JP2013212109A - Examination of schizophrenia - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for examining schizophrenia, and a testing reagent or testing equipment for schizophrenia used for the method.SOLUTION: A method for examining schizophrenia, targeting a biological sample of a subject, includes a step of measuring at least one selected from the group consisting of: (1) an amount of a protein modified product of a carbonyl compound (carbonyl protein modified product) in a biological sample; (2) an expression amount or activity of glyoxylase I in a biological sample; (3) an amount of gene abnormality of a glyoxylase I gene; and (4) an amount of pyridoxal in a biological sample.

Description

  The present invention relates to a method for testing schizophrenia, and a test reagent or test device for schizophrenia used in the method.

  Schizophrenia is a typical psychiatric disorder with hallucinations and delusions, formerly called schizophrenia. The incidence of disability is as high as 1%, and 700,000 patients are currently receiving treatment in Japan. The peak age of onset is from adolescence to adolescence from 17 to 27 years, and then chronically progresses, so 22% of all beds (1996) account for this disease. The male onset peak is 15 to 24 years old, whereas the female is 25 to 34 years old, the initial age is late, and there is another peak in menopause, so female hormones are associated with schizophrenia. It is also said that it works in a pathological manner, but the cause is still unknown. Further, as will be described later, the therapeutic method is mainly symptomatic drug administration, and a decisive therapeutic method has not yet been established.

  The coincidence rate of identical twins is 35-58%, which is higher than the coincidence rate of 13-27% of twins, suggesting that genetic factors are involved in the development of schizophrenia. Heritability is calculated to be about 80%, which is a disease with a large genetic factor compared to 30% of hypertension and 40-70% of obesity. For this reason, many candidate gene studies have been conducted since the 1990s, and the number of genes studied has reached three digits. A large-scale linkage study was also conducted, and several candidate genes were identified by a positional approach, but the pathological conditions including biochemistry could not be explained. The situation is not consistent. For this reason, schizophrenia is not a so-called genetic disease (single gene disease), but is considered to be a complex genetic disease that develops by adding environmental factors to a combination of a plurality of genes with weak pathogenic effects.

  Symptoms of schizophrenia are mainly positive symptoms (such as hallucinations, delusions, disruption of thoughts) in the acute phase, and negative symptoms that become conspicuous in the chronic phase (emotion dullness, lack of motivation / voluntaryness, social Withdrawal).

  At present, the diagnosis of schizophrenia is based on the interview with the patient, based on the contents of the statement, facial expressions, and sometimes family information. (ICD-10) (Fig. 1) or the American Psychiatric Association (APA) "Diagnostics / Statistics Manual, 4th Edition" (DSM-IV) (see Fig. 2). For this reason, the final decision must be based on subjectivity based on the experience of the attending physician, and the accuracy of diagnosis cannot be said to be sufficient. For this reason, chromosomal mapping and identification of genes responsible for schizophrenia and research using biological samples such as blood and urine of patients are actively conducted, and as a result, they can be used for diagnosis of schizophrenia. Several biological markers have been reported (for example, see Patent Documents 1 to 7). However, nothing has been established yet.

  The treatment of schizophrenia is centered on the administration of antipsychotic drugs (see FIG. 3), and medication is required for almost the entire life. The pharmacological effects of antipsychotics on hallucinations and delusions are based on the blocking action of dopamine receptors. However, by suppressing dopamine nerves, Parkinson's disease-like symptoms appear as a side effect, and thus antiparkinsonian drugs are usually used in combination. In recent years, atypical antipsychotics that are effective for negative symptoms have been developed in place of conventional antipsychotics that are effective for positive symptoms of schizophrenia but have little effect on negative symptoms It has been replaced by conventional drugs. The atypical antipsychotic has an advantage that it has less side effects like Parkinson's disease because the blocking action on dopamine receptors is not as strong as that of the conventional type.

  Treatment methods other than antipsychotic medication include electroconvulsive therapy (ECT), which is a treatment method that induces generalized convulsions by applying 100V alternating current to the head for 5 seconds, as well as agriculture, woodworking, handicrafts, recreation, etc. Mention psychiatric rehabilitation such as occupational therapy and social life skills training (SST). The former ECT is a method applied to refractory symptoms such as imminent suicide risk, malnutrition, catatonic conditions, and cases resistant to drug treatment. The latter type of psychiatric rehabilitation aims to reduce stress in social life and prevent recurrence by acquiring life skills.

  However, both treatments are only symptomatic treatments and are old data, but as a result of follow-up for 15 years in chronic intractable cases with no drug treatment effect, recovery was 6%. It is reported as 8% good, 23% moderate, 23% border, and 41% persistent disability (Non-patent Document 1).

  By the way, advanced glycation end products (hereinafter referred to as “AGEs”) react non-enzymatically with carbonyl compounds and proteins (amino groups) generated from sugars and lipids under high blood sugar and oxidative stress. It is a substance produced in the body by (Maillard reaction) (the "AGEs are included in the" protein modification products of carbonyl compounds (carbonyl protein modification products) "in the present invention). AGEs are heterogeneous aggregates composed of a number of structures. Pentosidine, one of the structures that make up the aggregates, was isolated from human brain dural collagen in 1989 by Sell et al. It is a substance. In recent years, analysis results using antibodies against these AGEs (anti-AGE antibodies) have revealed that AGEs in tissues and blood have increased in various pathological conditions. For example, due to hyperglycemia in diabetes, an increase in carbonyl compounds derived from sugar, which is a precursor of AGEs, and protein modifications (AGEs) thereof are observed. In renal failure, production of carbonyl compounds is increased and protein modifications (AGEs) are increased due to decreased excretion of carbonyl compounds and increased oxidative stress, and in inflammatory diseases, increased oxidative stress. Furthermore, it has been reported that AGEs are also increased in patients who lack glyoxylase, an enzyme that eliminates carbonyl compounds. For this reason, blood AGEs are actually used in clinical tests as indicators of diabetes and renal vascular complications. Methods for measuring AGEs include methods that measure the amount of pentosidine that is the structure of AGEs by ELISA using an anti-pentosidine antibody, and methods that measure the content of pentosidine in the skin with an AGE Reader device. Yes.

Japanese Patent Laid-Open No. 2001-245661 JP 2003-38198 A Japanese Patent Laid-Open No. 2003-212795 WO2004 / 005935 Brochure JP 2004-251865 A JP 2005-55227 A JP 2005-278490 A

McGlashan Schizophr Bull, 1988

  Schizophrenia has a special pathological condition and has a long medical history as described above, so that the mental pain and burden of the person and his family are great. In addition, since the number of affected people is high at 1%, social loss due to this mental illness cannot be measured. Furthermore, as mentioned above, the fact that 22% of all beds are occupied by patients with schizophrenia is a big problem from the viewpoint of medical economy. Therefore, early establishment of a comprehensive diagnosis and treatment system such as early diagnosis, treatment, rehabilitation activities, and prevention of recurrence is desired.

  The object of the present invention is to provide a novel technique that can be used effectively in the diagnosis and treatment of schizophrenia. Specifically, an object of the present invention is to provide a test method for schizophrenia and a test reagent and a test device that can be effectively used in the method.

  As a result of gene analysis for schizophrenic patient families, Itokawa, one of the present inventors, found that one allele was frame-shifted due to point mutations in schizophrenic patient families, and glyoxylase was We found that there are several patient families that are missing. Therefore, when Miyata, one of the present inventors, conducted biological and chemical analysis of such patients, in this schizophrenic family, the activity of erythrocyte glyoxylase dropped to about half that of healthy individuals. , High levels of carbonyl compounds and AGEs (ie carbonyl stress) despite the absence of other conditions such as renal failure, diabetes and inflammation, and consumption of vitamin B6 (pyridoxal) in the body to eliminate carbonyls It is possible to confirm that a series of biochemical abnormalities have occurred in patients with schizophrenia based on glyoxylase genetic abnormalities, such as increased levels of homocysteine. did it.

  Furthermore, through further studies by the present inventors, carbonyl stress and vitamin B6 deficiency could be confirmed not only in schizophrenic patients deficient in the above glyoxylase but also in some general schizophrenia. Specifically, in schizophrenic patients (5 out of 1099) having the 111th amino acid mutation (Glu → Ala) of glyoxylase I as a homozygote, the activity of erythrocyte glyoxylase is It was reduced to about 80%, and a high level of carbonyl compounds and AGEs in blood (carbonyl stress) and a low level of vitamin B6 were confirmed. The glyoxylase I gene is known to have polymorphism (with low activity polymorphism), but none of the above-mentioned 111 Ala / Ala homozygotes were detected from 854 healthy subjects Therefore, it was considered that the decrease in the activity of glyoxylase based on the mutation caused a series of biochemical abnormalities related to schizophrenia.

  Thus, the present inventors have (1) glyoxylase gene abnormality in a subject, (2) glyoxylase activity in a biological sample, (3) the amount of carbonyl compounds or protein modifications (AGEs) thereof, And (4) at least one selected from the amount of pyridoxal in the biological sample, preferably (1) measuring at least one of the glyoxylase gene abnormalities or (3) the amount of AGEs and indicating it as an index As a result, it was confirmed that the diagnosis of schizophrenia, which has conventionally relied on subjective judgments by doctors' interviews, can be performed simply and objectively, and the present invention has been completed. Furthermore, the present inventors have convinced from the above findings that a carbonyl scavenger or an AGEs production inhibitor that suppresses carbonyl stress is effective in the treatment of schizophrenia or the improvement of symptoms, thereby completing the present invention. It came to.

That is, the present invention relates to a method for examining schizophrenia and a test agent for schizophrenia used in the method described below.
I. Method for examining schizophrenia (I-1)
(1) Amount of protein modification product (carbonyl protein modification product) of a carbonyl compound in a biological sample,
(2) expression level or activity of glyoxylase I in a biological sample,
(3) a genetic abnormality of the glyoxylase I gene, and
(4) measuring at least one selected from the group consisting of the amount of pyridoxal in the biological sample,
Testing method for schizophrenia.
(I-2) The method for examining schizophrenia according to (I-1), comprising the following steps (A) and (B):
(A) For a biological sample of a subject, (1) the amount of a protein modification product of a carbonyl compound in the biological sample, (2) the expression level or activity of glyoxylase I in the biological sample, (3) glyoxylase A step of measuring at least one selected from the group consisting of a genetic abnormality of an I gene, and (4) a pyridoxal amount in a biological sample, and
(B) A step of comparing the measured value in the subject obtained in the step (A) with the measured value (control value) in a healthy subject corresponding to the measured value.
(I-3) When each measurement value measured on the biological sample of the subject falls under the following, the subject is recognized as having schizophrenia or having a high risk of developing it in the future, ( Testing method for schizophrenia described in I-2):
(1) The amount of the protein modification product of the carbonyl compound in the biological sample of the subject is greater than the amount in the healthy human biological sample,
(2) The expression level or activity of glyoxylase I in the biological sample of the subject is lower than the expression level or activity in the biological sample of a healthy person,
(3) The subject's glyoxylase I gene has an abnormality that causes a decrease in glyoxylase I activity,
(4) The amount of pyridoxal in the subject's biological sample is less than that in the healthy subject's biological sample.
(I-4) (4) In addition to the measurement of the amount of pyridoxal in the biological sample, (5) further comprising the step of measuring the amount of homocysteine in the biological sample of (I-1) to (I-3) The test method of schizophrenia described in any one.
(I-5) The subject has symptoms or findings characteristic of schizophrenia as defined in the diagnostic criteria for schizophrenia, or is a person suspected of having schizophrenia according to the criteria (I-1 The inspection method of schizophrenia as described in any one of) thru | or (I-4).
(I-6) The test method for schizophrenia according to any one of (I-1) to (I-5), wherein the subject is a person who does not have renal dysfunction, diabetes or inflammation.
(I-7) (3) A glyoxylase I gene abnormality is detected between the adenine at position 79 and the cytosine at position 80 in the base sequence of the glyoxylase I gene shown in SEQ ID NO: 1 (SEQ ID NO: 2). The method for examining schizophrenia according to any one of (I-1) to (I-6), which is a frameshift abnormality caused by adenine insertion.
(I-8) (3) The glyoxylase I gene abnormality is a base substitution mutation that results in a mutation of the 111th amino acid (Glu → Ala) in the amino acid sequence of glyoxylase I shown in SEQ ID NO: 3. (I-1) thru | or the inspection method of schizophrenia described in any one of (I-6)
(I-9) At least one selected from the group consisting of (1), (2) and (4) above for a subject having a glyoxylase I gene abnormality leading to a decrease in glyoxylase I activity The method for testing for schizophrenia according to any one of (I-1) to (I-6), comprising a step of measuring one.
(I-10) A frame in which the above glyoxylase I gene abnormality is caused by insertion of adenine between adenine at position 79 and cytosine at position 80 in the base sequence of the glyoxylase I gene shown in SEQ ID NO: 1 (SEQ ID NO: 2) The examination for schizophrenia according to (I-9), which is a shift abnormality or a base substitution mutation that causes a mutation of the 111st amino acid (Glu → Ala) in the amino acid sequence of glyoxylase I shown in SEQ ID NO: 3 Method.
(I-11) Schizophrenia according to any one of (I-1) to (I-10), comprising a step of measuring the amount of (1) a protein modification product (carbonyl protein modification product) of a carbonyl compound in a subject. Detection method for the carbonyl protein modification product, such as immunological technique, high performance liquid chromatography, gas chromatography / mass spectrometry, or AGE-Reader using anti-carbonyl protein modification antibody A method characterized by using a method.
(I-12) The method for examining schizophrenia according to any one of (I-1) to (I-11), wherein the modified carbonyl protein is pentosidine.

II. Test reagent or test device for schizophrenia and use thereof (II-1) In the glyoxylase I gene shown in SEQ ID NO: 1, the adenine at position 79 and the 80th position in the base sequence of the coding region (SEQ ID NO: 2) Consecutive oligos or polynucleotides having a length of 16 bases or more containing cytosine (however, when the oligonucleotide or polynucleotide is RNA, the base symbol “t” in the sequence listing should be read as “u”) An oligonucleotide having a length of 15 to 35 bases or a label thereof used for hybridizing and specifically amplifying the oligo or polynucleotide.
(II-2) Identifies and integrates the adenine insertion between the 79th position adenine and the 80th position cytosine in the base sequence of the glyoxylase I gene coding region shown in SEQ ID NO: 1 (SEQ ID NO: 2). A primer comprising the oligonucleotide or the label thereof described in (II-1), which is used for examining ataxia.
(II-3) In the glyoxylase I gene shown in SEQ ID NO: 1, a continuous oligo or poly having a length of 16 bases or more containing adenine at position 79 and cytosine at position 80 in the base sequence of the coding region (SEQ ID NO: 2) Nucleotides or continuous oligos or polynucleotides having a length of 16 bases or more including the base at position 332 (provided that these oligonucleotides or polynucleotides are RNA, the base symbol “t” in the sequence listing is “u”. The oligonucleotide having a length of 16 to 500 bases or a labeled product thereof, which is used for specifically detecting the oligo or polynucleotide.
(II-4) Insertion between adenine at position 79 and cytosine at position 80 or adenine at position 332 in the base sequence of the coding region of the glyoxylase I gene shown in SEQ ID NO: 1 (SEQ ID NO: 2) A probe comprising the oligonucleotide or the labeled product thereof according to (II-3), which is used for identifying a mutation from cysteine to cytosine and examining schizophrenia.
(II-5) A test reagent for schizophrenia comprising the primer described in (II-2) and / or the probe described in (II-4), or a kit containing the reagent.
(II-6) A reagent for use in a test for schizophrenia comprising a step of measuring the expression level or activity of glyoxylase I in a biological sample of a subject, and comprising at least an anti-glyoxylase I antibody or glycine A test reagent containing a reaction substrate for oxylase I.
(II-7) A reagent used for testing schizophrenia having a step of measuring the amount of (1) a carbonyl protein modification in a biological sample of a subject, (a) a carbonyl protein modification at a known concentration, or (b) A test reagent containing at least one of the anti-carbonyl protein modified antibodies which may be labeled.
(II-8) The test reagent according to (II-5), wherein the modified carbonyl protein is pentosidine.
(II-9) A reagent used in the examination of schizophrenia having a step of measuring the amount of pyridoxal in a biological sample of a subject, comprising (a) a known concentration of pyridoxal, or (b) a pyridoxal measurement reagent A test reagent containing at least one of the above.
(II-10) An AGE-Reader device for use in diagnosing schizophrenia, comprising means for measuring the amount of carbonyl protein modification in the skin of a subject.
(II-11) The AGE-Reader device according to (II-10), which is accompanied by a document stating that it can be used for diagnosis of schizophrenia.
(II-12) Use of an AGE-Reader device having a means for measuring the amount of carbonyl protein modification in the skin of a subject for diagnosis of schizophrenia.

According to the diagnostic method of the present invention, (1) the genetic abnormality of the glyoxylase I gene in the subject, (2) the expression level or activity of glyoxylase in the biological sample, (3) the carbonyl compound or the protein modification product thereof By using at least one selected from the group consisting of the amount and (4) the amount of pyridoxal in the biological sample as an index, diagnosis of schizophrenia that has traditionally relied on subjective judgments by doctors' interviews can be simplified and performed. Can be done objectively.

Diagnostic criteria for schizophrenia according to the World Health Organization (WHO) "International Disease Classification-10th Edition" (ICD-10) is shown. The diagnostic criteria of schizophrenia by the American Psychiatric Association (APA) "Diagnosis and Statistical Manual, 4th Edition" (DSM-IV) are shown. It is the figure which listed the antipsychotic drug currently used for the treatment of schizophrenia. It is a figure which shows the result of having compared the erythrocyte glyoxalase I activity with a serious schizophrenia patient (n = 1) and a healthy subject (n = 5) (Example 1). The glyoxalase I gene analysis result of a severe schizophrenic patient (patient A) is shown. In FIGS. 5A and 5B, gene M and gene X both mean glyoxalase I gene. In FIG. A, case 2 means a schizophrenic patient (patient A). It is a figure which shows the result of having compared the serum AGEs value (the amount of pentosidine) with a serious schizophrenia patient (n = 1) and a healthy subject (n = 5) (Example 1). It is a figure which shows the result of having compared the AGE content in skin with a healthy subject (n = 24) and a schizophrenia patient (n = 24) (Example 2). It is a figure which shows the result of having compared the AGEs content in skin according to age (50 years old or younger, older than 50 years old) by a healthy subject (n = 24) and a schizophrenia patient (n = 24) (Example) 2). It is a figure which shows the result of having measured the erythrocyte glyoxalase activity about the schizophrenia patient of Ala / Ala homozygote, the schizophrenia patient of Glu / Ala heterozygote, and a Glu / Glu heterozygote, and a healthy subject. Example 3). The COS cells express the mutant GLO-I and GFP construct (pAcGFP-GLO1-Ala) at position 111 and normal GLO-I and GFP (pAcGFP-GLO1-Glu) at position 111, respectively. (Example 3) which shows the result of having measured GLO-I activity. The figure which shows the result of having measured the expression level of mRNA of glyoxalase I about Ala / Ala homozygous schizophrenia patient (n = 4) and serious schizophrenia patient A (glyoxalase I deficiency by frame shift). is there. Patients with schizophrenia of each type (frameshift [n = 2], Ala / Ala homozygous [n = 5], Glu / Ala heterozygous [n = 1]), and healthy individuals (Glu / Glu homozygous) Conjugated type) [n = 7], Glyoxalase I activity (mUnit / 10 ⁶ RBC), pentosidine activity (pmol / mg protein), blood vitamin B6 content (ng / ml) (pyridoxal, pyridoxamine, pyridoxine), blood Vitamin B12 content (pg / ml), blood folate content (ng / ml), blood homocysteine content (nmol / ml), creatinine amount (mg / dl), and eGFR (ml / min / 1.73m ² ) The results of measurement are summarized. The result of having investigated the pentosidine production inhibitory effect about aminoguanidine, pyridoxamine, olmesartan, edaravone, and TM2002 is shown (test example 1 (2)).

