject is likely to suffer from schizophrenia or bipolar disorder, respectively.
　また、第１態様に記載の１群、３群、５群及び７群に属する精神疾患判定マーカーを組み合わせる場合が挙げられる。７群の精神疾患判定マーカーの結果から、健常状態ではなく、統合失調症、双極性障害又はうつ病のいずれかの精神疾患に罹患している可能性が高いと判定し、また５群の精神疾患判定マーカーの結果から、その被験者は、統合失調症又はうつ病に罹患している可能性が高いと判定することができる。続いて、１群及び３群の精神疾患判定マーカーの結果から、その被験者は、それぞれ統合失調症、又はうつ病に罹患している可能性が高いと判定することができる。 Moreover, the case where the mental disease determination markers belonging to the first group, the third group, the fifth group, and the seventh group described in the first aspect are combined is exemplified. From the results of the 7 groups of mental illness determination markers, it was determined that the patient is not in a healthy state and is likely to suffer from psychiatric disorders such as schizophrenia, bipolar disorder, or depression. From the result of the disease determination marker, it can be determined that the subject is likely to have schizophrenia or depression. Subsequently, from the results of the group 1 and group 3 mental disease determination markers, it can be determined that the subject has a high possibility of suffering from schizophrenia or depression, respectively.
　さらに、第１態様に記載の２群、３群、６群及び７群に属する精神疾患判定マーカーを組み合わせる場合が挙げられる。７群の精神疾患判定マーカーの結果から、健常状態ではなく、統合失調症、双極性障害又はうつ病のいずれかの精神疾患に罹患している可能性が高いと判定し、また６群の精神疾患判定マーカーの結果から、その被験者は、うつ病又は双極性障害に罹患している可能性が高いと判定することができる。続いて、２群及び３群の精神疾患判定マーカーの結果から、その被験者は、それぞれ双極性障害、又はうつ病に罹患している可能性が高いと判定することができる。 Furthermore, there may be mentioned a case where the mental disease determination markers belonging to the second group, the third group, the sixth group and the seventh group described in the first aspect are combined. Based on the results of the 7 groups of mental illness determination markers, it is determined that the patient is not in a healthy state and is likely to suffer from psychiatric disorder such as schizophrenia, bipolar disorder, or depression. From the result of the disease determination marker, it can be determined that the subject is highly likely to suffer from depression or bipolar disorder. Subsequently, it can be determined from the results of the mental disorder determination markers in groups 2 and 3 that the subject is highly likely to suffer from bipolar disorder or depression, respectively.
　精神疾患判定マーカーの組み合わせは、同一群内からであってもよい。例えば、１群から配列番号２７のIntegrin α1及び配列番号２８のIntegrin β1を選択する場合が挙げられる。これらは生体内で互いにヘテロ二量体を形成することから、組み合わせによってそれぞれの量を測定することで測定精度を高くすることができる。 The combination of psychiatric disorder determination markers may be from the same group. For example, there is a case where Integrin1α1 of SEQ ID NO: 27 and Integrin β1 of SEQ ID NO: 28 are selected from one group. Since these form heterodimers with each other in vivo, the measurement accuracy can be increased by measuring the respective amounts in combination.
＜精神疾患判定マーカーの単離（１）＞
１．検体の収集と試料の採取
　本発明の精神疾患判定マーカーを選択するための検体の提供者は、国立精神・神経医療研究センター（東京、日本）のホームページ、フリーペーパー上の広告、又はセンター病院内に掲示した募集ポスター等で募集した。その結果、300検体以上の提供者が得られた。表１２にその一部である統合失調症患者30例、うつ病患者30例、及び双極性障害患者16例、並びに対照としての健常者30例についての検体コホート情報を示す。 <Isolation marker for mental illness (1)>
1. Sample collection and sample collection The sample provider for selecting the psychiatric disorder determination marker of the present invention can be obtained from the National Psychiatric and Neurological Research Center (Tokyo, Japan) website, free paper advertisement, or in the center hospital. Recruited with recruitment posters posted on. As a result, more than 300 donors were obtained. Table 12 shows sample cohort information for 30 schizophrenia patients, 30 depression patients, 16 bipolar disorder patients, and 30 healthy individuals as controls.
Figure JPOXMLDOC01-appb-T000022
Figure JPOXMLDOC01-appb-T000022
　表中、Nは各精神疾患及び健常者の総数を、DFは、薬剤を服用していない患者（薬剤フリー）の数を示す。これら106人の脳脊髄液を本実施例における検体とした。患者は、二人以上の精神科医による面接と病歴からDSM-IV（Diagnostic and Statistical Manual of Mental Disorders 4th Ed.）に準拠した診断がなされた。健常者は、精神疾患簡易構造化面接法(MINI)にて精神疾患を除外した。被験者には、研究目的や検体の流れ、副作用の頻度などについて説明し、書面にて同意を得た。研究は国立精神・神経医療研究センター倫理委員会の承認が得られている。なお、被験者はいずれも血縁関係のない日本人である。 In the table, N indicates the total number of each mental illness and healthy person, and DF indicates the number of patients not taking the drug (drug free). These 106 cerebrospinal fluids were used as specimens in this example. Patients were diagnosed in accordance with DSM-IV (Diagnostic and Statistical, Manual, of Mental, Disorders, 4th, Ed.) From interviews and medical history of two or more psychiatrists. Healthy subjects excluded mental illnesses using the simplified structured interview method (MINI). The subjects explained the purpose of the study, the flow of specimens, the frequency of side effects, etc., and consent was obtained in writing. The study has been approved by the National Center for Psychiatry and Neurology Research Ethics Committee. The subjects are all unrelated Japanese.