I. Method for examining schizophrenia The method for examining schizophrenia of the present invention can be performed by measuring at least one of the following using a biological sample of a subject as a material:
(1) a genetic abnormality of the glyoxylase I gene,
(2) Glyoxylase I expression level or activity,
(3) Amount of carbonyl compound or protein modification product thereof (for example, “advanced glycation end products” (AGEs)),
(4) Amount of pyridoxal.

  Further, (4) when measuring the amount of pyridoxal in a biological sample, (5) the amount of homocysteine in the biological sample may be further measured.

  Examples of biological samples that can be used in the test of the present invention include blood and blood components (serum, plasma, blood cells, etc.), urine, spinal fluid, saliva, tears, sweat, and other biological fluid components; hair And a solid component derived from a living body such as a part of a tissue excised by biopsy or the like. Blood or blood components are preferable, and serum is particularly preferable. In addition, even if it uses directly the sample extract | collected from the biological body, the sample adjusted by adding a certain process can also be used as the object of the biological sample of this invention.

  The subject is preferably a human. However, the present invention is not limited to this, and mammals other than humans, such as monkeys, mice, rats, guinea pigs, etc. can also be targeted.

  In addition, the subject can have a symptom or a finding characteristic of schizophrenia as defined in the diagnostic criteria for schizophrenia, or can be a person suspected of having schizophrenia according to the diagnostic criteria. Here, diagnostic criteria for schizophrenia can include those commonly used in the industry. Specifically, the World Health Organization (WHO) International Disease Classification (ICD) and US Psychiatry You can list the “Diagnostic and Statistical Manual” (DSM) of the Association (APA). Fig. 1 shows the criteria for schizophrenia according to the former "International Disease Classification" 10th edition (ICD-10), and Fig. 1 shows the criteria for schizophrenia according to the latter "Diagnosis and Statistical Manual" 4th edition (DSM-IV). It is shown in 2.

  When examining schizophrenia using the above (2) to (4) or (2) to (5) as an index, the subject may have a glyoxylase I gene abnormality, and The person who received the test | inspection which measures the gene abnormality of the glyoxylase I gene demonstrated in (1) may be sufficient. The target glyoxylase I gene abnormality means a gene abnormality that results in a decrease in glyoxylase I activity, such as a deficiency in glyoxylase I or a poor expression of glyoxylase I. Specifically, at least one allele of the glyoxylase I gene is point-mutated (adenine insertion between positions 79 and 80 of the base sequence (SEQ ID NO: 2) of the coding region of the glyoxylase I gene) In this case, a case where the frame is shifted or a case where the mutation (Glu → Ala) at the 111st amino acid of the amino acid sequence of glyoxylase I (SEQ ID NO: 3) is homozygous can be mentioned. In the case of the former gene abnormality, the glyoxylase I activity in the blood is about 50% lower than that in the healthy subject. In the latter gene abnormality, the glyoxylase I activity in the blood is reduced. It has been confirmed that it is about 20% lower than that of healthy subjects (see Examples). In addition, not only these gene abnormalities but gene abnormalities that cause a decrease in glyoxylase I activity may be used.

  When schizophrenia is examined using the above (3) to (4) or (3) to (5) as an index, the subject is preferably a person who does not have at least renal dysfunction, diabetes and inflammation. Since (3) carbonyl compounds or protein modifications thereof (AGEs), (4) pyridoxal amount, and (5) homocysteine amount that the present invention uses as an index of examination vary depending on the presence or absence of renal dysfunction, diabetes or inflammation It is. Therefore, when examining schizophrenia using the above (3) to (4) or (3) to (5) as an index, renal dysfunction, diabetes or inflammation in advance or in parallel with the examination of the present invention. The presence or absence is preferably confirmed by an interview or / and biochemical examination.

(1) Measurement of genetic abnormality of glyoxylase I gene The genetic abnormality of the glyoxylase I gene (hereinafter also simply referred to as “GLO-I” gene) targeted by the present invention is expressed as glyoxylase I (GLO-I). ) Deficiency and poor expression of GLO-I, resulting in a genetic abnormality that leads to a decrease in GLO-I activity. Although not particularly limited, specifically, (1-1) at least one allele of the base sequence of the coding region of GLO-I gene (SEQ ID NO: 2) is point-mutated (adenine at position 79 and 80 Insertion of adenine between cytosine in position), gene abnormality shifted by this, and (1-2) mutation of amino acid 111 of GLO-I amino acid sequence (SEQ ID NO: 3) (Glu → Ala) Base substitution mutations that cause homozygosity. As described above, in the case of the former gene abnormality, the GLO-I activity in the blood is reduced by about 50% as compared with that in the healthy person. In the latter case, the GLO-I in the blood is decreased. It has been confirmed that I activity is reduced by about 20% compared to healthy subjects (see Examples).

  Specifically, the detection of such a gene abnormality includes (i) a method in which PCR is performed in a region containing a gene abnormality and detected by the SSCP method, (ii) PCR is performed in a region containing the gene abnormality, and a restriction enzyme for the PCR product is detected. (Iii) a method of directly sequencing the PCR product and determining the sequence; (iv) using an oligonucleotide that hybridizes to a region having a genetic abnormality as a probe, By using known methods such as ASO (allele specific oligonucleotide) method to hybridize with, and (v) a method using a oligonucleotide that hybridizes to a region having a gene abnormality as a probe and detecting with a mass spectrometer etc. It can be carried out.

As a more specific detection method, (1-1) 1 base insertion (adenine insertion) between adenine at position 79 and cytosine at position 80 in the base sequence of the coding region of GLO-I gene (SEQ ID NO: 2) (1-2) GLO-I gene, which is one of the polymorphisms that cause mutation (Glu → Ala) of the 111th amino acid of the amino acid sequence of GLO-I (SEQ ID NO: 3) A method for identifying a mutation at position 332 (a → c) in the base sequence of the coding region (SEQ ID NO: 2) can be exemplified. The detection method can be carried out by performing the following steps (a) and (b):
(a) extracting genomic DNA from a biological sample of a subject, and
(b) A step of identifying the base sequence of the region of positions 79-81 or the base of position 332 of the base sequence of the coding region for the GLO-I gene contained in the extracted genomic DNA.

  In the step (b), it is not actually necessary to identify and target the GLO-I gene on the extracted genomic DNA, but the extracted genomic DNA itself is the target and is present in the base sequence, SEQ ID NO: 4 It can also be carried out by identifying a base sequence sandwiched between the base sequence described and the base sequence described in SEQ ID NO: 5 or a base sequence sandwiched between the base sequence described in SEQ ID NO: 6 and the base sequence described in SEQ ID NO: 7.

  Here, the base sequence represented by SEQ ID NO: 4 is the positions 79-80 of the base sequence of the coding region (SEQ ID NO: 2) in the base sequence of human GLO-I gene (SEQ ID NO: 1) (Genbank accession number NM_006708). Corresponds to a base sequence of 200 bases located on the 5 ′ side (upstream side) of the base sequence (ac) located in FIG. 5, and the base sequence shown in SEQ ID NO: 5 is the base sequence (sequence) of the human GLO-I gene No. 1) corresponds to a base sequence having a length of 300 bases located on the 3 ′ side (downstream side) of the base sequence (ac) located at positions 79-80 of the base sequence of the coding region (SEQ ID NO: 2). In addition, the base sequences shown in SEQ ID NOs: 6 and 7 are those of the base (a) located at position 332 of the base sequence of the coding region (SEQ ID NO: 2) in the base sequence of the GLO-I gene (SEQ ID NO: 1). These correspond to a base sequence having a length of 300 bases located on the 5 ′ side (upstream side) and a base sequence having a length of 300 bases located on the 3 ′ side (downstream side), respectively.

The method (1-1) of the present invention preferably further comprises the following step (c-1):
(c-1) A step of detecting whether or not adenine is inserted between 79-position adenine and 80-position cytosine in the base sequence (SEQ ID NO: 2) of the coding region of the GLO-I gene.

  In the case where (1-1) adenine is inserted between 79-position adenine and 80-position cytosine of the base sequence (SEQ ID NO: 2) of the coding region of GLO-I gene by the detection, the genomic DNA sample is The provided subject has schizophrenia or is likely to develop schizophrenia in the future.

The method (1-2) of the present invention preferably further comprises the following step (c-2):
(c-2) A step of detecting whether the base at position 332 of the base sequence of the coding region of the GLO-I gene (SEQ ID NO: 2) is adenine or cytosine.

  (1-2) When the base at position 332 is a cytosine homozygote (C / C), the subject who has provided the genomic DNA sample has schizophrenia or a future schizophrenia. Is likely to develop.

  According to this method, the presence or absence of schizophrenia or the potential risk of the onset can be determined for the subject. The test can be performed mechanically without requiring the judgment of a person having specialized knowledge such as a doctor using the gene abnormality as a judgment criterion (determination index). For this reason, the method of this invention can also be called the detection method of schizophrenia.

  The step (a) (extraction of genomic DNA) and the step (b) (identification of target base) are carried out by known methods [for example, Bruce, et al., Geneme Analysis / A laboratory Manual (vol.4), Cold Spring Harbor Laboratory, NY. (1999)].

  The specimen from which genomic DNA is extracted in step (a) is any cell (including cultured cells, excluding germ cells), tissue (eg, liver, kidney, adrenal gland, uterus) isolated from subjects and clinical specimens. , Brain, etc., including cultured tissues) or body fluids (eg saliva, lymph, airway mucosa, semen, sweat, urine, etc.). As the material, leukocytes or mononuclear cells separated from peripheral blood are preferable, and leukocytes are particularly preferable. These materials can be isolated according to methods commonly used in clinical laboratory tests.

For example, when leukocytes are used as a material, leukocytes are first separated from peripheral blood isolated from a subject according to a conventional method. Subsequently, proteinase K and sodium dodecyl sulfate (SDS) are added to the obtained leukocytes to degrade and denature proteins, and then phenol / chloroform extraction is performed to obtain genomic DNA (including RNA). RNA can be removed by RNase as necessary. Incidentally, extraction of the genomic DNA is not limited to the above method, methods well known in the art (e.g., Sambrook J. et al,.. "Molecular Cloning:. A Laboratry Manual (2 nd Ed)" Cold Spring Harbor Laboratory, NY) and commercially available DNA extraction kits can be used. Further, if necessary, the GLO-I gene or a DNA containing it may be isolated. The DNA can be isolated by PCR using genomic DNA as a template, using a primer that hybridizes to the GLO-I gene.

  In step (b), the presence or absence of an inserted base between positions 79 and 80 in the base sequence (SEQ ID NO: 2) of the coding region of the GLO-I gene from the extract containing human genomic DNA obtained as described above Or the base at position 332 of the base sequence of the coding region of the GLO-I gene (SEQ ID NO: 2). The base is identified by directly determining the base sequence of the coding region of the GLO-I gene further isolated from a sample containing human genomic DNA, and identifying the base located between positions 79 and 80 of the base sequence. It may be determined by a method for examining the presence or absence or the type of base at position 332 (adenine or cytosine).

  For example, as a method for identifying the target base, in addition to the method for directly determining the gene sequence of the corresponding region as described above, if the polymorphic sequence is a restriction enzyme recognition site, the difference in restriction enzyme cleavage pattern is used. Then, a method for determining a genotype (hereinafter referred to as RFLP), a method based on hybridization using a polymorphism-specific probe (for example, attaching a specific probe on a chip, glass slide, nylon membrane, By detecting the difference in hybridization intensity for these probes, the type of polymorphism can be determined, or the efficiency of hybridization of a specific probe can be determined by the amount of probe that polymerase degrades during template duplex amplification. The method of identifying the genotype by detection, the fluorescence emitted by a certain double-strand specific fluorescent dye, By detecting the temperature difference of double-strand melting by following the change, a polymorphism is identified by this method, and complementary sequences are attached to both ends of the polymorphic site-specific oligo probe. And a method for specifying a genotype using a difference between whether a secondary structure is formed in the structure or hybridizing to a target region. In addition, a method in which a base-extension reaction is performed by polymerase from a template-specific primer and a base incorporated into a polymorphic site at that time is identified (by using dideoxynucleotide, each is fluorescently labeled, and each fluorescence is Detection method, method of detecting incorporated dideoxynucleotide by mass spectrometry), and further using template-specific primers followed by the presence or absence of base pairs complementary to the mutation site or non-complementary base pairs. There is a method to make it recognize.

  Hereinafter, conventionally known representative methods for detecting gene polymorphisms are listed, but the present invention is not limited to these.

  (a) RFLP (restriction fragment length polymorphism) method, (b) PCR-SSCP method (single-stranded DNA higher-order structure polymorphism analysis) [Biotechniques, 16, 296-297 (1994), and Biotechniques, 21 , 510-514 (1996)], (c) ASO (Allele Specific Oligonucleotide) hybridization method [Clin. Chim. Acta, 189, 153-157 (1990)], (d) Direct sequencing method [Biotechniques, 11, 246 -249 (1991)], (e) ARMS (Amplification Refracting Mutation System) method (Nuc. Acids. Res., 19, 3561-3567 (1991); Nuc. Acids. Res., 20, 4831-4837 (1992) (F) Denaturing Gradient Gel Electrophoresis (DGGE) method [Biotechniques, 27, 1016-1018 (1999)], (g) RNase A cleavage method [DNA Cell. Biol., 14, 87 -94 (1995)], (h) chemical cleavage method [Biotechniques, 21, 216-218 (1996)], (i) DOL (Dye-labeled Oligonucleotide Ligation) method [Genome Res., 8, 549-556 (1998) )], (J) T qMan-PCR method [Genet. Anal., 14, 143-149 (1999); J. Clin. Microbiol., 34, 2933-2936 (1996)], (k) Invader method [Science, 5109, 778-783 ( 1993); J. Biol. Chem., 30, 21387-21394 (1999); Nat. Biotechnol., 17, 292-296 (1999)], (l) MALDI-TOF / MS method (Matrix Assisted Laser Desorption-time of Flight / Mass Spectrometry) [Genome Res., 7, 378-388 (1997); Eur. J. Clin. Chem. Clin. Biochem., 35, 545-548 (1997)], (m) TDI (Template -directed Dye-terminator Incorporation method [Proc. Natl. Acad. Sci. USA, 94, 10756-10761 (1997)], (n) Molecular Beacons method [Nat. Biotechnol., 1, p49- 53 (1998); gene medicine, 4, p46-48 (2000)], (o) Dynamic Allele-Specific Hybridization (DASH) method [Nat. Biotechnol., 1.p.87 -88 (1999); gene medicine, 4, 47-48 (2000)], (p) padlock Probe (Padlock Probe) method [Nat. Genet., 3, p225-232 (1998); Gene medicine, 4, p50-51 (2000)], (q) UCAN method [Takara Shuzo Co., Ltd. home page (http: http://www.takara.co.jp)], (r) DNA chip or DNA microarray ("SNP gene polymorphism strategy" Kenichi Matsubara, Yoshiyuki Tsuji, Nakayama Shoten, p128-135), (s) ECA method [ Anal. Chem., 72, 1334-1341, (2000)].

  The above are typical gene polymorphism detection methods, but the schizophrenia test of the present invention is not limited to these, and other known or future developed gene abnormality detection methods can also be used. Moreover, when detecting a genetic abnormality of the present invention, these gene polymorphism detection methods may be used alone or in combination of two or more.

  For a subject who has been found to be schizophrenia or has a relatively high potential risk of developing schizophrenia by the method of the present invention, the fact is notified, and the treatment or improvement thereof , Or take appropriate measures to prevent its onset. Therefore, the present invention is extremely useful as an examination method for early treatment or improvement of schizophrenia, or for preventing the onset and further for preventing the progress (progress).

  The method of the present invention is not limited to a method for detecting a gene abnormality from a base sequence, but a mutation (Glu → Ala) at the 111th amino acid in the amino acid sequence of GLO-I (SEQ ID NO: 3) resulting from the gene abnormality is directly used. A detection method may be used. Examples of such a method include a method of measuring whether or not the amino acid mutation has occurred in a GLO-I gene expression product (eg, mRNA, GLO-I) of a subject. Such methods include a method for determining the sequence of the expression product of the GLO-I gene, a Western blotting method using an antibody against GLO-I or a mutant GLO-I (Glu111Ala), a dot blotting method, an immunoprecipitation method, an enzyme Examples thereof include a method for measuring the activity generated (or eliminated) by the linked immunoassay (ELISA) and the immunofluorescence method, or the 111th amino acid mutation (Glu → Ala) in the amino acid sequence of GLO-I. .

(2) Measurement of expression level or activity of glyoxylase I in a biological sample Such measurement can be preferably performed using blood, particularly serum or blood cells as a material.

  The expression level of GLO-I is usually determined by using known antibodies such as Western blotting, EIA method, RIA method, FIA method, chemiluminescence immunoassay, or ECLIA method using an antibody against GLO-I (polyclonal antibody or monoclonal antibody). Can be measured using conventional techniques. The Western blotting method using a monoclonal antibody against GLO-I is preferred. Specifically, the test sample was treated with mercaptoethanol, subjected to electrophoresis on an SDS-polyacrylamide gel, and then the protein was transferred to a PVDF membrane, followed by anti-GLO-I antibody, then labeled second antibody and The method of making it react and making it color and visualize can be mentioned. It should be noted that GLO-I mRNA can be quantified more accurately by using a protein derived from a housekeeping gene such as β-actin as an internal standard as necessary in Western blotting. An antibody against GLO-I can be prepared according to a conventional method using GLO-I, and can also be obtained commercially (for example, GLO1 polyclonal antibody [Abnova Corporation / Taipei City 114 Taiwan]). Catalog #: H00002739-A01]).

The activity of GLO-I can be performed using methods known in the art. Although not limited, for example, as described in Example 1, McLellan et al. (McLellan AC, Thornalley PJ: Glyoxalase activity in human red blood cells fractioned by age. Mech Aging Dev 48: 63-71, 1989) Can be mentioned. Such a method is a method for evaluating GLO-I activity in erythrocytes by reacting the broken erythrocytes with hemithioacetal generated from methylglyoxal and glutathione, and measuring the amount of SD-lactoyl glutathione produced. . Incidentally, 1 Unit erythrocytes 10 ⁶ per minute in such a method, means an amount for forming the SD-lactoylglutathion of 1 [mu] mol.

  More specifically, in the diagnostic method of the present invention, after measuring the expression level or activity of GLO-I in a biological sample of a subject by the above method, the measured value is expressed as the corresponding GLO-I expression in a healthy subject. This can be done by assessing the level compared to the amount (control expression amount) or activity (control activity). In this case, the determination that the subject is suffering from schizophrenia can be used as an indicator that the subject's expression level or activity of GLO-I is lower than the control expression amount or control activity.

(3) Measurement of a carbonyl compound or a protein modification product thereof in a biological sample The measurement can be performed preferably using blood, particularly serum or plasma.

  Target carbonyl compounds include arabinose, GO, MGO, 3-DG, glycol aldehyde, dehydroascorbic acid, hydroxynonenal, malondialdehyde, acrolein, 5-hydroxymethylfurfural, formaldehyde, acetaldehyde, repric acid, furfural, etc. Can be mentioned. These amounts can usually be measured by performing instrumental analysis such as high performance liquid chromatography (HPLC) or gas chromatography / mass spectrometry (GC / MS) using standard products of known concentrations.

  Similarly, protein modifications (for example, “AGEs”) (hereinafter referred to simply as “AGEs”) produced by the reaction of these carbonyl compounds with biological proteins are also analyzed using standard products of known concentrations. And can be measured by instrumental analysis such as GC / MS. AGEs are an aggregate of a plurality of structures (AGEs structures). Therefore, AGEs can be measured by measuring the AGEs structure. AGEs structures include pentosidine, croslin, pyropyridine, vesperlidine A, B, C, glyoxal-lysine dimmer (GOLD), methylglyoxal-lysine dimmer (MOLD) (above, fluorescent substance), Nε- (carboxymethyl) lysine ( CML), Nε- (carboxyethyl) lysine (CEL), argpyrimidine, pyralin, imidazolone, GA-pyridine (hereinafter, non-fluorescent substance) and the like. Pentosidine is preferred.

  Pentosidine has a structure in which pentose, equimolar lysine and arginine are cross-linked, and is a fluorescent substance stable to acid hydrolysis. It is known to accumulate in human skin in correlation with aging and the onset of diabetes, and it has been reported to increase especially in the onset of diabetes and end-stage nephropathy. Pentosidine contained in proteins such as blood can be quantified by HPLC after acid hydrolysis using the fluorescence (Ex: 335 nm, Em: 385 nm) as an index. Pentosidine can be quantified using an immunochemical method using a monoclonal antibody against pentosidine (for example, ELISA, particularly sandwich ELISA or competitive ELISA). In addition, although the monoclonal antibody and polyclonal antibody with respect to pentosidine can be prepared in accordance with a conventional method, they can also be obtained commercially easily (for example, (transgenic), Fushimi Pharmaceutical, etc.). In addition, carboxymethyl lysine, which is an AGEs structure, and antibodies against protein modifications of malondialdehyde and hydroxynonenal are also commercially available.

  In the competitive ELISA method, a specimen (plasma) pretreated with proteolytic enzyme and a standard AGEs structure are added to a microplate on which an AGEs structure such as pentosidine is immobilized, and an antibody against the AGEs structure is added and reacted. Then, after washing, the enzyme solution can be added and washed again, and a color former can be added to measure the absorbance. This method is a method defined as a blood chemistry test for pentosidine, and can be carried out simply by using a quantitative kit such as “FSK Pentosidine” (product name) (Fushimi Pharmaceutical Co., Ltd.). In addition, the amount of AGEs such as pentosidine accumulated in the skin can be easily measured using a commercially available AGE-Reader device (Diagn Optics).

  More specifically, after the amount of carbonyl compounds or protein modifications (AGEs) thereof in a biological sample of a subject is measured by the above method, the test method of the present invention is used to calculate the corresponding measured value in a healthy subject. This can be done by evaluating the height in comparison with the (control amount). In this case, in order to judge that the subject is suffering from schizophrenia or has a high risk of developing it in the future, the subject's amount of carbonyl compounds or AGEs may be used as an index. it can. It is known that the amount of AGEs structures such as pentosidine in vivo increases with age. Therefore, it is preferable to use a sample of a healthy person to be compared that has the same age or age as the subject.

(4) Measurement of pyridoxal content and (4) homocysteine content in a biological sample Such measurement can be performed according to a known method, preferably using blood, particularly serum or plasma. For example, it can be measured by performing an instrumental analysis such as HPLC or GC / MS using a standard product having a known concentration.

  More specifically, the test method of the present invention measures the amount of pyridoxal in a biological sample of a subject by the above method, and then compares the measured value with the corresponding amount (control amount) in a healthy person. This can be done by evaluating. In this case, the judgment that the subject is suffering from schizophrenia or has a high risk of developing it in the future can be used as an indicator that the subject's pyridoxal amount is lower than the control amount. The serum concentration of pyridoxal in healthy individuals is 6.0-40.0 ng / ml for men and 4.0-19.0 ng / ml for women.

  The pyridoxal value is linked to the homocysteine value, and when the pyridoxal value becomes low, the homocysteine value tends to become high. For this reason, when measuring the amount of pyridoxal, the amount of homocysteine may be measured together to confirm the accuracy and accuracy of the measured value of the amount of pyridoxal. In this case, if the subject's pyridoxal level is lower than the control level and the subject's homocysteine level is higher than the healthy person's homocysteine level (control level), then the subject is suffering from schizophrenia, or There is a high probability that there is a high risk of developing it in the future. The amount of homocysteine can be measured according to a conventional method. For example, it can usually be measured by performing instrumental analysis such as HPLC or GC / MS using a standard product having a known concentration. The serum concentration of homocysteine in healthy individuals is 3.7-13.5 nmol / ml.

  The inspection method of the present invention can be carried out by performing at least one of the above-mentioned (1) to (4), but these two or more, preferably 3 or more, more preferably 4 or more are combined. Can also be done. The diagnosis accuracy can be further improved by combining a plurality of combinations. Preferably, at least (1) measurement of a genetic abnormality of the GLO-I gene, or (3) measurement of a carbonyl compound or a protein modification product thereof. More preferably, it is a combination of the measurements (1) and (3), or a combination of the measurement (3) and (2) measurement of the expression level or activity of glyoxylase I. The inspection method of the present invention can also be performed in combination with conventional subjective inspection and diagnosis methods (for example, BPRS score determination according to ICD and DSM standards).

  According to the test method of the present invention, since it is possible to objectively evaluate the presence or absence of schizophrenia using a biological sample of a subject as a material, as one of auxiliary evidence when examining the presence or absence of legal responsibility Even if it is performed for purposes other than the determination of the presence or absence of legal responsibility, it can also be applied to psychological evaluation.

  Further, according to the test method of the present invention, determination of the cause and treatment method of a patient, determination of pathological condition and follow-up of the pathological condition, determination of presence or absence of therapeutic effect, prediction of prognosis, discrimination from other diseases including mental illness It can be carried out. In particular, according to the method of the present invention, patients with schizophrenia caused by carbonyl stress caused by decreased GLO-I activity can be selected. Therefore, carbonyl stress using a carbonyl scavenger, an AGEs production inhibitor or the like is used. It becomes possible to accurately select patients with schizophrenia for whom ablation therapy is effective, and to perform accurate and effective treatment.

II. The test reagent or test device for schizophrenia, the use thereof, and the present invention provide a test reagent and test device used in the test method for schizophrenia. Such reagents and devices can be appropriately selected according to the index used for the examination of schizophrenia.

(1) Test reagent in the case of using GLO-I gene abnormality as a test index For example, (1) As a test reagent in the case of using GLO-I gene abnormality as a test index, the probe described below or Primers (note that these may be labeled or may be immobilized on a solid phase). In addition to these probes or primers, the reagents include other reagents and instruments necessary for the inspection of the present invention, such as hybridization reagents, probe labels, label detection agents, and buffer solutions. You may have the form of the kit included suitably.

(1-1) Probe Insertion of adenine between positions 79 and 80 of the nucleotide sequence of the coding region of the target GLO-I gene (SEQ ID NO: 2) or mutation from adenine to cytosine at position 332 For detection, it is possible to specifically hybridize to an oligo or polynucleotide containing the base sequence at positions 79-80 or 332 of the base sequence of the coding region of the gene and detect the base sequence or base. Oligos or polynucleotides are used. Such an oligo or polynucleotide is present on the GLO-I gene (SEQ ID NO: 1) in the nucleotide sequence (positions 79 to 80 of the coding region (SEQ ID NO: 2)) or base (position 332 of the coding region (SEQ ID NO: 2)). As an oligo or polynucleotide having the above base length so as to specifically hybridize with a continuous gene region having a length of 16 to 500 bases, preferably 20 to 200 bases, more preferably 20 to 50 bases Designed.

  Here, “specifically hybridize” means normal hybridization conditions, preferably stringent hybridization conditions (for example, Sambrook et al., Molecular Cloning, Cold Spring Harbor Laboratory Press, New York, USA, In the second edition, conditions described in 1989), it means that cross-hybridization with other DNA does not occur significantly. Preferably, the oligo or polynucleotide has a base sequence complementary to the base sequence to be detected or the base sequence of the gene region containing the base. However, it is not necessary to be completely complementary if such specific hybridization is possible.

  As such an oligo or polynucleotide, specifically, a continuous sequence of 16 to 500 bases in length containing the base sequence (ac) located at positions 79-80 in the base sequence (SEQ ID NO: 2) of the coding region of the GLO-I gene An oligo or polynucleotide having a length of 16 to 500 bases that hybridizes to the oligo or polynucleotide (provided that the nucleotide symbol “t” in the sequence listing should be read as “u” when the nucleotide is RNA), Or a continuous oligo or polynucleotide having a length of 16 to 500 bases containing a base sequence located at position 332 (provided that the nucleotide symbol “t” in the sequence listing is read as “u” when the nucleotide is RNA) And oligo- or polynucleotides having a length of 16 to 500 bases.

  In order to determine whether or not a subject has schizophrenia or the possibility of its occurrence, the oligo or polynucleotide is positioned at positions 79 to 80 in the base sequence of the coding region of the GLO-I gene (SEQ ID NO: 2). Designed as an oligo or polynucleotide “probe” that specifically hybridizes to an oligonucleotide containing a region or position 332 base. These oligos or polynucleotides can be prepared based on the base sequence of the GLO-I gene (SEQ ID NO: 1) by, for example, a commercially available nucleotide synthesizer according to a conventional method.

  More preferably, as described later, the probe may be labeled with a radioactive substance, a fluorescent substance, a chemiluminescent substance, an enzyme, or the like.

  The probe (oligo or polynucleotide) can be used by immobilizing on an arbitrary solid phase. Therefore, the present invention also provides the probe (oligo or polynucleotide) as an immobilized probe (for example, a gene chip, a cDNA microarray, an oligo DNA array, a membrane filter, etc. on which the probe is immobilized). The probe can be preferably used as a DNA chip for schizophrenia testing.

  The solid phase used for immobilization is not particularly limited as long as it can immobilize oligos or polynucleotides. Examples thereof include glass plates, nylon membranes, microbeads, silicon chips, capillaries or other substrates. be able to. The immobilization of the oligo or polynucleotide to the solid phase is a method of placing a pre-synthesized oligo or polynucleotide on the solid phase, or a method of synthesizing the target oligo or polynucleotide on the solid phase. Also good. The immobilization method is well known in the art depending on the type of immobilization probe, for example, using a commercially available spotter (such as Amersham) in the case of a DNA microarray [for example, photolithographic technique (Affymetrix) In situ synthesis of oligonucleotides by inkjet technology (Rosetta Inpharmatics).

  The TaqMan PCR method (Livak KJ. Gene Anal 14, 143 (1999), Morris T et al., J Clin Microbiol 34, 2933 (1996)), which is an example of the ASO method, includes positions 79-80 or 332. An oligonucleotide having a length of about 20 bases complementary to the region is designed as a probe. The probe is labeled with a fluorescent dye at the 5 'end and a quenching substance at the 3' end and specifically hybridizes with the sample DNA, but does not emit light as it is, but is an extension reaction from the upstream of a separately added PCR primer. By this, the 5 'fluorescent dye bond is cleaved and detected by the released fluorescent dye. In Invade method [Lyamichev V. et al., Nat Biotechnol 17, 292 (1999)], which is another example of ASO method, is complementary to the sequence adjacent to the mutation site (two kinds of 3 'side and 5' side). Specific oligonucleotides are designed as probes. Detection is achieved by these two probes and a third probe that is independent of the analyte.

(1-2) Primer The present invention also provides a mutation (base insertion between positions 79 and 80, or adenine at position 332 to cytosine in the base sequence (SEQ ID NO: 2) of the coding region of the GLO-I gene. An oligonucleotide used as a primer for specifically amplifying a sequence region containing Such a primer (oligonucleotide) specifically hybridizes to a part of a continuous oligo or polynucleotide containing nucleotides 79 to 80 or 332 of the base sequence of the coding region in the GLO-I gene. The oligonucleotide is designed as an oligonucleotide having a length of 15 to 35 bases, preferably about 18 to 30 bases, for specifically amplifying the oligo or polynucleotide. The length of the oligo or polynucleotide to be amplified is appropriately set depending on the detection method used, but is generally 15 to 1000 bases, preferably 20 to 500 bases, more preferably 20 to 200 bases. It is.

  As such a primer, specifically, in the nucleotide sequence (SEQ ID NO: 2) of the coding region of the human GLO-I gene, a continuous oligo or polynucleotide having a length of 16 bases or more including a nucleotide located at positions 79-80 (however, And when the nucleotide is RNA, the base symbol “t” in the sequence listing shall be read as “u”) to specifically amplify the oligo or polynucleotide 15 to 35 bases Oligonucleotide having a length, preferably about 18 to 30 bases, and a continuous oligo or polynucleotide having a length of 16 bases or more including a nucleotide located at position 332 (provided that when the nucleotide is RNA, bases in the sequence listing) The symbol “t” shall be read as “u”) and the oligo or polynucleotide An oligonucleotide having a length of 15 to 35 bases, preferably about 18 to 30 bases, for specifically amplifying tide can be exemplified.

  These oligonucleotides can be prepared according to a conventional method by, for example, a commercially available nucleotide synthesizer based on the known base sequence (SEQ ID NO: 1) of the GLO-I gene.

(1-3) Labeled substance The above-described probe or primer of the present invention is added with an appropriate labeling substance for detecting a gene abnormality, such as a fluorescent dye, enzyme, protein, radioisotope, chemiluminescent substance, biotin, etc. Things are included.

  As the fluorescent dye used in the present invention, those generally labeled with nucleotides and used for detection and quantification of nucleic acids can be suitably used. For example, HEX (4, 7, 2 ′, 4 ′, 5 ′ , 7'-hexachloro-6-carboxylfluorescein, green fluorescent dye, fluorescein, NED (trade name, Applied Biosystems, yellow fluorescent dye), or 6-FAM (trade name, Applied Biosystems) , Yellow-green fluorescent dye), rhodamine or a derivative thereof [eg, tetramethylrhodamine (TMR)], but is not limited thereto. As a method for labeling nucleotides with a fluorescent dye, an appropriate one of known labeling methods can be used [see Nature Biotechnology, 14, 303-308 (1996)]. Commercially available fluorescent labeling kits can also be used (for example, Amersham Pharmacia, oligonucleotide ECL 3'-oligo labeling system, etc.).

  In addition, the primer of the present invention includes those to which a linker sequence for detecting a gene abnormality is added at the end. Examples of such a linker sequence include a flap (a sequence completely unrelated to the sequence near the polymorphism) added to the 5 ′ end of the oligonucleotide used in the above-described invader method.

  When PCR-direct sequence method is used as a gene abnormality measurement method, a forward primer (forward primer) for detecting a base sequence including positions 79-80 of the base sequence of the coding region of GLO-I gene (SEQ ID NO: 2) ) Is preferably an oligonucleotide having the base sequence shown in SEQ ID NO: 8, and the reverse primer (reverse primer) is preferably an oligonucleotide having the base sequence shown in SEQ ID NO: 9. be able to. In addition, as a forward primer for detecting a base sequence containing position 332 of the base sequence (SEQ ID NO: 2) of the coding region of the GLO-I gene, preferably an oligonucleotide having the base sequence shown in SEQ ID NO: 10, Moreover, as a reverse primer, the oligonucleotide which has a base sequence shown to sequence number 11 suitably can be illustrated.

  The above probes or primers (which may be labeled) can be used as reagents for detecting the presence or risk of developing schizophrenia.

(2) Reagent when GLO-I expression level is used as a test index (2) Anti-GLO-I antibody (polyclonal antibody or monoclonal antibody) is used as a reagent when GLO-I expression level is used as a test index. In addition, as a reagent when GLO-I activity is used as a diagnostic indicator, a reaction substrate of GLO-I (for example, hemithioacetal) can be mentioned. In the former case, in addition to the anti-GLO-I antibody, a reagent used for performing an immunological technique such as Western blotting, EIA method, RIA method, FIA method, chemiluminescence immunoassay, or ECLIA method ( For example, a labeled second antibody, SDS-PAGE, microplate, etc.) can also be included.

(3) Reagents or equipment when the amount of carbonyl compound or its protein modification product (AGEs) is used as a test indicator Further, (3) Reagents or equipment when the amount of carbonyl compound or AGEs is used as a test indicator are (a) Mention may be made of known concentrations of carbonyl compounds or AGEs, or (b) optionally labeled anti-AGE antibodies. Examples of the carbonyl compound and the anti-AGE antibody include those described above. In addition, when pentosidine is used as the protein modification product of (3) a carbonyl compound, the amount of pentosidine contained in the skin can be used as a test index as described above. In this case, a commercially available AGE-Reader device that can measure the amount of pentosidine in the skin can be suitably used as a schizophrenia testing device. That is, the present invention provides the AGE-Reader device as a testing device for testing schizophrenia. In this case, the device can be accompanied by a document stating that it can be used for schizophrenia testing, and the document should include a description of the diagnostic criteria for schizophrenia and how to use it for diagnosis. Can do.

(4) Reagents when pyridoxal amount is used as a test index, (5) Reagents when homocysteine amount is used as a test index, and (4) Reagents when pyridoxal amount is used as a test index, (a) A known concentration of pyridoxal or (b) a reagent for measuring pyridoxal can be mentioned. Furthermore, examples of (5) a reagent when the amount of homocysteine is used as a test index include (a) homocysteine at a known concentration, or (b) a homocysteine measuring reagent.

III. Treatment or improvement agent for schizophrenia The treatment or improvement agent for schizophrenia of the present invention is characterized by comprising a carbonyl scavenger or an AGEs production inhibitor as an active ingredient.