　被験者からの脳脊髄液の採取方法は、従来法に準じて行った。まず、眼底検査を含む神経学的検査の後、第3～4腰椎間乃至第4～5腰椎間に局所麻酔を行い、無外傷性針（22G、ユニシス社、UNIEVER穿刺針）にて腰椎穿刺を行い、約10mLの脳脊髄液を採取した。採取開始後、最初の約2mLを一般検体検査用（細胞数、総タンパク質、糖等）に使用し、次の約8mLを精神疾患判定マーカー単離用として低タンパク質吸着チューブ（住友ベークライト、プロテオセーブ15mL）に採取した。採取後、脳脊髄液を直ちに氷冷し、4000gで、15分間、4℃で遠心した。その後、上清を0.5mLずつ低タンパク質吸着チューブに分注し、使用するまで-80度の超低温槽で保存した。 The method of collecting cerebrospinal fluid from the subject was performed according to the conventional method. First, after neurological examinations including fundus examination, local anesthesia is performed between the 3rd and 4th lumbar vertebrae and the 4th and 5th lumbar vertebrae, and lumbar puncture is performed with an atraumatic needle (22G, Unisys, UNIEVER puncture needle) And about 10 mL of cerebrospinal fluid was collected. After the start of collection, the first approximately 2mL is used for general specimen testing (cell count, total protein, sugar, etc.), and the next approximately 8mL is used for isolating a psychiatric disorder marker (Sumitomo Bakelite, Proteo Save). 15 mL). After collection, the cerebrospinal fluid was immediately ice-cooled and centrifuged at 4000 g for 15 minutes at 4 ° C. Thereafter, 0.5 mL of the supernatant was dispensed into a low protein adsorption tube and stored in a -80 degree ultra low temperature tank until use.
２．脳脊髄液中のタンパク質の解析
　脳脊髄液中のタンパク質の解析は、SomaLogic社（USA）の提供するアプタマー解析サービス、SOMAscan premiumにより行った（Lollo et al., 2014, Proteomics doi: 10.1002/pmic.201300187. 又はhttp://www.somalogic.com/）。本解析では、1129種の改変型の核酸アプタマーSOMAmerを用いて、マイクロアレイにより脳脊髄液中のタンパク質と結合したSOMAmerを測定し、各々のタンパク質の定量を行った。同解析はダイナミックレンジの中央値が4.2 log、%CV（変動係数）の中央値が5.1%（上掲論文）と報告されている。本発明者らが行ったELISAによる比較解析でもR2 = 0.60と高い相関が得られた（図示せず）。なお、統計解析は、SPSS(IBM, Ver 20)を用いて行った。 2. Analysis of protein in cerebrospinal fluid Analysis of protein in cerebrospinal fluid was performed by SOMAscan premium, an aptamer analysis service provided by SomaLogic (USA) (Lollo et al., 2014, Proteomics doi: 10.1002 / pmic. 201300187. or http://www.somalogic.com/). In this analysis, 1129 kinds of modified nucleic acid aptamer SOMAmers were used to measure SOMAmers bound to proteins in cerebrospinal fluid using a microarray, and each protein was quantified. The analysis reports a median dynamic range of 4.2 logs and a median% CV (coefficient of variation) of 5.1% (the above paper). In the comparative analysis by ELISA conducted by the present inventors, a high correlation of R 2 = 0.60 was obtained (not shown). Statistical analysis was performed using SPSS (IBM, Ver 20).
３．結果
　1129種の各タンパク質の測定結果と診断及び症状評価との関連を解析した。その結果、健常者群の5～95パーセンタイルを正常値としたとき、各疾患群で20%以上の症例で異常値を示したタンパク質が延べ250以上、各患者群と健常者群との間に有意差のあるタンパク質が延べ300以上、それぞれの重症度と相関するタンパク質が延べ150以上得られた。その中で、特に強い有意差が認められ、バイオマーカーとしての有用性が極めて高い140種を選択した。それらのタンパク質を表１に示す。各タンパク質をその属性について分類したものが、本明細書において表２～５、７、９及び１１に示した１～７群である。 3. Results The relationship between the measurement results of 1129 proteins and the diagnosis and symptom evaluation was analyzed. As a result, when the 5th to 95th percentile of the healthy group was regarded as a normal value, a total of 250 or more proteins that showed abnormal values in 20% or more cases in each disease group, between each patient group and the healthy group A total of more than 300 proteins with significant differences and more than 150 proteins correlated with their severity were obtained. Among them, 140 species that were found to have particularly strong significant differences and extremely useful as biomarkers were selected. These proteins are shown in Table 1. Each protein is classified according to its attributes into groups 1 to 7 shown in Tables 2 to 5, 7, 9 and 11 in this specification.
　図２は、３群に属する配列番号７０で示す精神疾患判定マーカーの一つSCFsRの測定値分布を示す図である。うつ病患者群は、健常者群と比べて脳脊髄液中のSCFsRが有意に減少していた(性・年齢を考慮したANCOVAでp=0.013)。この有意差は、薬剤フリー（DF)でも顕著に認められた(p<0.001)。一方、統合失調症患者群や双極性障害患者群では、そのような有意差は認められなかった。したがって、３群に属するタンパク質は、うつ病を判定する精神疾患判定マーカーとなり得る。これと同様の結果は、１群に属する統合失調症用の精神疾患判定マーカー及び２群に属する双極性障害用の精神疾患判定マーカーについても見られた。 FIG. 2 is a diagram showing a measured value distribution of one of the mental disease determination markers SCFsR represented by SEQ ID NO: 70 belonging to Group 3. In the depression patient group, SCFsR in cerebrospinal fluid was significantly reduced compared with the healthy group (p = 0.013 in ANCOVA considering gender and age). This significant difference was also noticeable in drug-free (DF) (p <0.001). On the other hand, such a significant difference was not recognized in the schizophrenia patient group and the bipolar disorder patient group. Therefore, proteins belonging to Group 3 can serve as a psychiatric disease determination marker for determining depression. Similar results were observed for the psychiatric disorder determination marker for schizophrenia belonging to Group 1 and the mental illness determination marker for bipolar disorder belonging to Group 2.
　図３は、１群に属する配列番号７で示す統合失調症用の精神疾患判定マーカーの中で、統合失調症を亜型に細分類することのできるUNC5H3の測定値分布を示す図である。この図では、健常者群における95パーセンタイルをカットオフ値とし、95パーセンタイル以上を陽性とした。95パーセンタイル以上の異常値を示した30例中6例の統合失調症患者は、統合失調症のUNC5H3亜型として細分類することができる。なお、6例中4例は、薬剤フリー検体であり、治療薬による影響の可能性は低いことが示された。同様の結果は、２群に属する双極性障害用の精神疾患判定マーカーや第３群に属するうつ病用の精神疾患判定マーカーについても見られた。 FIG. 3 is a diagram showing a measured value distribution of UNC5H3 that can subdivide schizophrenia into subtypes among the psychiatric psychiatric disorder determination markers indicated by SEQ ID NO: 7 belonging to Group 1. In this figure, the 95th percentile in the healthy group was defined as the cut-off value, and the 95th percentile or greater was defined as positive. Six of 30 patients with schizophrenia who showed an abnormal value of 95th percentile or higher can be subdivided as UNC5H3 subtype of schizophrenia. In addition, 4 out of 6 cases were drug-free specimens, and it was shown that the possibility of the influence of the therapeutic agent is low. Similar results were observed for the mental disorder determination marker for bipolar disorder belonging to Group 2 and the mental disorder determination marker for depression belonging to Group 3.
　図４は、１群に属する配列番号２９で示す統合失調症用の精神疾患判定マーカーあるST4S6による統合失調症の測定値と評価尺度PANSSによる評価値との関係を示す図である。この図で示すようにST4S6濃度と症状評価尺度であるPANSSとの間には相関があり、ST4S6濃度が高いほど、症状が重かった。したがって、ST4S6を精神疾患判定マーカーに用いることで、統合失調症の有無とその重症度を判定することができる。これと同様の特徴を示す精神疾患判定マーカーが、２群に属する双極性障害用の精神疾患判定マーカーや３群に属するうつ病用の精神疾患判定マーカーの一部についても確認された。 FIG. 4 is a diagram showing the relationship between the measured value of schizophrenia by ST4S6 which is a psychiatric psychiatric disorder determination marker indicated by SEQ ID NO: 29 belonging to group 1 and the evaluation value by the evaluation scale PANSS. As shown in this figure, there is a correlation between ST4S6 concentration and PANSS, which is a symptom evaluation scale, and the higher the ST4S6 concentration, the more severe the symptoms. Therefore, the presence and severity of schizophrenia can be determined by using ST4S6 as a psychiatric disorder determination marker. Mental illness determination markers having the same characteristics as above were confirmed for some of the mental illness determination markers for bipolar disorder belonging to Group 2 and some of the mental illness determination markers for depression belonging to Group 3.
　図５は、６群に属する配列番号１０８で示す精神疾患判定マーカーの一つLYN Bの測定値分布を示す図である。うつ病患者群と双極性障害患者群は、健常者群と比べてLYN Bが有意に増加していた(性・年齢を考慮したANCOVAでp<0.01)。一方、統合失調症患者群では、そのような有意差は認められなかった。したがって、６群に属するタンパク質は、うつ病／双極性障害を判定する精神疾患判定マーカーとなり得る。これと同様の結果は、４群に属する統合失調症／双極性障害用の精神疾患判定マーカー及び５群に属する統合失調症／うつ病用の精神疾患判定マーカーについても見られた。 FIG. 5 is a diagram showing a distribution of measured values of LYN B, one of the psychiatric disorder determination markers indicated by SEQ ID NO: 108 belonging to Group 6. In the depression patient group and the bipolar disorder patient group, LYN B was significantly increased compared with the healthy group (ANCOVA considering gender and age, p <0.01). On the other hand, such a significant difference was not recognized in the schizophrenic patient group. Therefore, proteins belonging to Group 6 can be a psychiatric disease determination marker for determining depression / bipolar disorder. Similar results were observed for the psychiatric disease determination marker for schizophrenia / bipolar disorder belonging to Group 4 and the psychiatric disease determination marker for schizophrenia / depression belonging to Group 5.