(1) Carbonyl scavenger The carbonyl scavenger targeted by the present invention is a carbonyl compound (for example, arabinose, GO, MGO, 3-DG, glycolaldehyde, produced from sugar, lipid or amino acid in the body by oxidative stress or the like. Dehydroascorbic acid, hydroxynonenal, malondialdehyde, acrolein, 5-hydroxymethylfurfural, formaldehyde, acetaldehyde, repric acid, furfural, and the like). The action mechanism is not particularly limited, and examples thereof include an action of suppressing the production of a carbonyl compound and an action of capturing the produced carbonyl compound and consequently reducing the amount of the body. As the carbonyl scavenger having such action, pyridoxamine or a pharmaceutically acceptable salt or ester thereof is known, and these can be widely used.

(2) AGEs production inhibitor The carbonyl compounds described above are highly reactive and non-enzymatically react with proteins in the living body to produce advanced glycation end products (AGEs) (carbonyl stress). AGEs are an assembly of a plurality of structures, such as pentosidine, croslin, pyropyridine, vesperlidine A, B, C, glyoxal-lysine dimmer (GOLD), methylglyoxal-lysine dimmer (MOLD) (above, Fluorescent substances), Nε- (carboxymethyl) lysine (CML), Nε- (carboxyethyl) lysine (CEL), argypirimidine, pyralin, imidazolone, GA-pyridine (and so on) are known.

  The AGEs production inhibitor targeted by the present invention has an action of inhibiting the production of such AGEs in the body. Here, “suppressing the production of AGEs” may be due to the action of trapping carbonyl compounds, or by inhibiting the reaction that produces protein modifications (AGEs). However, it is only necessary to finally suppress the generation of AGEs, and the action mechanism is not limited.

In addition, it can be confirmed by the following test (i) or (ii) whether the target compound has an AGEs production inhibitory effect:
(I) Using pentosidine as a representative AGEs as an index, plasma is collected from a nondiabetic renal failure dialysis patient, a test compound is added, and the amount of pentosidine produced after a certain period of time is measured.
(Ii) Phenylalanine combines with OH radicals in the presence of hydroxy radicals to generate o- or m-tyrosine. Furthermore, tyrosine reacts with NO radicals in the presence of peroxynitrite to produce nitrotyrosine. On the other hand, radicals are known to damage the kidney in vivo. Therefore, the radical scavenging ability of the test compound in the phenylalanine-radical reaction system is verified.

Furthermore, it is preferable that the target compound does not cause vitamin B6 deficiency. Whether or not the target compound causes vitamin B6 deficiency can be confirmed by the following test (a) or (b).
(A) A test compound is added to the vitamin B6 solution, and the residual amount of vitamin B6 after a predetermined time is measured.
(B) The test compound is administered to normal rats, and the presence or absence of the onset of vitamin B6 deficiency after a certain period is confirmed.

  The AGEs production inhibitor targeted by the present invention is only required to have at least an AGEs production inhibitory action, and as an example of its active ingredient, 1-substituted or unsubstituted-3-substituted or non-free or salt form A compound having a structure in which a substituent (including one derived from vitamin B6 molecule itself) that prevents binding of vitamin B6 molecule is introduced at the 4-position of the substituted-2-pyrazolin-5-one compound or a rearranged form of the compound Can be mentioned. These compounds can eventually inhibit the generation of AGEs, whether in vivo, ex vivo or / and in vitro. Specific examples of such compounds include edaravone (3-methyl-1-phenyl-2-pyrazolin-5-one) represented by the following general formulas (1) and (2) and analogs thereof. Moreover, the active ingredient of the AGEs production inhibitor targeted by the present invention includes a phenylene derivative represented by the general formula (I); and a phenylene derivative represented by the general formula (II).

(2-1) Formula (1) of edaravone and its analog free form or salt form:

Or formula (2):

[In the formula, Ra is a substituted or unsubstituted aromatic ring group, Rb, Rc and Rd are each a hydrogen atom or a monovalent organic group, or Rb and Rc combine to form a condensed ring. Or Rc and Rd together represent a divalent organic group. ]
The compound shown by.

  In formula (1) or (2), Ra represents a hydrogen atom or a substituted or unsubstituted aromatic ring (including hetero ring) group. In the “aromatic ring group”, the number of ring-constituting atoms that does not exceed 20 (including heteroatoms such as oxygen, sulfur, nitrogen, etc. may be present, but the number of these atoms exceeds 4 In particular, aryl having 6 to 10 ring carbon atoms (for example, phenyl, naphthyl) is preferred.

  As the substituent, lower alkyl, lower alkenyl, lower alkoxy, lower alkenyloxy, lower alkanoyl, halo (lower) alkyl, carboxyl, lower alkoxycarbonyl, carboxy (lower) alkyl, halo (for example, chlorine, bromine, iodine, fluorine) , Nitro, amino, lower alkylamino, di (lower) alkylamino, lower alkanoylamino, hydroxy, thiol, hydroxysulfonyl, aminosulfonyl, aryl (lower) alkanoyl, aryloxyamino, aryl, aryl (lower) alkyl, cyclo ( One or more selected from (lower) alkyl, cyclo (lower) alkenyl, cyclo (lower) alkyl (lower) alkyl, 3-7 membered heterocycle (eg oxadiazolyl, thiadiazolyl), etc. . The number of substituents is not limited, but usually does not exceed three.

  Specific examples of the substituted or unsubstituted aromatic ring group represented by Ra are as follows: phenyl, naphthyl, o-, m- or p-lower alkylphenyl (for example, o-methylphenyl, p- Methylphenyl, p-ethylphenyl), o-, m- or p-lower alkoxyphenyl (eg o-, m- or p-methoxyphenyl, o-, m- or p-ethoxyphenyl), o-, m- Or p-aminophenyl, o-, m- or p-nitrophenyl, o-, m- or p-halophenyl (eg o-, m- or p-chlorophenyl, o-, m- or p-fluorophenyl), o-, m- or p-halo (lower) alkylphenyl (eg o-, m- or p-trifluoromethylphenyl), o-, m- or p-sulfa Ylphenyl, o-, m- or p-carboxyphenyl, o-, m- or p-lower alkoxycarbonylphenyl (eg o-, m- or p-methoxycarbonylphenyl, o-, m- or p-ethoxycarbonyl) Phenyl, o-, m- or p-isopropoxycarbonylphenyl), o-, m- or p-lower alkanoylphenyl (eg o-, m- or p-acetylphenyl), di (lower) alkylphenyl (eg 3 , 4-dimethylphenyl), dihydroxyphenyl (eg 2,4-dihydroxyphenyl), 2-amino-4-carboxyphenyl, 3-amino-5-carboxyphenyl, 3-lower alkoxy-4-hydroxyphenyl (eg 3- Methoxy-4-hydroxyphenyl), 3-carboxy Such as 4-halophenyl (e.g. 3-carboxy-4-chlorophenyl).

  Rb, Rc and Rd each independently represent a hydrogen atom or a monovalent organic group. The “monovalent organic group” includes a substituted or unsubstituted hydrocarbon group, halo group, nitro group, amino group, hydroxy group, thiol group, carboxy group, carboxy (lower) alkyl group, lower alkoxycarbonyl group, formyl Group, lower alkanoyl group, lower alkylamino group, di (lower) alkylamino group, lower alkanoylamino group, aryl (lower) alkanoyl group, aryloxyamino group, sulfonic acid group, 3-7 membered heterocyclic group, etc. Is done. “Hydrocarbon group” includes a linear or cyclic, aliphatic, alicyclic or aromatic hydrocarbon group not exceeding 30 carbon atoms (preferably not exceeding 8), specifically Examples include an alkyl group, an alkenyl group, an alkynyl group, a cycloalkyl group, a cycloalkenyl group, and an aryl group. The “3- to 7-membered heterocyclic group” contains not more than 3 heteroatoms as ring-constituting atoms, and examples thereof include pyrrolidino, piperidino, morpholino, thiamorpholino and the like. The type and number of substituents are the same as those described for Ra.

  Rb and Rc can be combined to form a condensed ring. The condensed ring is preferably a 5- or 6-membered saturated carbocycle (that is, Rb + Rc = trimethylene or tetramethylene), and a substituent may be present. Rc and Rd can be combined to represent a divalent organic group. Examples of the divalent organic group include methylene type and spiro type. Examples of methylene type include phenylmethylene, phenylalkenylmethylene, quinolinylmethylene, furanylmethylene, diazolylmethylene, aminomethylene, di (lower) alkylaminomethylene, pyridylmethylene, thiophenylmethylene, etc. May optionally have a substituent. The kind and number of substituents that may be present in these condensed rings and divalent organic groups may be the same as in the case of Ra.

  In the above, the term “lower” used in connection with terms such as alkyl, alkoxy, alkanoyl, etc. is usually used to refer to a group having up to 8 carbon atoms, preferably up to 5 carbon atoms. Is done.

Specific examples of the compound (1) or (2) of the present invention are as follows:
1. 2- (3-amino-5-oxo-1-phenyl-4,5-hydro-1H-pyrazol-4-yl) -2-oxo-N-phenyl-acetamide;
2. 2- (3-amino-5-oxo-1-phenyl-4,5-hydro-1H-pyrazol-4-yl) -2-oxo-N-thiazol-2-yl-acetamide;
3. 2- (3-amino-5-oxo-1-phenyl-4,5-hydro-1H-pyrazol-4-yl) -2-oxo-acetamide;
4). 2- (3-amino-5-oxo-1-phenyl-4,5-hydro-1H-pyrazol-4-yl) -N- (3,4-dimethyl-phenyl) -4-oxo-butyramide;
5. 2- (4-amino-phenyl) -4- (2-hydroxy-ethyl) -5-methyl-2,4-hydro-pyrazol-3-one;
6). 5-amino-2-phenyl-4- (1-phenyl-1H-tetrazol-5-ylsulfanyl) -2,4-dihydro-pyrazol-3-one;
7). 3- (3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-yl) -propionic acid;
8). N- (3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-yl) -acetamide;
9. 4-[(5-hydroxy-3-methyl-1-phenyl-1Hpyrazol-4-yl) -phenyl-methyl] -5-methyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
10. 2-phenyl-3a, 4,5,6-tetrahydro-2H-cyclopentapyrazol-3-one;
11. 4-methyl-N- (3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-yl) -benzenesulfonamide;
12 N- (3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-yl) -acetamide;
13. 5-methyl-2- (3-nitro-phenyl) -4- (1-phenyl-1H-tetrazol-5-ylsulfanyl) -2,4-dihydro-pyrazol-3-one;
14 N- [5-oxo-1-phenyl-4- (1-phenyl-1H-tetrazol-5-ylsulfanyl) -4,5-dihydro-1H-pyrazol-3-yl] -benzamide;
15. 4- (hydroxy-phenyl-methyl) -2-phenyl-5-trifluoromethyl-2,4-dihydro-pyrazol-3-one;
16. 4- (1-hydroxyimino-ethyl) -2,5-diphenyl-2,4-dihydro-pyrazol-3-one;
17. 5,5′-dimethyl-2,2′-diphenyl-2,4,2 ′, 4′-tetrahydro- [4,4 ′] bipyrazole-3,3′-dione;
18. 2- (4-chloro-phenyl) -4-ethyl-5-methyl-2,4-dihydro-pyrazol-3-one;
19. 4- [4- (4-methoxy-phenyl) -thiazol-2-ylsulfanyl] -5-methyl-5-phenyl-2,4-dihydro-pyrazol-3-one;
20. 4- (2-oxo-2-phenyl-ethyl) -2-phenyl-5-propyl-2,4-dihydro-pyrazol-3-one;
21. 5-methyl-2-phenyl-4- (4-p-toluyl-thiazol-2-ylsulfanyl) -2,4-dihydro-pyrazol-3-one;
22. 2- (4-Fluoro-phenyl) -4-[[1- (4-fluoro-phenyl) -5-hydroxy-3-methyl-1H-pyrazol-4-yl]-(2-hydroxy-phenyl) -methyl ] -5-methyl-2,4-dihydro-pyrazol-3-one;
23. N- (3,4-dimethyl-phenyl) -2- (3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazol-4-yl) -2-oxo-acetamide;
24. 5- (4-chloro-benzoyl) -4,4-dihydroxy-2-phenyl-2,4-dihydro-pyrazol-3-one;
25. Sodium; sodium 4-hydroxy-3-methyl-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazole-4-sulfonate;
26. 5-methyl-4,4-di-morpholin-4-yl-2-phenyl-2,4-dihydro-pyrazol-3-one;
27. Sodium 3-benzoylamino-4-hydroxy-5-oxo-1-phenyl-4,5-dihydro-1H-pyrazole-4-sulfonate;
28. 3-methyl-1-phenyl-5-oxo-4-spiro (3oxo-2,3-dihydro-benzo [b] thiophen-2-yl) -4,5-dihydro-1H-pyrazole;
29. 4,4,5-trimethyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
30. 4,10-dimethyl-2,8,11-triphenyl-2,3,8,9-tetraza-dispiro [4.0.4.1] undeca-3,9-diene-1,7-dione;
31. 2- (2-chloro-phenyl) -4- (3-ethoxy-4-hydroxy-benzylidene) -5-methyl-2,4-dihydro-pyrazol-3-one;
32. 2- (2-chloro-phenyl) -4- (4-dimethylamino-benzylidene) -5-methyl-2,4-dihydro-pyrazol-3-one;
33. 5-methyl-4- (3-phenyl-arylidene) -2- (3-trifluoromethyl-phenyl) -2,4-dihydro-pyrazol-3-one;
34. 3- {5- [3-Methyl-5-oxo-1- (4-sulfamoyl-phenyl) -1,5-dihydro-pyrazol-4-ylidenemethyl] -furan-2-yl} -benzoic acid;
35. 4- (4-dimethylamino-benzylidene) -2- (3-fluoro-phenyl) -5-methyl-2,4-dihydro-pyrazol-3-one;
36. 3- {4- [4- (3-Chloro-4,5-dihydro-pyrazol-1-yl) -benzylidene] -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl}- benzoic acid;
37. 3- [4- (2-hydroxy-benzylidene) -5-oxo-3-phenyl-4,5-dihydro-pyrazol-1-yl] -benzoic acid;
38. 3- [1- (3-chloro-phenyl) -3-methyl-5-oxo-1,5-dihydro-pyrazol-4-ylidenemethyl] -1H-quinolin-2-one;
39. 3- {5- [3-methyl-5-oxo-1- (4-sulfamoyl-phenyl) -1,5-dihydro-pyrazol-4-ylidenemethyl] -furan-2-yl} -methyl benzoate;
40. 4- (4-Benzo [1,3] dioxol-5-ylmethylene-3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl) -methyl benzoate;
41. 4- {3-methyl-5-oxo-4- [5- (4-sulfamoyl-phenyl) -furan-2-ylmethylene] -4,5-dihydro-pyrazol-1-yl} -methylbenzoate;
42. 2- (4-chloro-phenyl) -4- (2,4-dihydroxy-benzylidene) -5-methyl-2,4-dihydro-pyrazol-3-one;
43. 2- (4-chloro-phenyl) -4- (3-hydroxy-benzylidene) -5-methyl-2,4-dihydropyrazol-3-one;
44. 4- (3,4-dihydroxy-benzylidene) -5-methyl-2-p-toluyl-2,4-dihydro-pyrazol-3-one;
45. 3- [1- (4-acetyl-phenyl) -3-methyl-5-oxo-1,5-dihydro-pyrazol-4-ylidene] -1,3-dihydro-indol-2-one;
46. 2- (4-Fluoro-phenyl) -4- (5-hydroxy-3-methyl-1-o-toluyl-1H-pyrazol-4-ylmethylene) -5-methyl-2,4-dihydro-pyrazole-3- on;
47. 2- (4-Chloro-phenyl) -4- (4-hydroxy-3-methoxy-benzylidene) -5-trifluoromethyl-2,4-dihydro-pyrazol-3-one;
48. 2- (4-ethyl-phenyl) -4- (4-hydroxy-benzylidene) -5-methyl-2,4-dihydro-pyrazol-3-one;
49. 4- [4- (4-hydroxy-benzylidene) -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl] -benzenesulfonamide;
50. 4- (5-oxo-4-thiophen-2-ylmethylene-3-trifluoromethyl-4,5-dihydro-pyrazol-1-yl) -ethyl benzoate;
51. 4- [4- (4-dimethylamino-benzylidene) -5-oxo-3-trifluoromethyl-4,5-dihydro-pyrazol-1-yl] -benzenesulfonamide;
52. 4-isopropylidene-5-methyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
53. 4- (4-hydroxy-benzylidene) -2-phenyl-5-trifluoromethyl-2,4-dihydro-pyrazol-3-one;
54. 4- (2,4-dihydroxy-benzylidene) -2- (3,4-dimethyl-phenyl) -5-methyl-2,4-dihydro-pyrazol-3-one;
55. 3- [4- (3-Ethoxy-4-hydroxy-benzylidene) -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl] -benzoic acid;
56. 4- [4- (3,5-di-tert-butyl-4-hydroxy-benzylidene) -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl] -benzoic acid;
57. 3- [3- (4-hydroxy-3-methoxy-benzylidene) -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl] -benzoic acid;
58. 3- [3-hydroxy-4- (4-hydroxy-3-methoxy-benzylidene) -5-oxo-pyrazolidin-1-yl] -benzoic acid;
59. 4- (3-hydroxy-2,4-dimethoxy-benzylidene) -5-methyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
60. 4- [4- (4-Hydroxy-3-methoxy-benzylidene) -3-methyl-5-oxo-pyrazolidin-1-yl] -isopropyl benzoate;
61. 2-chloro-5- [4- (2-chloro-4-hydroxy-5-methoxy-benzylidene) -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl] -benzoic acid;
62. 4- [4- (4-hydroxy-benzylidene) -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl] -ethyl benzoate;
63. 4- [4- (4-hydroxy-benzylidene) -5-oxo-3-trifluoromethyl-4,5-dihydro-pyrazol-1-yl] -ethyl benzoate;
64. 4- [4- (4-hydroxy-benzylidene) -3-methyl-5-oxo-4,5-dihydro-pyrazol-1-yl] -benzoic acid;
65. 4-dimethylaminomethylene-5-methyl-2-phenyl-2,4-dihumanlo-pyrazol-3-one;
66. 4- (5-hydroxy-3-methyl-1-phenyl-1H-pyrazol-4-ylmethylene) -5-methyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
67. 4- (4-chloro-benzylidene) -5-methyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
68. 1- (5-hydroxy-3-methyl-1-phenyl-1H-pyrazol-4-yl) -6-methyl-1,3-dihydro-furo [3,4-c] pyridin-7-ol;
69. 1- (5-Hydroxy-3-methyl-1-phenyl-1H-pyrazol-4-yl) -6-methyl-1,3-dihydro-furo [3,4-c] pyridin-7-ol (hydrochloride salt) );
70. 4- (4-hydroxy-benzylidene) -5-methyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
71. 2- (3-chloro-phenyl) -4- (4-hydroxy-benzylidene) -5-methyl-2,4-dihydropyrazol-3-one;
72. 4- (4-benzyloxy-benzylidene) -5-methyl-2-phenyl-2,4-dihydropyrazol-3-one;
73. 2- (3-chloro-phenyl) -5-methyl-2H-pyrazole-3,4-dione 4-oxime;
74. Ethyl 5- (5-oxo-1,3-diphenyl-1,5-dihydro-pyrazole-4-ylidene) -4-phenyl-4,5-dihydro- [1,3,4] thiazole-2-carboxylate ;
75. 4- [1,3] dithiolane-2-ylidene-5-methyl-2-phenyl-2,4-dihydro-pyrazol-3-one;
76. 5- (4-chloro-phenylsulfanylmethyl) -2-phenyl-4- [N ′-(3-trifluoromethyl-phenyl) -hydrazino] -2,4-dihydro-pyrazol-3-one;
77. 4- (5-benzoyl-3-phenyl-3H- [1,3,4] thiadiazol-2-ylidene) -2,5-diphenyl-2,4-dihydro-pyrazol-3-one;
78. Phosphoric acid mono- [5-hydroxy-6-methyl-4- (3-methyl-5-oxo-1-phenyl-1,5-dihydro-pyrazol-4-ylidenemethyl) -pyridin-3-ylmethyl] ester ;
79.3-Methyl-1-phenyl-2-pyrazolin-5-one.