　図６は、図５に示したLYN Bの測定値分布図にカットオフ値を記入した図である。カットオフ値は、健常者群における95パーセンタイルとした。うつ病患者30例中7例及び双極性障害16例中6例でLYN Bがカットオフ値以上の陽性であった。LYN Bのような６群に属する精神疾患判定マーカーの一部は、カットオフ値を設定することにより、うつ病／双極性障害の亜型を判定する精神疾患判定マーカーとなり得、またそのような精神疾患判定マーカーを使用することで、うつ病／双極性障害を細分類することも可能なことが示された。同様の結果は、４群に属する統合失調症／双極性障害用の精神疾患判定マーカーや５群に属する統合失調症／うつ病用の精神疾患判定マーカーについても見られた。 FIG. 6 is a diagram in which cut-off values are entered in the measured value distribution diagram of LYN B shown in FIG. The cutoff value was the 95th percentile in the healthy group. LYN B was more than the cut-off value in 7 of 30 patients with depression and 6 of 16 patients with bipolar disorder. Some psychiatric disease markers belonging to group 6 such as LYN B can be used as psychiatric disease markers for determining depression / bipolar disorder subtypes by setting a cut-off value. It was shown that depression / bipolar disorder can be subdivided by using a psychiatric disorder determination marker. Similar results were also found for schizophrenia / bipolar disorder psychiatric disease markers belonging to group 4 and schizophrenia / depressive psychiatric disease markers belonging to group 5.
　図７は、統合失調症とうつ病のいずれに罹患しているかを判定する精神疾患判定マーカーPTHの測定値分布を示す図である。この図で示すように、脳脊髄液中のPTHは、うつ病患者群と比べて統合失調症患者群で有意に減少していた(性・年齢を考慮したANCOVAでp<0.002)。統合失調症群を薬剤フリー例に限定しても、うつ病群と比べて有意に減少していた(p<0.05)。一方、脳脊髄液中のPTHは、統合失調症患者群もうつ病患者群も健常者群に対しては有意差が認められなかった。したがって、このマーカーは、例えば、統合失調症又はうつ病と判定された被験者に対して、統合失調症とうつ病のいずれであるかを適切に判定する得る補助的精神疾患判定マーカーとなり得る。 FIG. 7 is a diagram showing a distribution of measured values of a psychiatric disorder determination marker PTH for determining whether the patient suffers from schizophrenia or depression. As shown in this figure, cerebrospinal fluid PTH was significantly decreased in the schizophrenic patient group compared to the depression patient group (p <0.002 in ANCOVA considering gender and age). Even if the schizophrenia group was limited to drug-free cases, it was significantly reduced compared to the depression group (p <0.05). On the other hand, PTH in cerebrospinal fluid was not significantly different from the group of healthy subjects in the schizophrenia patient group and the depression patient group. Therefore, this marker can be an auxiliary psychiatric disease determination marker that can appropriately determine whether it is schizophrenia or depression for a subject determined to be schizophrenia or depression, for example.
　図８は、７群に属する精神疾患判定マーカーあるVEGF sR3の測定値分布を示す図である。カットオフ値は、健常者群における95パーセンタイルとした。統合失調症の30例中10例、うつ病の30例中8例、及び双極性障害16例中3例でVEGF sR3が陽性（カットオフ値以上）であった。このマーカーを用いれば被験者が健常状態か精神疾患かを判定することができる。 FIG. 8 is a diagram showing the measured value distribution of VEGF sR3, which is a psychiatric disorder determination marker belonging to Group 7. The cutoff value was the 95th percentile in the healthy group. 10 out of 30 cases of schizophrenia, 8 out of 30 cases of depression, and 3 out of 16 cases of bipolar disorder were positive (above cut-off value). If this marker is used, it can be judged whether a test subject is in a healthy state or a mental illness.
＜精神疾患判定マーカーの単離（２）＞
１．検体の収集と試料の採取
　国立精神・神経医療研究センターで募集した前述の300以上の検体から、表１２に示した検体コホートとは異なる表１３に示す検体コホートを用いて、実施例１と同じ手順で試料の採取を行った。 <Isolation marker for mental illness (2)>
1. Sample Collection and Sample Collection Same as Example 1 using the sample cohort shown in Table 13, which is different from the sample cohort shown in Table 12, from the above 300 or more samples recruited at the National Center for Neurology and Psychiatry Samples were collected according to the procedure.