  In addition, you may use these compounds (1) or (2) in a free form or a salt form. Examples of the salt form generally include pharmaceutically acceptable salts such as salts with inorganic bases and organic bases, acid addition salts such as inorganic acids, organic acids, basic or acidic amino acids, and the like. Examples of the salt with an inorganic base include alkali metal (sodium, potassium, etc.) salts, alkaline earth metal (calcium, magnesium, etc.) salts, aluminum salts, ammonium salts and the like. Examples of salts with organic bases include primary amines (such as ethanolamine), secondary amines (such as diethylamine, diethanolamine, dicyclohexylamine, N, N′-dibenzylethylenediamine), and tertiary amines (trimethylamine, triethylamine). , Pyridine, picoline, triethanolamine, etc.). Examples of salts with inorganic acids include salts with hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid, etc. Examples of salts with organic acids include formic acid, acetic acid, lactic acid, trifluoroacetic acid, fumaric acid, Examples thereof include salts with oxalic acid, tartaric acid, maleic acid, benzoic acid, citric acid, succinic acid, malic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid and the like. Furthermore, examples of salts with basic amino acids include salts with arginine, lysine, ornithine, and examples of salts with acidic amino acids include salts with aspartic acid, glutamic acid, and the like.

  These compounds (1) and (2), their pharmacologically acceptable salts and their pharmacologically acceptable esters can be prepared using, for example, known compounds as starting materials according to the method described in International Publication WO2005 / 054205 Pamphlet. Can be manufactured. In addition, the manufacturing method of these compounds described in the said publication | presentation gazette is used as content of this specification as it is.

(2-2) Phenylene derivatives (I)
Formula (I)

[Wherein, A represents the following general formula (A1), (A2) or (A3)

B represents a 1H-tetrazol-5-yl group or a 2,4-dioxothiazolidin-5-yl group, X represents a methylene, oxygen atom or sulfur atom, and Y represents a single bond. Or a C6-10 arylene group, R ^1A represents a hydrogen atom or a C 1-6 alkyl group, R ^2A and R ^3A are the same or different and represent a hydrogen atom, a carboxyl group or a C 1-6 alkyl group, R ^4A , R ^5A and R ^6A are the same or different and each represents a hydrogen atom or a C 1-6 alkyl group, and R ^7A represents a C 1-10 alkylcarbonyl group. However, when A is (A2), B represents a 2,4-dioxothiazolidin-5-yl group. ] The compound represented by this.

Here, the “C 1-6 alkyl group” is a linear or branched alkyl group having 1 to 6 carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, s -Butyl, t-butyl, pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, hexyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,2-dimethylbutyl, 2, 3-dimethylbutyl, 3,3-dimethylbutyl, 1-ethylbutyl, 2-ethylbutyl, 1,1,2-trimethylpropyl, 1,2,2-to It may be mentioned methyl propyl. R ^1A is preferably a propyl group. R ^2A and R ^3A are preferably a C 1-3 alkyl group. R ^4A is preferably a C 1-3 alkyl group, and more preferably an ethyl group or a propyl group. R ^5A and R ^6A are preferably a C 1-3 alkyl group, and more preferably a methyl group.

  The “C6-10 arylene group” is a divalent aromatic hydrocarbon group having 6 to 10 carbon atoms, and examples thereof include phenylene, indenylene, and naphthylene groups. Y is preferably a phenylene group. is there.

The “C 1-10 alkylcarbonyl group” is a group in which a linear or branched alkyl group having 1 to 10 carbon atoms is bonded to a carbonyl group. For example, acetyl, propionyl, butyryl, isobutyryl And alkylcarbonyl groups such as pentanoyl, pivaloyl, valeryl, isovaleryl, octanoyl, nonanoyl, decanoyl, 3-methylnonanoyl, 8-methinolenanoyl, 3-ethyloctanoyl, 3,7-dimethyloctanoyl and undecanoyl R ^7A is preferably a C 4-7 alkylcarbonyl group, more preferably an octanoyl group.

  In the present invention, A is preferably a group represented by (A1) or (A3), and more preferably a group represented by (A3).

  In the present invention, B is preferably a 1H-tetrazol-5-yl group.

  In the present invention, X is preferably methylene or an oxygen atom, and more preferably methylene.

  In the present invention, Y is preferably a single bond or phenylene, and more preferably phenylene.

In the present invention, R ^1A is preferably a propyl group or a hydrogen atom, and more preferably a propyl group.

In the present invention, R ^2A and R ^3A are the same or different, and are preferably a hydrogen atom, a carboxyl group, or a pharmacologically acceptable ester thereof.

In the present invention, R ^4A is preferably an ethyl group or a propyl group.

In the present invention, R ^4A and R ^6A are preferably a methyl group or a hydrogen atom.

In the present invention, R ^7A is preferably a C4-7 alkylcarbonyl group, and more preferably an octanoyl group.

  When the phenylene derivative (I) of the present invention has a basic group, it can be converted into an acid addition salt according to a conventional method. Such salts include, for example, hydrohalic acid salts such as hydrofluoric acid, hydrochloric acid, hydrobromic acid and hydroiodic acid; inorganics such as nitrates, perchlorates, sulfates and phosphates. Acid salts; salts of lower alkanesulfonic acids such as methanesulfonic acid, trifluoromethanesulfonic acid, ethanesulfonic acid; salts of arylsulfonic acids such as benzenesulfonic acid and P-toluenesulfonic acid; such as glutamic acid and aspartic acid Amino acid salts; examples include salts of carboxylic acids such as acetic acid, fumaric acid, tartaric acid, succinic acid, maleic acid, malic acid, succinic acid, benzoic acid, mandelic acid, ascorbic acid, lactic acid, gluconic acid, and citric acid. . Preferred is a salt of hydrohalic acid.

  Furthermore, when the phenylene derivative (I) has a carboxyl group, it can be converted into a metal salt according to a conventional method. Examples of such salts include alkali metal salts such as lithium, sodium and potassium; alkaline earth metal salts such as calcium, barium and magnesium; and aluminum salts. Alkali metal salts are preferred.

  The phenylene derivative (I) of the present invention can be converted into a pharmacologically acceptable ester according to a conventional method. Such ester is not particularly limited as long as it is medically used and pharmacologically acceptable.

  Examples of the ester residue of the ester of the phenylene derivative (I) of the present invention include, for example, a linear or branched alkyl group having 1 to 6 carbon atoms (the alkyl group is substituted with a trialkylsilyl group). An aralkyl group having 7 to 19 carbon atoms, a linear or branched chain alkanoyloxy having 1 to 6 carbon atoms, and a straight chain having 1 to 5 carbon atoms. Or a linear or branched alkyl group having 1 to 5 carbon atoms substituted with a branched alkyl group, a linear or branched alkyloxycarbonyloxy having 1 to 6 carbon atoms Alkyl group, linear or branched alkyl group having 1 to 5 carbon atoms substituted by cycloalkylcarbonyloxy having 5 to 7 carbon atoms, having 5 to 7 carbon atoms A linear or branched alkyl group having 1 to 5 carbon atoms substituted by cycloalkyloxycarbonyloxy and arylcarbonyloxy having 6 to 10 carbon atoms substituted with 1 to 5 carbon atoms A linear or branched alkyl group having 1 to 5 carbon atoms substituted by an aryloxycarbonyloxy having 6 to 10 carbon atoms, 5-position And (2-oxo-1,3-dioxolen-4-yl) methyl group having a linear or branched alkyl having 1 to 6 carbon atoms as a substituent.

  Here, examples of the linear or branched alkyl group having 1 to 6 carbon atoms include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, s-butyl, t-butyl, pentyl, 1- Methylbutyl, 2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, hexyl, 1-methylpentyl, 2-methylpentyl, 3 -Methylpentyl, 4-methylpentyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl, 3,3-dimethylbutyl 1-ethylbutyl, 2-ethylbutyl, 1,1,2-trimethylpropyl or 1,2,2-trimethylpropyl. Preferably, it is a linear or branched alkyl group having 1 to 4 carbon atoms, more preferably methyl, ethyl, propyl, isopropyl, butyl or isobutyl, and most preferably Is methyl or ethyl.

  Examples of the aralkyl group having 7 to 19 carbon atoms include benzyl, phenethyl, 3-phenylpropyl, 4-phenylbutyl, 1-naphthylmethyl, 2-naphthylmethyl and diphenylmethyl, preferably benzyl It is.

  Examples of the cycloalkyl group having 5 to 7 carbon atoms include cyclopentyl, cyclohexyl and cycloheptyl, and cyclohexyl is preferred.

  Examples of the aryl group having 6 to 10 carbon atoms include phenyl and naphthyl, and is preferably phenyl.

  Specific examples of suitable ester residues include, for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, benzyl, acetoxymethyl, 1- (acetoxy) ethyl, propionyloxymethyl, 1- (propionyloxy) Ethyl, butyryloxymethyl, 1- (butyryloxy) ethyl, isobutyryloxymethyl, 1- (isobutyryloxy) ethyl, valeryloxymethyl, 1- (valeryloxy) ethyl, isovaleryloxymethyl, 1- (Isovaleryloxy) ethyl, pivaloyloxymethyl, 1- (pivaloyloxy) ethyl, methoxycarbonyloxymethyl, 1- (methoxycarbonyloxy) ethyl, ethoxycarbonyloxymethyl, 1- (ethoxycarbonyloxy) ethyl, propoxy Carbonyloxymethyl, 1- ( Propoxycarbonyloxy) ethyl, isopropoxycarbonyloxymethyl, 1- (isopropoxycanolenyloxy) ethyl, butoxycarbonyloxymethyl, 1- (butoxycarbonyloxy) ethyl, isobutoxycarbonyloxymethyl, 1- (isobutoxycarbonyl) Oxy) ethyl, t-butoxycarbonyloxymethyl, 1- (t-butoxycarbonyloxy) ethyl, cyclopentanecarbonyloxymethyl, 1- (cyclopentanecarbonyloxy) ethyl, cyclohexanecarbonyloxymethyl, 1- (cyclohexanecarbonyloxy) Ethyl, cyclopentyloxycarbonyloxymethyl, 1- (cyclopentyloxycarbonyloxy) ethyl, cyclohexyloxycarbonyloxymethyl, 1- (cyclohexyloxycarbonyl) Bonyloxy) ethyl, benzoyloxymethyl, 1- (benzoyloxy) ethyl, phenoxycarbonyloxymethyl, 1- (phenoxycarbonyloxy) ethyl, (5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl 2-trimethylsilylethyl or phthalidyl, more preferably (5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl, pivaloyloxymethyl or 1- (isopropoxycarbonyloxy) Ethyl.

  In addition, when the phenylene derivative represented by the above general formula (I), a salt or an ester thereof forms a solvate (for example, a hydrate), all of these solvates are also included in the present invention.

  Furthermore, the present invention includes all phenylene derivatives (I) expressed in the above and converted into salts or esters thereof (so-called prodrugs such as amide derivatives).

  Specific examples of the phenylene derivative represented by the above general formula (1) of the present invention or a pharmacologically acceptable salt or ester thereof include the compounds exemplified below. However, the present invention is not limited to the following exemplary compounds.

  In Tables 1 to 3 below, “Me” represents a methyl group, “Et” represents an ethyl group, “Pr” represents a propyl group, “Bu” represents a butyl group, and “t-Bu” represents t- “Hex” represents a hexyl group, “-Ph-” represents a phenylene group, “DMDO” represents a (5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl group, “PHT” represents a phthalidyl group, “Tez” represents a 1H-tetrazolyl-5-yl group, “Tzd” represents a 2,4-dioxothiazolidin-5-yl group, and “−” represents a single bond.

  In the above table, preferred compounds (I) are 1-1, 1-8, 1-10, 1-19, 1-20, 1-25, 1-29, 1-30, 2-2, 2- 3, 2-12, 2-18, 3-3-3, 3-4, 3-5, 3-6, 3-7, 3-8, 3-9, 3-10, 3-13, 3-25 , 3-29, 3-30, more preferably 1-1, 2-2, 3-7, 3-6, 3-3, 3-9, 3-13, 3-25 or 3- 30.

  These phenylene derivatives (I), their pharmacologically acceptable salts and their pharmacologically acceptable esters are produced, for example, using known compounds as starting materials according to the method described in the pamphlet of International Publication WO2005 / 030737. can do. In addition, the manufacturing method of the phenylene derivative (I) described in the said publication | presentation gazette is used as it is as the content of this specification as it is.

(2-3) Phenylene derivatives (II)
Formula (II)

[Wherein, A represents the following general formula (A4), (A5), (A6) or (A7)

Represents a group represented by
B represents a 1H-tetrazol-5-yl group or a 2,4-dioxo-1,3-thiazolidin-5-yl group;
X represents a single bond or a C6-10 arylene group;
Y1 represents carbonyl, sulfonyl or a single bond;
Y2 represents a C1-6 alkylene group or a single bond;
Y3 represents methylene, carbonyl or a single bond;
Y4 represents a methine or nitrogen atom;
Y5 represents methylene, carbonyl or a single bond;
R ¹ is
A C1-8 alkyl group (the C1-8 alkyl group may be substituted with 1 to 3 groups selected from the following substituent group α);
A C6-14 aryl group (the C6-14 aryl group may be substituted with 1 to 5 groups selected from the following substituent group β);
A C6-14 aryl-C1-6 alkyl group (the C6-14 aryl-C1-6 alkyl group may be substituted with 1 to 5 groups selected from the following substituent group β);
A 4- to 10-membered heterocyclic group containing 1 to 3 heteroatoms which are the same or different and selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocyclic group is a group selected from the following substituent group β) 1 to 5 may be substituted),
A 4- to 10-membered heterocyclic-C1-6 alkyl group containing 1 to 3 heteroatoms identical or different selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocycle-C1-6 alkyl group is: 1 to 5 groups may be substituted with a group selected from substituent group β),
A C3-7 cycloalkyl group (the C3-7 cycloalkyl group may be substituted with 1 to 5 groups selected from the following substituent group β);
A C3-7 cycloalkyl-C1-6 alkyl group (the C3-7 cycloalkyl-C1-6 alkyl group may be substituted with 1 to 5 groups selected from the following substituent group β);
C6-14 aryl-carbonyl-C1-6 alkyl group (the C6-14 aryl-carbonyl-Cl-6 alkyl group may be substituted with 1 to 5 groups selected from the following substituent group β) Or
A C1-6 aliphatic acyl group (the C1-6 aliphatic acyl group may be substituted with 1 to 5 groups selected from the following substituent group β);
R ² represents a hydrogen atom, a group selected from the following substituent group β, a carboxyl group, or a group represented by —C (O) NR ¹³ R ¹⁴ ;
R ³ is
Hydrogen atom,
A C1-8 alkyl group (the C1-8 alkyl group may be substituted with 1 to 3 groups selected from the following substituent group α);
A C6-14 aryl group (the C6-14 aryl group may be substituted with 1 to 5 groups selected from the following substituent group β), or a nitrogen atom, an oxygen atom and a sulfur atom. A 4- to 10-membered heterocyclic group containing the same or different 1 to 3 heteroatoms (the heterocyclic group may be substituted with 1 to 5 groups selected from the following substituent group β) Show;
R ⁴ and R ⁵ are the same or different and each is a hydrogen atom or a C 6-14 aryl group (the C 6-14 aryl group may be substituted with 1 to 5 groups selected from the following substituent group β). Good);
R ⁶ and R ⁷ together form a C 3-7 cycloalkane, or are the same or different and each is a hydrogen atom or a C 1-8 alkyl group (the C 1-8 alkyl group is selected from the following substituent group α). 1 to 3 may be substituted with a selected group);
R ⁸ and R ⁹ are the same or different and each represents a hydrogen atom, a group selected from the following substituent group β, a carboxyl group, or a group represented by —C (O) NR ¹³ R ¹⁴ ;
R ¹⁰ is
A hydrogen atom,
-C1-8 alkyl group (the C1-8 alkyl group may be substituted by 1 to 3 groups selected from the following substituent group α);
-C6-14 aryl group (the C6-14 aryl group may be substituted with 1 to 5 groups selected from the following substituent group β), or-selected from a nitrogen atom, an oxygen atom and a sulfur atom 4- to 10-membered heterocyclic group containing 1 to 3 heteroatoms which are the same or different (the heterocyclic group may be substituted with 1 to 5 groups selected from the following substituent group β) );
R ¹¹ and R ¹² are the same or different and each represents a hydrogen atom, a group selected from the following substituent group β, a carboxyl group, or a group represented by —C (O) NR ¹³ R ¹⁴ ;
R ¹³ and R ¹⁴ together form a 4- to 10-membered nitrogen-containing heterocycle (the nitrogen-containing heterocycle may be substituted with 1 to 5 groups selected from the following substituent group β). Or are the same or different from each other, and each is a hydrogen atom or a C1-8 alkyl group (the C1-8 alkyl group may be substituted with 1 to 3 groups selected from the following substituent group α). Show.