Figure JPOXMLDOC01-appb-T000023
Figure JPOXMLDOC01-appb-T000023
２．脳脊髄液中のタンパク質の解析
　脳脊髄液中のタンパク質の解析も実施例１と同様に行った。 2. Analysis of protein in cerebrospinal fluid Analysis of protein in cerebrospinal fluid was performed in the same manner as in Example 1.
３．結果
　1129種の各タンパク質の測定結果と診断及び症状評価との関連を解析した結果、バイオマーカーとしての有用性が極めて高い138種を選択した。このうち、実施例１で得たバイオマーカーと重複するタンパク質が29種存在したことから、新たなバイオマーカーとして109種（配列番号１４１～２４９）が得られた。これらの新たに得られたタンパク質についても表１に示す。各タンパク質を実施例１と同様にその属性について分類したものが、本明細書において表２～５、７、９及び１１に示した１～７群である。 3. Results As a result of analyzing the relationship between the measurement results of 1129 kinds of proteins and the diagnosis and symptom evaluation, 138 kinds having extremely high usefulness as biomarkers were selected. Among these, 29 proteins (SEQ ID NOs: 141 to 249) were obtained as new biomarkers because there were 29 proteins overlapping with the biomarkers obtained in Example 1. These newly obtained proteins are also shown in Table 1. In the present specification, each protein is classified into groups 1 to 7 shown in Tables 2 to 5, 7, 9, and 11 in the same manner as in Example 1.
＜精神疾患判定マーカーによる判定制度の検証＞
　実施例１及び実施例２で得られた各精神疾患判定マーカーが、異なる検体コホートについて精神疾患判定を行った場合にも再現性がみられることを検証した。 <Verification of judgment system by mental disease judgment marker>
Each psychiatric disease determination marker obtained in Example 1 and Example 2 was verified to be reproducible even when psychiatric disease determination was performed for different sample cohorts.
　本発明の249種の精神疾患判定マーカーのうち、任意に40種を選択し、前述の表１２に示す第１検体コホート、及び表１３に示す第２検体コホートのそれぞれについて、実施例１と同様の方法で独立に解析し、測定分布を検証した。さらに第１検体コホートの測定結果と第２検体コホートの測定結果をZ変換により結合し、その分布を検証した。 Of the 249 psychiatric disorder determination markers of the present invention, 40 are arbitrarily selected, and each of the first sample cohort shown in Table 12 and the second sample cohort shown in Table 13 is the same as in Example 1. The analysis was independently performed by the method described above to verify the measurement distribution. Furthermore, the measurement result of the first sample cohort and the measurement result of the second sample cohort were combined by Z conversion, and the distribution was verified.
　図９～４８に結果を示す。 Figures 9 to 48 show the results.
　図９は、第１検体コホートから選択されたうつ病の精神疾患判定マーカーであるFibrinogenについて検証している。（ａ）は第２検体コホートにおける各精神疾患患者の髄液中におけるFibrinogen値の測定分布である。第２検体コホートでも、うつ病患者（MDD）でFibrinogen値が顕著に亢進しており、Fibrinogenのうつ病の精神疾患判定マーカーとしての再現性が確認された。また、（ｂ）からSOMAscanによるFibrinogen測定値はELISAによる測定値と、よく相関していることが立証された。（ｃ）は、第１検体コホートの測定結果と第２検体コホートの測定結果をZ変換により結合した結果である。この図でもFibrinogenは、うつ病患者（MDD)で顕著な亢進が認められた。したがって、脳脊髄液中のFibrinogenの測定により、うつ病の亜型を判定できる。 FIG. 9 verifies Fibrinogen, a mental illness determination marker for depression selected from the first sample cohort. (A) is a measurement distribution of fibrinogen values in the cerebrospinal fluid of each psychiatric patient in the second specimen cohort. In the second sample cohort, fibrinogen levels were significantly increased in depressed patients (MDD), confirming the reproducibility of fibrinogen as a psychiatric disease marker for depression. In addition, from (b), it was proved that the fibrinogen measurement value by SOMAscan correlated well with the measurement value by ELISA. (C) is the result of combining the measurement results of the first sample cohort and the measurement results of the second sample cohort by Z conversion. In this figure, fibrinogen was also markedly elevated in depressed patients (MDD). Therefore, depression subtypes can be determined by measuring fibrinogen in cerebrospinal fluid.
　図１０は、統合失調症用の精神疾患判定マーカーであるPIGFによる統合失調症検体の測定値と統合失調症認知機能簡易評価尺度(BACS)の符号課題の評価値との関係を示す図である。（ａ）で示すように、PIGF濃度とBACS符号課題の評価値との間には相関があり、PIGF濃度が高いほど、符号課題の評価値が低い、すなわち症状が重かった。また、PIGFは統合失調症群で高い傾向が見られた。したがって、PIGFを精神疾患判定マーカーに用いることで、統合失調症に罹患している可能性とその重症度を判定することができる。 FIG. 10 is a diagram showing a relationship between a measured value of a schizophrenia sample by PIGF which is a psychiatric disorder determination marker for schizophrenia and an evaluation value of a sign task of a schizophrenia cognitive function simple evaluation scale (BACS). . As shown in (a), there is a correlation between the PIGF concentration and the evaluation value of the BACS code task. The higher the PIGF concentration, the lower the evaluation value of the code task, that is, the more severe the symptoms. PIGF showed a high tendency in the schizophrenia group. Therefore, by using PIGF as a psychiatric disorder determination marker, it is possible to determine the possibility and severity of schizophrenia.
　図１１～４８は、各精神疾患判定マーカーの測定分布図で、（ａ）は第１検体コホートにおける測定結果、（ｂ）は第２検体コホートにおける測定結果、（ｃ）は（ａ）と（ｂ）の測定結果をZ変換により結合した結果を示している。各図の精神疾患判定マーカー名については、上記「図面の簡単な説明」において記述した。　いずれの図においても（ａ）～（ｃ）において、マーカーの対象疾患となる患者群では顕著な差が認められ、精神疾患判定マーカーとしての検出精度の再現性を確認することができた。この結果から、本発明の精神疾患判定マーカーは、任意の検体に対して精神疾患の判定を補助する有効な精神疾患マーカーであることが示された。 FIGS. 11 to 48 are measurement distribution diagrams of each psychiatric disorder determination marker, where (a) shows the measurement results in the first sample cohort, (b) shows the measurement results in the second sample cohort, and (c) shows (a) and ( The result of combining the measurement results of b) by Z conversion is shown. The name of the psychiatric disorder determination marker in each figure was described in the above “Simple Description of Drawing”. In any of the figures, in (a) to (c), a significant difference was observed in the patient group as the target disease of the marker, and the reproducibility of detection accuracy as a psychiatric disease determination marker could be confirmed. From this result, it was shown that the mental illness determination marker of the present invention is an effective mental illness marker for assisting the determination of mental illness for any specimen.
　本明細書で引用した全ての刊行物、特許及び特許出願をそのまま参考として本明細書にとり入れるものとする。 All publications, patents and patent applications cited in this specification shall be incorporated into this specification as they are.

Claims (36)

  1. 　脳脊髄液中に含まれる配列番号１～２４９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる精神疾患判定マーカー。 A psychiatric disease determination marker comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 1 to 249 contained in cerebrospinal fluid, a variant thereof, or a fragment thereof.
  2. 　配列番号１、３、５、６、７、８～１３、１５～３３、３５～５４、５６～６０、１４１～１４４、１４７～１６２、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症である、請求項１に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 1, 3, 5, 6, 7, 8 to 13, 15 to 33, 35 to 54, 56 to 60, 141 to 144, 147 to 162, and 249 The psychiatric disorder determination marker according to claim 1, wherein the psychiatric disorder is schizophrenia.
  3. 　統合失調症を亜型に細分類する、配列番号７～１３、１５～２５、１４１～１４４、１４７、１４８、及び２４９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、請求項２に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 7-13, 15-25, 141-144, 147, 148, and 249, or a variant thereof, which subdivides schizophrenia into subtypes The psychiatric disorder determination marker according to claim 2, comprising a fragment.
  4. 　統合失調症の重症度を同定する、配列番号９、１３、２６～２９、３１～３３、３５～５４、５６～６０、及び１５９～１６２で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、請求項２に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 9, 13, 26-29, 31-33, 35-54, 56-60, and 159-162, which identifies the severity of schizophrenia, The psychiatric disorder determination marker according to claim 2, comprising a mutant or a fragment thereof.
  5. 　配列番号４７、６１～６９、１１４、及び１６３～１７３で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が双極性障害である、請求項１に記載の精神疾患判定マーカー。 The protein according to claim 1, comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 47, 61 to 69, 114, and 163 to 173, a variant thereof, or a fragment thereof, wherein the mental illness is bipolar disorder. The psychiatric disorder determination marker described.
  6. 　双極性障害を亜型に細分類する、配列番号６３、６４、１１４、及び１６３～１６９で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、請求項５に記載の精神疾患判定マーカー。 The protein according to claim 5, consisting of a protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 63, 64, 114, and 163-169, a variant thereof, or a fragment thereof, which subdivides bipolar disorder into subtypes. The psychiatric disorder determination marker described.
  7. 　双極性障害の重症度を同定する、配列番号４７、６５～６９、及び１７３で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、請求項５に記載の精神疾患判定マーカー。 The spirit according to claim 5, consisting of a protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 47, 65 to 69, and 173, a variant thereof, or a fragment thereof, which identifies the severity of bipolar disorder Disease determination marker.
  8. 　配列番号１３、１４、１７、４４、５６、６６、７０～９３、１２３、１７４～１７８、及び１８０～２３２で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患がうつ病である、請求項１に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 13, 14, 17, 44, 56, 66, 70 to 93, 123, 174 to 178, and 180 to 232, a variant thereof, or a fragment thereof The psychiatric disorder determination marker according to claim 1, wherein the psychiatric disorder is depression.
  9. 　うつ病を亜型に細分類する、配列番号６６、７６～８７、１７４～１７８、及び１８０～１９２で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、請求項８に記載の精神疾患判定マーカー。 Subclassifying depression into subtypes, consisting of a protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 66, 76-87, 174-178, and 180-192, variants thereof, or fragments thereof, Item 9. A psychiatric disorder determination marker according to Item 8.