However,
(1) When R ¹ is a C 1-8 alkyl group and Y 1 is a single bond, B is a 2,4-dioxo-1,3-thiazolidin-5-yl group,
(2) When Y2 is methylene, R ⁶ and R ⁷ are not both hydrogen atoms,
(3) When R ¹ is an unsubstituted C 1-8 alkyl group, Y 1 is not carbonyl,
(4) When R ¹ is an unsubstituted C 1-6 aliphatic acyl group, Y 1 is not a single bond.
(Substituent group α)
A halogen atom, a C1-6 alkylthio group, a C1-6 aliphatic acyl group, a C1-6 alkylsulfonyl group, a C1-6 alkoxy group, a cyano group and a nitro group;
(Substituent group β)
Halogen atom, C1-6 alkyl group, halo C1-6 alkyl group, C1-6 alkoxy group, C1-6 alkylthio group, C1-6 alkylsulfonyl group, C1-6 aliphatic acyl group, oxo group, cyano group, nitro 1 to 3 selected from the group consisting of a nitrogen atom, an oxygen atom and a sulfur atom, a C3-7 cycloalkyl group, a C6-14 aryl group, a C1-6 alkoxyimino group, a C6-14 aryl-carbol group 4- to 10-membered heterocyclic group containing 1 heteroatom. ]
A compound represented by

  In the phenylene derivative represented by the general formula (II), the “C 6-10 arylene group” is a divalent aromatic hydrocarbon group having 6 to 10 carbon atoms, such as 1,2-phenylene. 1,3-phenylene, 1,4-phenylene, 1,2-naphthylene, 1,3-naphthylene, 1,4-naphthylene, 1,5-naphthylene, 1,6-naphthylene, 1,7-naphthylene, , 8-naphthylene, 2,3-naphthylene, 2,4-naphthylene and the like.

  X is preferably phenylene.

  In the phenylene derivative (II), the “C 1-6 alkylene group” is a linear or branched alkylene group having 1 to 6 carbon atoms such as methylene, ethylene, propylene, trimethylene, 1 -Ethyl-1,2-ethylene, 1-propyl-1,2-ethylene, 1-isopropyl-1,2-ethylene, 1-butyl-1,2-ethylene, 1,2-dimethyl-1,2-ethylene , Tetramethylene, pentamethylene, hexamethylene and the like.

  Y2 is preferably a C1-3 alkylene group, and more preferably methylene or ethylene.

  In the phenylene derivative (II), the “C 1-8 alkyl group” is a linear or branched alkyl group having 1 to 8 carbon atoms, such as methyl, ethyl, propyl, isopropyl, Butyl, isobutyl, sec-butyl, t-butyl, pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethyl Propyl, hexyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,2-dimethyl Butyl, 2,3-dimethylbutyl, 3,3-dimethylbutyl, 1-ethylbutyl, 2-ethylbutyl, 1,1,2-trimethyl Lopyl, 1,2,2-trimethylpropyl, heptyl, 1-methylpentyl, 2-methylpentyl, n-heptyl, isoheptyl, sec-heptyl, tert-heptyl, n-octyl, isooctyl, tert-octyl, 2-methyl Examples include hexyl and 2-ethylhexyl.

In R ¹
(1) When Y1 is a sulfonyl group, it is preferably a C1-6 alkyl group, more preferably methyl, ethyl, propyl, butyl, pentyl, hexyl, and further preferably methyl or butyl.
(2) When Y1 is a single bond or a carbonyl group, it is preferably a C4-8 alkyl group, more preferably butyl, pentyl, hexyl, heptyl, octyl, and even more preferably pentyl.

R ³ is preferably a C 1-5 alkyl group, and more preferably methyl, ethyl, propyl, butyl, or pentyl.

R ⁶ and R ⁷ are preferably a C 1-3 alkyl group, and more preferably methyl, ethyl, or propyl.

R ¹⁰ is preferably a C 1-4 alkyl group, and more preferably methyl, ethyl, propyl, or butyl.

R ¹³ and R ¹⁴ are preferably a C 1-3 alkyl group, and more preferably methyl or ethyl.

  In the phenylene derivative (II), the “C 1-6 alkyl group” is a linear or branched alkyl group having 1 to 6 carbon atoms, for example, methyl, ethyl, propyl, isopropyl, butyl , Isobutyl, sec-butyl, t-butyl, pentyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl Hexyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,2-dimethylbutyl 2,3-dimethylbutyl, 3,3-dimethylbutyl, 1-ethylbutyl, 2-ethylbutyl, 1,1,2-trimethylpropyl Pills, 1,2,2-methylpropyl, and the like.

  The substituent β is preferably a C 1-3 alkyl group, and more preferably methyl or ethyl.

  In the phenylene derivative (II), the “C 6-14 aryl group” is an aromatic hydrocarbon group having 6 to 14 carbon atoms, and examples thereof include phenyl, naphthyl, anthryl, phenanthryl, acenaphthylenyl and the like.

R ¹ is preferably a C 6-10 aryl group, more preferably phenyl or naphthyl.

R ³ is preferably a C 6-10 aryl group, more preferably phenyl or naphthyl.

R ⁴ and R ⁵ are preferably a C 6-10 aryl group, and more preferably phenyl.

R ¹⁰ is preferably a C 6-10 aryl group, and more preferably phenyl.

  The substituent β is preferably a C6-10 aryl group, and more preferably phenyl.

  In the phenylene derivative (II), the “C 6-14 aryl-C 1-6 alkyl group” is a group in which the above “C 6-14 aryl group” is bonded to the above “C 1-6 alkyl group”. Benzyl, 1-phenethyl, 2-phenethyl, 3-phenylpropyl, 4-phenylbutyl, 5-phenylpentyl, 5-phenylhexyl, 1-naphthylmethyl, 2-naphthylethyl, anthrylmethyl, phenanthrylmethyl, asena Examples include butylenylmethyl.

R ¹ is preferably a C 6-10 aryl-C 1-3 alkyl group, more preferably benzyl, 1-phenethyl, 2-phenethyl, 3-phenylpropyl, 1-naphthylmethyl, 2-naphthylmethyl, 2- (1-naphthyl) ethyl, 2- (2-naphthylethyl).

  In the phenylene derivative (II), examples of the “4- to 10-membered heterocyclic group containing 1 to 3 heteroatoms selected from the nitrogen atom, oxygen atom and sulfur atom” include pyrrolyl, furyl and thienyl. , Pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, triazolyl, tetrazolyl, pyranyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, azetidinyl, pyrrolidinyl, pyrrolidinyl, pyrrolidinyl, pyrrolidinyl, pyrrolidinyl, pyrrolidinyl Imidazolinyl, pyrazolidinyl, pyrazolinyl, piperidyl, piperazinyl, morpholinyl, thiomorpholyl, perhydroazevinyl, perhydroazoseal, 1,4,5,6-the La tetrahydropyrimidinyl, 1,2,3,6 etc. tetrahydropyridyl, the unsaturated heterocyclic group is partially if <can be completely reduced group. Indolyl, indolinyl, benzofuranyl, benzothienyl, benzimidazolyl, benzisoxazolyl, benzisoxazolyl, benzothiazolyl, benzisothiazolyl, quinolyl, isoquinolyl, quinazolinyl, quinoxalinyl, benzoxiadiazolyl, benzothiadiazolyl , Pyrrolopyrrolyl, pyrrolooxazolyl, pyrrolothiazolyl, pyrrolopyridyl, furopyrrolyl, furopyridyl, thienopyrrolyl, thienopyridyl, imidazopyrrolyl, imidazolimidazolyl, imidazoloxazolyl, imidazoisoxazolyl, imidazolothiazolyl, imidazoisothiazolyl , Imidazopyridazinyl, imidazopyrimidinyl, imidazopyrazinyl, oxazooxazolyl, oxazoisoxazolyl, oxazo A group in which the above unsaturated heterocycles are condensed, such as azolyl, oxazoisothiazolyl, oxazopyridyl, thiazooxazolyl, thiazoisoxazolyl, thiazothiazolyl, thiazoisothiazolyl, thiazopyridyl, etc. A group condensed with a saturated heterocyclic ring may be used.

In R ¹ , a 5- or 6-membered heterocyclic group containing 1 or 2 heteroatoms which are the same or different, preferably selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocycle is condensed with a benzene ring) More preferred are pyridyl, oxazolyl, thiazolyl and quinolyl.

In R ³ , a 5- or 6-membered heterocyclic group containing 1 or 2 heteroatoms, which are the same or different, preferably selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocycle is condensed with a benzene ring) And may be pyridyl or quinolyl.

In R ¹⁰ , a 5- or 6-membered heterocyclic group containing 1 or 2 heteroatoms, which are the same or different, preferably selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocycle is condensed with a benzene ring). And may be pyridyl or quinolyl.

  In the substituent β, a 5- or 6-membered heterocyclic group containing 1 or 2 heteroatoms which are the same or different, preferably selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocycle is condensed with a benzene ring) And may be imidazolyl, thiazolyl, oxazolyl, oxadiazolyl, or pyridyl. The heterocyclic group may be substituted with 1 to 3 of the above “C 1-6 alkyl group”.

  In the phenylene derivative (II), the “4- to 10-membered nitrogen-containing heterocycle” is the above-mentioned “4 to 10 containing 1 to 3 heteroatoms which are the same or different and are selected from nitrogen, oxygen and sulfur atoms”. Among the rings constituting the “membered heterocyclic group”, the ring contains at least one nitrogen and may further contain the same or different 1 or 2 heteroatoms selected from a nitrogen atom, an oxygen atom and a sulfur atom For example, unsaturated heterocyclic rings such as pyrrole, pyrazole, imidazole, triazole, tetrazole; azetidine, pyrrolidine, pyrroline, imidazolidine, imidazoline, pyrazolidine, pyrazoline, piperidine, piperazine, morpholine, thiomorpholine, perhydroazepine, par Hydroazocine, 1,4,5,6-tetrahydropyri Jin, 1,2,3,6 etc. tetrahydropyridine, the unsaturated heterocyclic ring may be partially or fully reduced ring. Indole, indoline, benzimidazole, pyrrolopyrrole, pyrrolooxazole, pyrrolothiazole, pyrrolopyridine, furopyrrole, thienopyrrole, imidazopyrrole, imidazoimidazole, imidazoxazole, imidazoisoxazole, imidazothiazole, imidazoisothiazole, imidazopyridine, imidazopyridine A ring in which the unsaturated heterocyclic rings are condensed or a ring in which the benzene ring and the unsaturated heterocyclic ring are condensed, such as pyridazine, imidazopyrimidine, and imidazopyrazine, may be used.

R ¹³ and R ¹⁴ are preferably imidazole, pyrrolidine, piperidine, morpholine and indole ring, and more preferably pyrrolidine, piperidine and morpholine ring.

  In the phenylene derivative (II), “a 4- to 10-membered heterocyclic-C 1-6 alkyl group containing 1 to 3 heteroatoms which are the same or different and selected from a nitrogen atom, an oxygen atom and a sulfur atom” A group in which the “4- to 10-membered heterocyclic group containing 1 to 3 heteroatoms identical or different selected from a nitrogen atom, an oxygen atom and a sulfur atom” is bonded to the “C1-6 alkyl group” For example, pyrrolylmethyl, furylmethyl, thienylmethyl, thienylethyl, thienylpropyl, pyrazolylmethyl, imidazolylmethyl, oxazolylmethyl, isoxazolylmethyl, thiazolylmethyl, isothiazolylmethyl, oxadiazolylmethyl, thiadiazoyl Rylmethyl, triazolylmethyl, tetrazolylmethyl, pyranylmethyl, pyridylmethyl, pyriyl Dinylmethyl, pyrimidinylmethyl, pyrazinylmethyl, azepinylmethyl, azosinylmethyl, azetidinylmethyl, pyrrolidinylmethyl, pyrrolinylmethyl, imidazolidinylmethyl, imidazolinylmethyl, pyrazolidinylmethyl, pyrazolinylmethyl, piperidylmethyl, pipepe Razinylmethyl, morpholinylmethyl, thiomorpholinylmethyl, perhydroazepinylmethyl, perhydroazosinylmethyl, 1,4,5,6-tetrahydropyrimidinylmethyl, 1,2,3,6-tetrahydropyridyl Methyl, indolylmethyl, indolinylmethyl, benzofuranylmethyl, benzothienylmethyl, benzimidazolylmethyl, benzisoxazolylmethyl, benzisoxazolylmethyl, benzothiazolylmethyl, benzisothi Azolylmethyl, quinolylmethyl, isoquinolylmethyl, quinazolinylmethyl, quinoxalinylmethyl, benzoxiadiazolylmethyl, benzothiadiazolylmethyl, pyrrolopyrrolylmethyl, pyrrolooxazolylmethyl, pyrrolothiazolylmethyl, pyrrolo Pyridylmethyl, furopyrrolylmethyl, furopyridylmethyl, thienopyrrolylmethyl, thienopyridylmethyl, imidazopyrrolylmethyl, imidazoimidazolylmethyl, imidazooxazolylmethyl, imidazoisoxazolylmethyl, imidazothiazolylmethyl, imidazo Isothiazolylmethyl, imidazopyridylmethyl, imidazopyridazinylmethyl, imidazopyrimidinylmethyl, imidazopyrazinylmethyl, oxazooxazolylmethyl, oxazoisoxazolylmethyl, oxa Zothiazolylmethyl, oxazoisothiazolylmethyl, oxazopyridylmethyl, thiazooxazolylmethyl, thiazoisoxazolylmethyl, thiazothiazolylmethyl, thiazoisothiazolylmethyl, thiazopyridylmethyl Etc.

In R ¹ , a 5- or 6-membered heterocyclic-C 1-3 alkyl group containing 1 or 2 heteroatoms, which are the same or different, preferably selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocycle is And may be condensed with a benzene ring), and more preferably oxazolylmethyl, thiazolylmethyl, pyridylmethyl, quinolylmethyl, thienylethyl, and thienylpropyl.

  In the phenylene derivative (II), the “C 3-7 cycloalkyl group” is a cyclic alkyl group having 3 or 7 carbon atoms, and examples thereof include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and the like.

R ¹ is preferably a C 5-6 cycloalkyl group, and more preferably cyclopentyl or cyclohexyl.

  The substituent β is preferably a C 4-6 cycloalkyl group, and most preferably cyclopentyl or cyclohexyl.

  In the phenylene derivative (II), the “C 3-7 cycloalkyl-C 1-6 alkyl group” is a group in which the above “C 3-7 cycloalkyl group” is bonded to the above “C 1-6 alkyl group”. , Cyclopropylmethyl, 2-cyclopropylethyl, cyclobutylmethyl, 2-cyclobutylethyl, cyclopentylmethyl, 2-cyclopentylethyl, 3-cyclopentylpropyl, 4-cyclopentylbutyl, 5-cyclopentylpentyl, 6-cyclopentylhexyl, cyclohexyl Examples include methyl, 2-cyclohexylethyl, 3-cyclohexylpropyl, 4-cyclohexylbutyl, 5-cyclohexylpentyl, 6-cyclohexylhexyl, cycloheptylmethyl, 2-cycloheptylethyl and the like.

R ¹ is preferably a C 5-6 cycloalkyl-C 1-3 alkyl group, and more preferably cyclohexylmethyl or 2-cyclohexylethyl.

  In the phenylene derivative (II), the “C 6-14 aryl-carbonyl-C 1-6 alkyl group” is a group in which the above “C 6-14 aryl group” is bonded to the carbonyl group, and the above “C 1-6 alkyl group”. For example, benzoylmethyl, 2-benzoylethyl, 3-benzoylpropyl, 4-benzoylbutyl, 5-benzoylpentyl, 6-benzoylhexyl, naphthoylmethyl, 2-naphthoylethyl and the like. It is done.

R ¹ is preferably a C 6-10 aryl-carbonyl-C 1-3 alkyl group, and more preferably benzoylmethyl or naphthoylmethyl.

  In the phenylene derivative (II), “C 1-6 aliphatic acyl group” refers to a group in which a hydrogen atom or a saturated or unsaturated chain hydrocarbon group having 2 to 5 carbon atoms is bonded to a carbonyl group. Examples include formyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, pivaloyl, hexanoyl, acryloyl, methacryloyl, crotonoyl and the like.

R ¹ is preferably a C 1-4 aliphatic acyl group, and more preferably butyryl.

  The substituents α and β are preferably C1-4 aliphatic acyl groups, more preferably acetyl and propionyl, and most preferably acetyl.

  In the phenylene derivative (II), “C 3-7 cycloalkane” is a cyclic alkane having 3 to 7 carbon atoms, and examples thereof include cyclopropane, cyclobutane, cyclopentane, cyclohexane, cycloheptane and the like.

R ⁶ and R ⁷ together form preferably a C 3-5 cycloalkane, more preferably cyclopropane.

  Examples of the “halogen atom” in the phenylene derivative (II) include a fluorine atom, a chlorine atom, a bromine atom, and an iodine atom.

  The substituents α and β are each preferably a fluorine atom, a chlorine atom, or a bromine atom, more preferably a fluorine atom or a chlorine atom, and most preferably a fluorine atom.

  In the phenylene derivative (II), the “C 1-6 alkoxy group” is a hydroxyl group substituted with the above C 1-6 alkyl group. For example, methoxy, ethoxy, 1-propoxy, 2-propoxy, 1-butoxy, 2-butoxy 2-methyl-1-propoxy, 2-methyl-2-propoxy, 1-pentyloxy, 2-pentyloxy, 3-pentyloxy, 2-methyl-2-butoxy, 3-methyl-2-butoxy, 1- Hexyloxy, 2-hexyloxy, 3-hexyloxy, 2-methyl-1-pentyloxy, 3-methyl-1-pentyloxy, 2-ethyl-1-butoxy, 2,2-dimethyl-1-butoxy, 2 , 3-dimethyl-1-butoxy and the like.

  The substituents α and β are each preferably a C1-4 alkoxy group, more preferably methoxy or ethoxy, and most preferably methoxy.

  In the phenylene derivative (II), the “C 1-6 alkylthio group” is a mercapto group substituted with the above C 1-6 alkyl group. For example, methylthio, ethylthio, 1-propylthio, 2-propylthio, 1-butylthio, 2- Butylthio, 2-methyl-1-propylthio, 2-methyl-2-propylthio, 1-pentylthio, 2-pentylthio, 3-pentylthio, 2-methyl-2-butylthio, 3-methyl-2-butylthio, 1-hexylthio, 2-hexylthio, 3-hexylthio, 2-methyl-1-pentylthio, 3-methyl-1-pentylthio, 2-ethyl-1-butylthio, 2,2-dimethyl-1-butylthio, 2,3-dimethyl-1 -A butylthio group etc. are mentioned.

  The substituents α and β are each preferably a C1-4 alkylthio group, more preferably methylthio or ethylthio, and most preferably methylthio.

In the phenylene derivative (II), the “C 1-6 alkylsulfonyl group” is a sulfonyl group (—SO ₂ —) substituted with the above C 1-6 alkyl group, and examples thereof include methanesulfonyl, ethanesulfonyl, 1-propanesulfonyl, 2-propanesulfonyl, 1-butanesulfonyl, 2-butanesulfonyl, 2-methyl-1-propanesulfonyl, 2-methyl-2-propanesulfonyl, 1-pentanesulfonyl, 2-pentanesulfonyl, 3-pentanesulfonyl, 2- Methyl-2-butanesulfonyl, 3-methyl-2-butanesulfonyl, 1-hexanesulfonyl, 2-hexanesulfonyl, 3-hexanesulfonyl, 2-methyl-1-pentanesulfonyl, 3-methyl-1-pentanesulfonyl, 2 -Ethyl-1-butanesulfonyl, 2,2-dimethyl And lu-1-butanesulfonyl and 2,3-dimethyl-1-butanesulfonyl groups.

  The substituents α and β are preferably C 1-4 alkylsulfonyl groups, more preferably methanesulfonyl and ethanesulfonyl, and most preferably methanesulfonyl.

  In the phenylene derivative (II), the “halo C 1-6 alkyl group” is the C 1-6 alkyl group substituted by 1 to 7 halogen atoms, and examples thereof include fluoromethyl, difluoromethyl, dichloromethyl, dibromomethyl. , Trifluoromethyl mitrichloromethyl, 2-fluoroethyl, 2-bromoethyl, 2-chloroethyl, 2-iodoethyl, 2,2-difluoroethyl, 2,2,2-trifluoroethyl, 2,2,2-trichloroethyl , Pentafluoroethyl, 3-fluoropropyl, 3-chloropropyl, 4-fluorobutyl, 5-fluoropentyl, 6-fluorohexyl and the like.

  The substituent β is preferably a C1-4 alkyl group substituted with 1 to 5 halogen atoms, more preferably fluoromethyl, chloromethyl, difluoromethyl, trifluoromethyl, pentafluoroethyl. Most preferred is trifluoromethyl.

  In the phenylene derivative (II), the “C 6-14 aryl-carbonyl group” is a group in which the above “C 6-14 aryl group” is bonded to a carbonyl group. For example, benzoyl, naphthylcarbonyl, phenanthrylcarbonyl, etc. Can be mentioned.

  The substituent β is preferably a C 6-10 aryl-carbonyl group, and most preferably benzoyl.

  In the phenylene derivative (II), the “C 1-6 alkoxyimino group” is an imino group substituted with the above C 1-6 alkoxy group, for example, methoxyimino, ethoxyimino, 1-propoxyimino, 2-propoxyimino, 1-butoxyimino, 2-butoxyimino, 2-methyl-1-propoxyimino, 2-methyl-2-propoxyimino, 1-pentyloxyimino, 2-pentyloxyimino, 3-pentyloxyimino, 2-methyl- 2-butoxyimino, 3-methyl-2-butoxyimino, 1-hexyloxyimino, 2-hexyloxyimino, 3-hexyloxyimino, 2-methyl-1-pentyloxyimino, 3-methyl-1-pentyloxy Imino, 2-ethyl-1-butoxyimino, 2,2-dimethyl-1-butoxyimino 2,3-dimethyl-1-Butokishiimino like.

  The substituent β is preferably a C 1-4 alkoxyimino group, more preferably methoxyimino or ethoxyimino, and most preferably methoxyimino.

  In the phenylene derivative (II), A is preferably a group represented by the general formula (A4), (A5) or (A6).

  In the phenylene derivative (II), X is preferably a single bond or phenylene. Here, when X is phenylene, B is preferably a 1H-tetrazol-5-yl group, and when X is a single bond, B is preferably 2,4-dioxo-1, 3-thiazolidin-5-yl group.

  In the phenylene derivative (II), Y1 is preferably sulfonyl or carbonyl.

  In the phenylene derivative (II), Y2 is preferably a C1-3 alkylene group or a single bond, more preferably methylene, ethylene or a single bond.

  In the phenylene derivative (II), Y3 is preferably carbonyl or a single bond.

  In the present invention, Y4 is preferably a nitrogen atom.

  In the phenylene derivative (II), Y5 is preferably carbonyl or a single bond.

In the phenylene derivative (II), R ¹ is preferably a C 1-8 alkyl group (the C 1-8 alkyl group may be substituted with 1 to 5 C 1-6 aliphatic acyl groups), C 6 -14 aryl group (the C6-14 aryl group may be substituted with 1 to 5 groups selected from the group consisting of C1-6 alkyl group and C6-14 aryl group), C6-14 aryl- A C1-6 alkyl group (the C6-14 aryl-C1-6 alkyl group is a group selected from the group consisting of a C6-14 aryl group, a halogen atom, a C1-6 alkoxy group and a C6-14 aryl-carbonyl group); 1 to 5 optionally substituted), a 5- to 10-membered heterocyclic group containing the same or different heteroatoms selected from a nitrogen atom, an oxygen atom and a sulfur atom (the heterocyclic group is , Selected from the group consisting of C1-6 alkyl groups and C6-14 aryl groups And a 5- to 10-membered heterocyclic ring containing the same or different, or two heteroatoms selected from a nitrogen atom, an oxygen atom and a sulfur atom -C1- 6 alkyl group (the heterocyclic-C 1-6 alkyl group is substituted with 5 to 5 groups selected from the group consisting of C 1-6 alkyl group, C 6-14 aryl group and C 1-6 alkoxyimino group). C3-7 cycloalkyl group (the C3-7 cycloalkyl group may be substituted with an oxo group), C3-7 cycloalkyl-C1-6 alkyl group, C6-14 aryl-carbonyl- A C1-6 alkyl group, or a C1-6 aliphatic acyl group, more preferably a C4-8 alkyl group (preferably butyl, pentyl, hexyl, heptyl, octyl) (the C4-8 alkyl group is C1-4 aliphatic acyl group (preferably acetyl, 1-6 substituted with propionyl), C6-10 aryl group (preferably phenyl, naphthyl) (the C6-10 aryl group is C1-3 alkyl group (preferably methyl, ethyl)) And a C6-10 aryl group (preferably substituted with 1 to 5 groups selected from the group consisting of phenyl), a C6-10 aryl-C1-3 alkyl group (preferably benzyl, 1 -Phenethyl, 2-phenethyl, 3-phenylpropyl, 1-naphthylmethyl, 2-naphthylmethyl, 2- (1-naphthyl) ethyl, 2- (2-naphthylethyl)) (the C6-10 aryl-C1-3 The alkyl group includes C6-10 aryl group (preferably phenyl), halogen atom (preferably fluorine atom, chlorine atom, bromine atom), C1-4 alkoxy group (preferably methoxy, ethoxy) and C6-10 aryl. -Carbonyl group (preferably Suitably substituted with 1 to 5 groups selected from the group consisting of benzoyl)), the same or different heteroatoms selected from nitrogen, oxygen and sulfur atoms. 5- or 6-membered heterocyclic group containing (this heterocyclic ring may be condensed with a benzene ring. Preferably a group selected from the group consisting of pyridyl, oxazolyl, thiazolyl, quinolyl) (wherein the heterocyclic group is a C1-3 alkyl group (preferably methyl, ethyl) and a C6-10 aryl group (preferably phenyl)) A 5- or 6-membered heterocyclic-C1-3 alkyl group containing 1 or 2 heteroatoms which are the same or different and are selected from a nitrogen atom, an oxygen atom and a sulfur atom (The heterocyclic ring may be condensed with a benzene ring. Preferably, oxazolylmethyl, thiazolylmethyl, pyridylmethyl, quinolylmethyl, thienylethyl, thienylpropyl) (the heterocyclic-C1-3 alkyl group is C1- 3 alkyl group (preferably methyl, ethyl), C6-10 aryl group (preferably phenyl), C1-4 alkoxyimino group (preferably methoxyimino, ethoxyimino ), A C5-6 cycloalkyl group (preferably cyclopentyl, cyclohexyl) (the C5-6 cycloalkyl group is substituted with an oxo group). C5-6 cycloalkyl-C1-3 alkyl group (preferably cyclohexylmethyl, 2-cyclohexylethyl), C6-10 aryl-carbonyl-C1-3 alkyl group (preferably benzoylmethyl, Naphthoylmethyl), or a C1-4 aliphatic acyl group (preferably butyryl).

In the phenylene derivative (II), R ² is preferably a hydrogen atom, a carboxyl group, a phenyl group, or a group represented by 1C (O) NR ¹³ R ¹⁴ .

In the phenylene derivative (II), R ³ is preferably a hydrogen atom or a C 6-14 aryl group, more preferably a hydrogen atom or a C 6-10 aryl group (preferably phenyl, naphthyl).

In the phenylene derivative (II), R ⁴ and R ⁵ are preferably the same or different and each is a hydrogen atom or a C 6-10 aryl group, more preferably a hydrogen atom or phenyl.

In the phenylene derivative (II), R ⁶ and R ⁷ are preferably combined together to form a C 3-5 cycloalkane, or the same or different, each being a hydrogen atom or a Cl-3 alkyl group, More preferably, they are the same or different and each is a hydrogen atom or a C1-3 alkyl group (preferably methyl, ethyl, propyl).

In the phenylene derivative (II), R ⁸ and R ⁹ are preferably the same or different and each is a hydrogen atom or a carboxyl group, more preferably one is a hydrogen atom and the other is a carboxyl group.

In the phenylene derivative (II), R ¹⁰ is preferably a hydrogen atom, a C 1-4 alkyl group, a C 6-10 aryl group, or the same or different 1 selected from a nitrogen atom, an oxygen atom and a sulfur atom. A 5- or 6-membered heterocyclic group containing 2 heteroatoms (the heterocyclic group may be condensed with a benzene ring), more preferably a C1-4 alkyl group (preferably methyl, ethyl , Propyl, butyl), a C6-10 aryl group (preferably phenyl), or a 5- or 6-membered heterocycle containing the same or different heteroatoms selected from nitrogen, oxygen and sulfur atoms A group (the heterocyclic group may be condensed with a benzene ring) (preferably pyridyl, quinolyl).

In the phenylene derivative (II), R ¹¹ and R ¹² are preferably the same or different and each is a hydrogen atom or a carboxyl group, and more preferably a carboxyl group.

In the phenylene derivative (II), R ¹³ and R ¹⁴ are preferably combined together as a 5- or 6-membered nitrogen-containing heterocycle (the nitrogen-containing heterocycle may be condensed with a benzene ring) (preferably Imidazole, pyrrolidine, piperidine, morpholine, indole) or the same or different hydrogen atom or C1-3 alkyl group (preferably methyl, ethyl).

Specifically, the following compounds are suitable as the phenylene derivative (II).
(II-1) 3- {N- (3-phenylpropionyl) -N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-2) 3- {N-phenylacetyl-N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-3) 3- {N- (4-phenylbutyryl) -N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-4) 3- {N-cyclohexanecarbonyl-N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-5) 3- {N- (pyridin-3-carbonyl) mono-N- [2 '-(lH-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-6) 3- {N-pentyl-N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-7) 1- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -2-methyl-1,2,3,4-tetrahydroquinoline-7-carboxylic acid,
(II-8) 5- [4- (N-methylsulfonyl-N-phenylaminomethyl) phenyl] -1,3-thiazolidine-2,4-dione,
(II-9) 5- [4- (N-butylsulfonyl-N-phenylaminomethyl) phenyl] -1,3-thiazolidine-2,4-dione,
(II-10) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (methylsulfonyl) amino} benzoic acid,
(II-11) 3- {N-butylsulfonyl-N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-12) 5- {4-[(2-oxo-2H-quinolin-1-yl) methyl] phenyl} -1,3-thiazolidine-2,4-dione,
(II-13) 5- {4-[(2-methylbenzimidazol-1-yl) methyl] phenyl} -1,3-thiazolidine-2,4-dione,
(II-14) 5- {4-[(2-propylbenzimidazol-1-yl) methyl] phenyl} -1,3-thiazolidine-2,4-dione,
(II-15) 5- {4- [2- (2-pyridyl) benzimidazol-1-ylmethyl] phenyl} -1,3-thiazolidine-2,4-dione,
(II-16) 5- [4- (2-phenylimidazol-1-ylmethyl) phenyl] -1,3-thiazolidine-2,4-dione,
(II-17) 1- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-3-propyl-1,2-dihydroquinoline-7-carboxylic acid,
(II-18) 3- {N-cyclohexanecarbonyl-N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-19) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (pyridine-3-carbonyl) amino} benzoic acid,
(II-20) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (3-phenylpropionyl) amino} benzoic acid,
(II-21) 3- {N- (biphenyl-4-carbonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-22) 3- {N- (phenylsulfonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-23) 3- {N- (4-methylphenylsulfonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl) amino) benzoic acid,
(II-24) 3- {N- (biphenyl-4-sulfonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-25) 3- {N- (2-naphthylsulfol) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-26) 3- {N- (benzylsulfonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-27) 1- [4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-3-propyl-2,3-1H-indole-6-carboxylic acid ,
(II-28) 1- [4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-3-propyl-2,3,4,5-tetrahydro-1H- Benzo [b] azepine-8-carboxylic acid,
(II-29) 3,3-Dimethyl-1- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-1,2,3,4-tetrahydroquinoline -7-carboxylic acid,
(II-30) 1 ′-[4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -2′-oxo-1 ′, 4′-dihydro-2′H-spiro [ Cyclopropane-1,3′-quinoline] -7′-carboxylic acid,
(II-31) 1- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -2-propyl-1,2,3,4-tetrahydroquinoline-7-carboxylic acid,
(II-32) 3- {N-phenethyl N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-33) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N-[(5-methyl-2-phenyl 1,3-oxazole- 4-yl) acetyl] amino} benzoic acid,
(II-34) 3- {N-[(2,5-dimethyl-1,3-oxazol-4-yl) carbonyl] -N- [4- (2,4-dioxo-1,3-thiazolidine-5 -Yl) benzyl] amino} benzoic acid,
(II-35) 2-Oxo-3-propyl-1- [2,-(1H-tetrazol-5-ylmethyl) biphenyl-4-ylmethyl] -2,3,4,5-tetrahydro-H-benzo [b] Azepine-8-carboxylic acid,
(II-36) 3- {N- (3-cyclohexylpropanoyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-37) 3- {N-[(5-Methyl-2-phenyl-1,3-oxazol-4-yl) acetyl] -N- [2 '-(1H-tetrazol-5-yl) biphenyl 4- Ylmethyl] amino} benzoic acid,
(II-38) 5- [4 ′-(2-phenylimidazol-1-ylmethyl) biphenyl-2-yl] -1H-tetrazole,
(II-39) 5- [4 ′-(4-phenylimidazol-1-ylmethyl) biphenyl-2-yl] -1H-tetrazole,
(II-40) 5- [4 ′-((5-phenylimidazol-1-ylmethyl) biphenyl-2-yl] -1H-tetrazole,
(II-41) 2-propyl-1- [2 ′-(lH-tetrazol-5-yl) biphenyl-4-ylmethyl] -1,2,3,4-tetrahydroquinoline-7-carboxylic acid,
(II-42) 3- {N- (2-oxo-2-phenylethyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-43) 3- {N- (3-quinolinecarbonyl) -N- [2 '-(lH-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-44) 3- {N- (2-naphthoyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-45) 3- {N- (4-biphenylcarbonyl) -N- [2 '-(lH-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-46) 3- {N-[(2,5-Dimethyl-1,3-oxazol-4-yl) carbonyl] -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl Amino} benzoic acid,
(II-47) 3- {N- [4- (5-Methyl-1,2,4-oxadiazol-3-yl) benzoyl] -N- [2 '-(1H-tetrazol-5-yl) Biphenyl 4-ylmethyl] amino} benzoic acid,
(II-48) 3- {N- [4- (2-methylthiazol-4-yl) benzenesulfonyl] -N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid acid,
(II-49) 3- {N- [4- (2-pyridyl) benzoyl] mono-N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-50) N- [3- (morpholinocarbonyl) phenyl] -N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] -4- (2-pyridyl) benzamide,
(II-51) 3- {N- [4- (2-pyridyl) benzoyl] -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzamide,
(II-52) N- [3- (morpholinocarbonyl) phenyl] -N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] quinoline-3-carboxamide,
(II-53) 3- {N- (3-quinolinecarbonyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzamide,
(II-54) 3- {N-benzyl N- [2 ′-(1H-tetrazol-5-yl) biphenyl-ylmethyl] amino} benzoic acid,
(II-55) 3- {N- (biphenyl-4-ylmethyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-56) 3- {N- (4-chlorobenzyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-57) 3- {N- (3,4-dimethoxybenzyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-58) 3- {N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] mono-N- [2- (2-thienyl) ethyl] amino} benzoic acid,
(II-59) 3- {N- [2- (3,4-dimethoxyphenyl) ethyl] -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-60) 3- {N- (2-cyclohexylethyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-61) N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N-phenyl 1-naphthalenesulfonamide,
(II-62) N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N-phenyl 2-naphthalenesulfonamide,
(II-63) 3- {N- [4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -N- [4- (2-methylthiazol-4-yl) benzenesulfonyl Amino} benzoic acid,
(II-64) 3- {N- [4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -N- [4- (5-methyl-1,2,4-oxa Diazol-3-yl) benzoyl] amino} benzoic acid,
(II-65) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- [4- (2-pyridyl) benzoyl] amino} benzoic acid,
(II-66) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (quinoline-3-carbonyl) amino} benzoic acid,
(II-67) N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (2-piphenylyl) benzenesulfonamide,
(II-68) 3- {N- (biphenyl-4-ylmethyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-69) 3- {N- (2-cyclohexylethyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-70) 3- {N-butylsulfonyl-N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzamide,
(II-71) 5- {4- [N- (3-morpholinocarbonylphenyl) -N- (2-naphthalenesulfonyl) aminomethyl] phenyl} -1,3-thiazolidine-2,4-dione,
(II-72) 5- {4-[{2- (2-naphthyl) imidazol-1-yl} methyl] phenyl} -1,3-thiazolidine-2,4-dione,
(II-73) 5- {4-[{2- (4-biphenylyl) imidazol-1-yl} methyl]} phenyl 1,3-thiazolidine-2,4-dione,
(II-74) 2-oxo-N-phenyl N- [2 ′-(1H-tetrazol-5-yl) biphenyl 4-ylmethyl] valeramide,
(II-75) 3- {N- (7-oxooctanoyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-76) 3- {N- [4- (E) -methoxyimino-4- (2-thienyl) butyryl] -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-77) 3- {N- [4- (Z) methoxyimino-4- (2-thienyl) butyryl] -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino }benzoic acid,
(II-78) 3- {N- [2- (3-benzoylphenyl) propionyl] -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
and
(II-79) 3- {N- (3-oxo-1-cyclopentanecarbonyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid.

  Among the phenylene derivatives (II), (II-1), (II-6), (II-7), (II-8), (II-9), (II-10), (II -11), (II-15), (II-16), (II-17), (II-18), (II-19), (II-20), (II-21), (II-22) ), (II-23), (II-24), (II-25), (II-26), (II-27), (II-28), (II-29), (II-30), The compounds listed in (II-32), (II-33), (II-36), (II-42), (II-63) and (II-68).

  These phenylene derivatives (II), and pharmacologically acceptable salts and pharmacologically acceptable esters thereof are produced using, for example, known compounds as starting materials according to the method described in International Publication WO2007 / 026962 Pamphlet. can do. In addition, the manufacturing method of the phenylene derivative described in the said publication | presentation gazette is used as content of this specification as it is.

  As in the case of the phenylene derivative (I), when the phenylene derivative (II) has a basic group, it can be converted into an acid addition salt according to a conventional method. Examples of the salt include the same salts as the phenylene derivative (I). Preferred is a salt of hydrohalic acid. Further, when the phenylene derivative (II) has a carboxyl group, it can be converted into a metal salt according to a conventional method, like the phenylene derivative (I). Examples of such a salt include those similar to the phenylene derivative (I). Alkali metal salts are preferred.