  10. 　うつ病の重症度を同定する、配列番号４４、６６、及び８８～９３で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなる、請求項８に記載の精神疾患判定マーカー。 The psychiatric disorder according to claim 8, comprising a protein selected from the group consisting of the amino acid sequences shown in SEQ ID NOs: 44, 66, and 88 to 93, a variant thereof, or a fragment thereof, which identifies the severity of depression Judgment marker.
  11. 　配列番号９４～９８で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症／双極性障害である、請求項１に記載の精神疾患判定マーカー。 The psychiatric disorder determination according to claim 1, comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 94 to 98, a variant thereof, or a fragment thereof, and the psychiatric disorder is schizophrenia / bipolar disorder. marker.
  12. 　配列番号２、４、５５、９９～１０３、１４５、１４６、１７９、及び２３３～２４０で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症／うつ病である、請求項１に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 2, 4, 55, 99 to 103, 145, 146, 179, and 233 to 240, a variant thereof, or a fragment thereof, and psychiatric disorder is schizophrenia The psychiatric disorder determination marker according to claim 1, which is symptom / depression.
  13. 　配列番号１０４～１２２、及び２４１で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患がうつ病／双極性障害である、請求項１に記載の精神疾患判定マーカー。 The psychiatric disorder according to claim 1, which comprises a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 104 to 122 and 241; a variant thereof; or a fragment thereof; and the mental illness is depression / bipolar disorder. Disease determination marker.
  14. 　配列番号１２４～１２６で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症及びうつ病のいずれであるかを判定する、請求項１に記載の精神疾患判定マーカー。 The protein comprising a protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 124 to 126, a variant thereof, or a fragment thereof, and determining whether the mental illness is schizophrenia or depression The psychiatric disorder determination marker described.
  15. 　配列番号１２７～１３１、及び２４２～２４６で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症及び双極性障害のいずれであるかを判定する、請求項１に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 127 to 131 and 242 to 246, a variant thereof, or a fragment thereof, and determines whether the mental disorder is schizophrenia or bipolar disorder The psychiatric disorder determination marker according to claim 1.
  16. 　配列番号１３１～１３６、２４７、及び２４８で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患がうつ病及び双極性障害のいずれであるかを判定する、請求項１に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 131 to 136, 247, and 248, a variant thereof, or a fragment thereof, and determining whether the mental illness is depression or bipolar disorder The psychiatric disorder determination marker according to claim 1.
  17. 　配列番号１３、３４、及び１３７～１４０で示すアミノ酸配列からなる群から選択されるタンパク質、その変異体、又はその断片からなり、精神疾患が統合失調症、うつ病又は双極性障害である、請求項１に記載の精神疾患判定マーカー。 A protein selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 13, 34, and 137 to 140, a variant thereof, or a fragment thereof, wherein the mental illness is schizophrenia, depression, or bipolar disorder Item 2. A psychiatric disorder determination marker according to Item 1.
  18. 　精神疾患判定方法であって、
    　被験者及び健常者から採取された試料の単位量あたりに含まれる請求項２、３、５、６、８、９、１１～１３、及び１７に記載のいずれか一以上の精神疾患判定マーカーの量を測定してその測定値を得る測定工程、
    　前記測定工程で得られた被験者及び健常者の測定値を比較して、その結果に基づいて被験者における精神疾患を判定する比較判定工程を含む前記方法。     A method for determining mental illness,
    18. The amount of one or more mental disease determination markers according to claim 2, 3, 5, 6, 8, 9, 11 to 13, and 17, which are contained per unit amount of a sample collected from a subject and a healthy person. Measuring process to obtain the measured value,
    The said method including the comparison determination process which compares the measured value of the test subject obtained by the said measurement process, and a healthy person, and determines the mental disease in a test subject based on the result.
  19. 　前記測定工程において請求項２又は３に記載の精神疾患判定マーカーから統合失調症を判定する、請求項１８に記載の方法。 The method according to claim 18, wherein schizophrenia is determined from the mental disease determination marker according to claim 2 or 3 in the measurement step.
  20. 　請求項３に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて統合失調症の亜型をさらに判定する、請求項１９に記載の方法。 20. The method according to claim 19, wherein a subtype of schizophrenia is further determined based on the measured value of the psychiatric disorder determination marker according to claim 3 and a predetermined cutoff value.
  21. 　前記測定工程において請求項５又は６に記載の精神疾患判定マーカーから双極性障害を判定する、請求項１８に記載の方法。 The method according to claim 18, wherein bipolar disorder is determined from the mental disease determination marker according to claim 5 or 6 in the measurement step.
  22. 　請求項６に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて双極性障害の亜型をさらに判定する、請求項２１に記載の方法。 The method according to claim 21, wherein a subtype of bipolar disorder is further determined based on the measured value of the psychiatric disorder determination marker according to claim 6 and a predetermined cutoff value.
  23. 　前記測定工程において請求項８又は９に記載の精神疾患判定マーカーからうつ病を判定する、請求項１８に記載の方法。 The method according to claim 18, wherein depression is determined from the mental disease determination marker according to claim 8 or 9 in the measurement step.
  24. 　請求項９に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいてうつ病の亜型をさらに判定する、請求項２３に記載の方法。 The method according to claim 23, wherein a depression subtype is further determined based on the measured value of the mental illness determination marker according to claim 9 and a predetermined cutoff value.
  25. 　前記測定工程において請求項１１に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて統合失調症又は双極性障害を判定する、請求項１８に記載の方法。 The method according to claim 18, wherein in the measurement step, schizophrenia or bipolar disorder is determined based on the measured value of the psychiatric disorder determination marker according to claim 11 and a predetermined cut-off value.
  26. 　前記測定工程において請求項１２に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいて統合失調症又はうつ病を判定する、請求項１８に記載の方法。 The method according to claim 18, wherein in the measurement step, schizophrenia or depression is determined based on a measured value of the psychiatric disorder determination marker according to claim 12 and a predetermined cut-off value.
  27. 　前記測定工程において請求項１３に記載の精神疾患判定マーカーの測定値と所定のカットオフ値に基づいてうつ病又は双極性障害を判定する、請求項１８に記載の方法。 The method according to claim 18, wherein in the measurement step, depression or bipolar disorder is determined based on the measured value of the psychiatric disorder determination marker according to claim 13 and a predetermined cutoff value.
  28. 　前記測定工程において請求項１７に記載の精神疾患判定マーカーから統合失調症、うつ病又は双極性障害を判定する、請求項１８に記載の方法。 The method according to claim 18, wherein schizophrenia, depression, or bipolar disorder is determined from the mental disease determination marker according to claim 17 in the measurement step.
  29. 　前記測定工程において請求項４に記載の精神疾患判定マーカーの量を測定し、
    　該測定工程で得られた測定値に基づいて統合失調症の重症度を判定する重症度判定工程をさらに含む、請求項１９又は２０に記載の方法。     In the measurement step, the amount of the psychiatric disorder determination marker according to claim 4 is measured,
    The method according to claim 19 or 20, further comprising a severity determination step of determining the severity of schizophrenia based on the measurement value obtained in the measurement step.
  30. 　前記測定工程において請求項７に記載の精神疾患判定マーカーの量を測定し、
    　該測定工程で得られた測定値に基づいて双極性障害の重症度を判定する重症度判定工程をさらに含む、請求項２１又は２２に記載の方法。     In the measurement step, the amount of the psychiatric disorder determination marker according to claim 7 is measured,
    The method according to claim 21 or 22, further comprising a severity determination step of determining the severity of bipolar disorder based on the measurement value obtained in the measurement step.
  31. 　前記測定工程において請求項１０に記載の精神疾患判定マーカーの量を測定し、
    　該測定工程で得られた測定値に基づいてうつ病の重症度を判定する重症度判定工程をさらに含む、請求項２３又は２４に記載の方法。     In the measurement step, the amount of the psychiatric disorder determination marker according to claim 10 is measured,
    The method according to claim 23 or 24, further comprising a severity determination step of determining the severity of depression based on the measurement value obtained in the measurement step.
  32. 　前記測定工程において請求項１５に記載の精神疾患判定マーカーの量を測定し、
    　前記比較判定工程において測定工程で得られた測定値に基づいて統合失調症及び双極性障害のいずれであるかを判定する、請求項２５に記載の方法。     In the measurement step, the amount of the psychiatric disease determination marker according to claim 15 is measured,
    The method according to claim 25, wherein in the comparison and determination step, it is determined whether the disease is schizophrenia or bipolar disorder based on the measurement value obtained in the measurement step.
  33. 　前記測定工程において請求項１４に記載の精神疾患判定マーカーの量を測定し、
    　前記比較判定工程において測定工程で得られた測定値に基づいて統合失調症及びうつ病のいずれであるかを判定する、請求項２６に記載の方法。     The amount of the psychiatric disorder determination marker according to claim 14 is measured in the measurement step,
    The method according to claim 26, wherein in the comparison and determination step, it is determined whether the schizophrenia or depression is based on the measurement value obtained in the measurement step.
  34. 　前記測定工程において請求項１６に記載の精神疾患判定マーカーの量を測定し、
    　前記比較判定工程において測定工程で得られた測定値に基づいてうつ病及び双極性障害のいずれであるかを判定する、請求項２７に記載の方法。     The amount of the psychiatric disorder determination marker according to claim 16 is measured in the measurement step,
    28. The method according to claim 27, wherein in the comparison and determination step, it is determined whether the disease is depression or bipolar disorder based on the measurement value obtained in the measurement step.
  35. 　前記測定工程において二以上の異なる精神疾患判定マーカーを測定する、請求項１８～３４のいずれか一項に記載の方法。 The method according to any one of claims 18 to 34, wherein two or more different psychiatric disease determination markers are measured in the measurement step.
  36. 　前記試料が脳脊髄液である、請求項１８～３５のいずれか一項に記載の方法。 The method according to any one of claims 18 to 35, wherein the sample is cerebrospinal fluid.
PCT/JP2015/064233 2014-05-16 2015-05-18 Mental illness determination marker WO2015174544A1 (en)

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