  The phenylene derivative (II) of the present invention can be converted into a pharmacologically acceptable ester according to a conventional method. Such ester is not particularly limited as long as it is medically used and pharmacologically acceptable. Specific examples of suitable ester residues include, for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, benzyl, acetoxymethyl, 1- (acetoxy) ethyl, propionyloxymethyl, 1- (propionyloxy) Ethyl, butyryloxymethyl, 1- (butyryloxy) ethyl, isobutyryloxymethyl, 1- (isobutyryloxy) ethyl, valeryloxymethyl, 1- (valeryloxy) ethyl, isovaleryloxymethyl, 1- (Isovaleryloxy) ethyl, pivaloyloxymethyl, 1- (pivaloyloxy) ethyl, methoxycarbonyloxymethyl, 1- (methoxycarbonyloxy) ethyl, ethoxycarbonyloxymethyl, 1- (ethoxycarbonyloxy) ethyl, propoxy Carbonyloxymethyl, 1- ( Propoxycarbonyloxy) ethyl, isopropoxycarbonyloxymethyl, 1- (isopropoxycanolenyloxy) ethyl, butoxycarbonyloxymethyl, 1- (butoxycarbonyloxy) ethyl, isobutoxycarbonyloxymethyl, 1- (isobutoxycarbonyl) Oxy) ethyl, t-butoxycarbonyloxymethyl, 1- (t-butoxycarbonyloxy) ethyl, cyclopentanecarbonyloxymethyl, 1- (cyclopentanecarbonyloxy) ethyl, cyclohexanecarbonyloxymethyl, 1- (cyclohexanecarbonyloxy) Ethyl, cyclopentyloxycarbonyloxymethyl, 1- (cyclopentyloxycarbonyloxy) ethyl, cyclohexyloxycarbonyloxymethyl, 1- (cyclohexyloxycarbonyl) Bonyloxy) ethyl, benzoyloxymethyl, 1- (benzoyloxy) ethyl, phenoxycarbonyloxymethyl, 1- (phenoxycarbonyloxy) ethyl, (5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl 2-trimethylsilylethyl or phthalidyl, more preferably (5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl, pivaloyloxymethyl or 1- (isopropoxycarbonyloxy) Ethyl.

  In addition, when the phenylene derivative represented by the general formula (II), its salt or ester forms a solvate (for example, hydrate), all of these solvates are also included in the present invention. Furthermore, all compounds that are metabolized in vivo and converted to the above-mentioned phenylene derivative (II), salts or esters thereof (for example, so-called prodrugs such as amide derivatives) are also included in the present invention.

  Any of the above compounds (1) and (2), and phenylene derivatives (I) and (II), if necessary, aminoguanidine, pyridoxamine derivatives, OPB-9195, biguanide compounds, cross-linking inhibitors, amadori compounds May be used together with known drugs such as enzymes that degrade GSH, cysteine, acetylcysteine, vitamin E, ubiquinol, aldose reductase inhibitors, carbonyl compound trapping agents, etc., thereby enhancing the sustainability of AGEs production inhibitory action be able to.

  The carbonyl scavenger and the AGEs production inhibitor of the present invention are administered in various forms. The administration form is not particularly limited, and is determined according to various preparation forms, patient age, sex and other conditions, the degree of disease, and the like. For example, in the case of tablets, pills, powders, granules, syrups, solutions, suspensions, emulsions and capsules, they are orally administered. In the case of an injection, it is administered intravenously alone or mixed with a normal fluid such as glucose or amino acid, and further administered alone intramuscularly, intradermally, subcutaneously or intraperitoneally as necessary. In the case of a suppository, it is administered intrarectally. Oral administration is preferred.

  These various preparations are prepared by using known adjuvants that can be usually used in the field of pharmaceutical preparations such as excipients, binders, disintegrants, lubricants, solubilizers, flavoring agents, coating agents, etc. Can be

  In molding into tablets, conventionally known carriers can be widely used as carriers, such as lactose, sucrose, sodium chloride, glucose, urea, starch, calcium carbonate, kaolin, crystalline cellulose, silicic acid and the like. Form, Water, Ethanol, Propanol, Glucose solution, Starch solution, Gelatin solution, Carboxymethylcellulose, Shellac, Methylcellulose, Potassium phosphate, Polyvinylpyrrolidone and other binders, Dry starch, Sodium alginate, Agar powder, Laminaran powder, Carbonic acid Sodium hydride, calcium carbonate, polyoxyethylene sorbitan fatty acid esters, sodium lauryl sulfate, stearic acid monoglyceride, starch, lactose and other disintegrants, sucrose, stearic acid, cocoa butter, hydrogenated oil and other disintegration inhibitors, quaternary Ammonium base, LA Absorption promoters such as sodium rilsulfate, humectants such as glycerin and starch, adsorbents such as starch, lactose, kaolin, bentonite and colloidal silicic acid, purified talc, stearate, shelf acid powder, polyethylene glycol, etc. A lubricant can be exemplified. Further, the tablets can be made into tablets with ordinary coatings as necessary, for example, sugar-coated tablets, gelatin-encapsulated tablets, enteric-coated tablets, film-coated tablets, double tablets, and multilayer tablets.

  In molding into the form of pills, those conventionally known in this field can be widely used as carriers, for example, glucose, lactose, starch, cocoa butter, hydrogenated vegetable oil, kaolin, talc and other excipients, gum arabic powder, Examples thereof include binders such as tragacanth powder, gelatin and ethanol, and disintegrants such as lamina lankanten.

  In the case of forming into a suppository, conventionally known carriers can be widely used as carriers, such as polyethylene glycol, cacao butter, higher alcohols, higher alcohol esters, gelatin, semi-synthetic glycerides and the like. Can do.

  When prepared as an injection, solutions and suspensions are preferably sterilized and isotonic with blood. When forming into these liquids, emulsions and suspensions, this is used as a diluent. Anything customary in the field can be used. Examples of the diluent include water, ethyl alcohol, propylene glycol, ethoxylated isostearyl alcohol, polyoxylated isostearyl alcohol, polyoxyethylene sorbitan fatty acid esters and the like. In this case, a sufficient amount of sodium chloride, glucose, or glycerin to prepare an isotonic solution may be contained in the pharmaceutical preparation. Ordinary solubilizers, buffers, soothing agents, etc. may be added. It may be added.

  Furthermore, you may contain a coloring agent, a preservative, a fragrance | flavor, a flavoring agent, a sweetening agent, and other pharmaceuticals as needed.

  The amount of the compound as an active ingredient contained in the pharmaceutical preparation is not particularly limited and is appropriately selected over a wide range, but is usually 1 to 70 parts by weight, preferably 1 to 30 parts by weight in the total composition. Is appropriate. The dose varies depending on symptoms, age, body weight, administration method and dosage form, but is usually 0.01 mg (preferably 0.1 mg, more preferably 1 mg) as the lower limit and 2,000 as the upper limit per day for adults. mg (preferably 1,000 mg, more preferably 200 mg) can be administered in one or several divided doses.

  For the purpose of treating or improving schizophrenia, such a pharmaceutical preparation is used by administering an effective amount to a patient determined to be schizophrenia. Therefore, the pharmaceutical preparation of the present invention may be accompanied by specifications or instructions describing the usage when used for the treatment or improvement of schizophrenia.

IV. Method for treating or improving schizophrenia The method for treating or improving schizophrenia according to the present invention comprises an effective amount of a therapeutic or ameliorating agent for schizophrenia containing the aforementioned carbonyl scavenger or AGEs production inhibitor as an active ingredient. It is performed by administering to patients who are judged to have symptoms.

  The carbonyl scavenger or AGEs production inhibitor is an effective amount for removing carbonyl stress in schizophrenic patients and improving or treating schizophrenia with a pharmaceutically acceptable carrier or other additive. In addition, it can be used in the form of a pharmaceutical composition. The administration subject (subject), administration mode, administration route, administration method, and administration dose (administration amount of the active ingredient) of the pharmaceutical composition are as described above.

V. The present invention relates to a method effective for developing a drug or food useful for improving or preventing the development of schizophrenia, specifically, schizophrenia. Provided is a method (screening method) for selecting a candidate substance effective for improving treatment or preventing onset.

  The method can be carried out by selecting a substance having a carbonyl elimination action or a carbonyl protein modification product production inhibitory action (AGEs production inhibitory action) from among the test substances. Preferably, it can be carried out by selecting a substance having the action from the test substance using the AGEs production inhibitory action as an index.

  The test substance used in the method is not particularly limited. For example, natural compounds, organic compounds, inorganic compounds, single compounds such as proteins and peptides; and compound libraries, gene library expression products, cell extracts, cell culture supernatants, fermented microorganism products, marine organism extracts Examples of the composition include plant extracts.

The presence or absence of an AGEs production inhibitory effect can be confirmed by the test (i) below
(I) Using pentosidine as a representative AGEs as an index, plasma is collected from a nondiabetic renal failure dialysis patient, a test substance is added, and the amount of pentosidine produced after a certain period of time is measured.

  For the detailed method, the description in Experimental Example 1 can be referred to. In this case, the AGEs production inhibitory effect of the test substance can be evaluated by comparing with the amount of pentosidine production measured using pyridoxamine as a positive control instead of the test substance. Specifically, it can be determined that a test substance having an equivalent or small amount of pentosidine produced compared to the amount of pyridoxamine, which is a positive control, has an AGEs production inhibitory effect.

Moreover, the presence or absence of the AGEs production | generation inhibitory effect can also be confirmed by the test of the following (ii):
(Ii) The radical scavenging ability of the test substance in the phenylalanine-radical reaction system is measured.

  This method utilizes the fact that the hydroxy radical generated by the reaction between hydrogen peroxide and copper sulfate hydroxylates phenylalanine to produce o-tyrosine or m-tyrosine. Specifically, the method involves the production of o-tyrosine or m-tyrosine when a test substance is added to a reaction system containing hydrogen peroxide, copper sulfate and phenylalanine, and o when no test substance is added. -It can be carried out by comparing the amount of tyrosine or m-tyrosine produced, and if the suppression of o-tyrosine or m-tyrosine production is observed when the test substance is added, the test substance is free of radicals. It can be determined that it has a capturing ability and has an inhibitory effect on AGE generation. A specific measurement method is shown below.

<Measurement method of radical scavenging ability>
Preparation dissolved in 200 mM phosphate buffer (pH 7.4) so that the final concentration is 1 mM phenylalanine, 0.1 mM, 0.5 mM and 2.5 mM test substance, 5 mM hydrogen peroxide, 0.1 mM copper sulfate (total volume 500 μl) The solution is incubated at 37 ° C. for 4 hours to react. After completion of the reaction, the reaction is stopped by adding DTPA (diethylene triamine pentaacetic acid) and 260 units of catalase to a final concentration of 1 mM. Next, the production amounts of o-tyrosine and m-tyrosine are measured using HPLC under the following conditions.

HPLC conditions Column: C18 reverse phase column (4.6 × 250 mm, 5 μm: manufactured by Nomura Chemical Co., Ltd.)
Mobile phase: Buffer A: 0.10% trifluoroacetic acid,
Buffer B: Tolyl gradient in 80% aceto containing 0.08% trifluoroacetic acid: Increase buffer B concentration from 6.5% to 10% in 25 minutes Flow rate: 0.6ml / min Detection: Fluorescence detector (RF-10A, Shimadzu) Detected at an excitation wavelength of 275 nm and a fluorescence wavelength of 305 nm.

  The candidate substance preferably does not cause vitamin B6 deficiency. Therefore, it is preferable to confirm whether or not the test substance further causes vitamin B6 deficiency by the following test (a) or (b).

  (A) A test substance is added to the vitamin B6 solution, and the residual amount of vitamin B6 after a predetermined time is measured.

  (B) The test substance is administered to normal rats, and the presence or absence of the onset of vitamin B6 deficiency after a certain period is confirmed.

  Candidate substances selected by the above method (selection of candidate substances) are subjected to further pharmacological tests, clinical tests, and toxicity tests as necessary, so that schizophrenia is more effective and safe for humans. It can be obtained as an active ingredient of an agent for improving treatment or preventing onset.

  The candidate substance thus obtained can be provided as a therapeutic improvement or prevention agent for schizophrenia by formulation and formulation by a known method.

  Examples are given below to illustrate the present invention in more detail. However, the embodiment is an example of the present invention, and the present invention is not limited to the embodiment.

Example 1 Gene and Biochemical Analysis of Severe Schizophrenia Patients glyoxalase I gene analysis of schizophrenia patients (male, 60 years old, weight 75.5 kg) (hereinafter referred to as patient A) who are admitted to the psychiatric ward In addition, erythrocyte glyoxalase activity, serum AGEs value, and serum vitamin B6 were measured. In addition, the above-mentioned patient A is diagnosed as a patient with severe familial schizophrenia because 2 out of 4 siblings are suicide, 2 are currently hospitalized in the psychiatric ward.

<Measurement method>
(1) Measurement of erythrocyte glyoxalase activity
It was measured according to the method of McLellan et al. (McLellan AC, Thornalley PJ: Glyoxalase activity in human red blood cells fractioned by age. Mech Ageing Dev 48: 63-71, 1989). Specifically, the disrupted erythrocytes were added to a hemi-thioacetal solution produced from methylglyoxal and glutathione, and the amount of SD-lactoylglutathion formed was measured at an absorption wavelength of 240 nm using a spectrophotometer. From the measured value, the SD-lactoylglutathion concentration was calculated based on the molar extinction coefficient Δε ₂₄₀ = 2.86 Mm ⁻¹ cm ⁻¹ and converted into units. Incidentally, 1 Unit erythrocytes 10 ⁶ per minute, is an amount to form a SD-lactoylglutathion of 1 [mu] mol.

(2) Measurement of serum AGEs
Pentosidine was used as an index as an AGE value. Pentosidine was measured by hydrolyzing the collected serum in 6N HCl at 110 ° C for 16 hours under nitrogen, and then excited by reverse-phase high performance liquid chromatography (HPLC) using a known concentration of synthetic pentosidine as a standard. -Quantitative analysis at absorption wavelength (335 / 385nm).

(3) Measurement of serum vitamin B6 amount As serum vitamin B6 amount, serum Pyridoxamine value, Pyridoxine value, and Pyridoxal value were measured. In addition, the measurement was requested to a contract inspection company (SRL Co., Ltd.), and quantitative analysis was performed by reversed phase high performance liquid chromatography (HPLC).

<Measurement results>
(1) Red blood cell glyoxalase activity (Fig. 4)
When the red blood cell count of 10% hematocrit was calculated as 0.6 × 10 ⁹ / ml, the red blood cell Glyoxalase I activity of patient A was 2.9 mUnit / 10 ⁶ RBC, and the red blood cell Glyoxalase I activity (6.1 ± 0.7 mUnit / 10 ⁶ RBC) (Fig. 4).

(2) glyoxalase I gene analysis results (Figure 5)
The results of glyoxalase I gene analysis for patient A are shown in FIG. As can be seen from the comparison with the healthy person's glyoxalase I gene, the patient A's glyoxalase I gene has an A (adenine) inserted at position 80 (point mutation), resulting in a frameshift and normal It was found that no glyoxalase I was expressed. From this, it was found that the decrease in (1) erythrocyte glyoxalase activity in patient A was caused by glyoxalase I gene abnormality (frame shift due to point mutation).

(3) Measurement of serum AGEs (Figure 6)
FIG. 6 shows the results of comparing serum AGEs (pentosidine levels) with healthy subjects. As can be seen from this result, the serum AGEs value of patient A was 0.368 nmol / ml, and the serum AGEs value of healthy subjects (n = 5) was significantly higher than 0.128 ± 0.04 nmol / ml. When converted to the amount of pentosidine per mg of protein, the value of patient A was found to be 5 pmol / mg proten, 3 times higher than that of 1.7 ± 0.4 pmol / mg proten of healthy subjects.

  The estimated glomerular filtration rate (eGFR) of this patient A (60 years old, body weight 75.5 kg, creatinine 1.05 mg / dl) is 76.7 ml / min by MDRD method, and estimated creatinin clearance (eCcr) is 79.9 by Cockcroft-Gault method. ml / min, and a slight decrease in kidney function was observed. However, this decrease in kidney function cannot cause an increase in AGEs (pentosidin) as described above.Therefore, in schizophrenia, there are factors other than diabetes and renal failure that increase AGEs (pentosidin). It was. Considering the aforementioned Glyoxalase I deficiency, it is considered that the high serum AGEs in schizophrenic patients are caused by abnormal AGE precursor detoxification due to decreased Glyoxalase I activity.

(4) Serum vitamin B6 level Among serum vitamin B6, Pyridoxamine and Pyridoxine levels were below the detection limit as in healthy subjects (Pyridoxamine detection limit was 0.2 ng / ml, Pyridoxine detection limit was 3.0 ng / ml) ). On the other hand, the Pyridoxal value was 2.8 ng / ml, which was significantly lower than that of healthy individuals (14.8 ± 0.3 ng / ml, n = 2). By the way, the standard value for men is 6.0-40.0 ng / ml. Pyridoxal has the effect of scavenging carbonyls in the blood. Therefore, schizophrenic patients needed to erase carbonyl, so Pyridoxal was consumed in the body, and as a result, the amount of serum Pyridoxal (vitamin B6) was thought to have decreased.

  From the above, it was considered that carbonyl stress based on Glyoxalase I deficiency is related to the pathology of schizophrenia. In other words, in patients with schizophrenia, glyoxalase I activity decreased due to Glyoxalase I gene abnormality, which increased carbonyl stress (increased AGEs), thereby depleting vitamin B6 and worsening the pathology of schizophrenia It is thought that. Therefore, decreased glyoxalase I activity, increased carbonyl stress (increased AGEs), and decreased vitamin B6 can be indicators for the diagnosis of schizophrenia. In addition, AGEs inhibitors (AGEs production inhibitors) and carbonyl scavengers such as vitamin B6 and edaravone are considered to be effective as schizophrenia treatments or disease condition improving agents.

Example 2 Measurement of AGE content in skin Healthy subjects (12 women, 12 men, 24 in total; average age 48.88 + 3.17 years) and schizophrenic patients (12 women, 12 men, 24 in total) ;; AGEs content in skin was measured using AGE-Reader (Diagn Optics: The Netherlands) with a mean age of 48.38 + 2.23 years old).

  FIG. 7 shows the results of comparing the AGE content in the skin between healthy subjects and schizophrenic patients. From this result, it can be seen that the AGE content in the skin is significantly increased in schizophrenic patients compared to healthy individuals. FIG. 8A and FIG. 8B show the results of comparing the AGE content in the skin according to age between healthy subjects and schizophrenic patients. From this result, although a strong positive correlation was seen between age and AGE content in healthy subjects, it was not observed in schizophrenic patients (FIG. 8A). However, as shown in FIG. 8B, the AGE content in the skin of schizophrenic patients tended to be higher than that of healthy subjects even at 50 years of age or younger and even older than 50 years.

  Therefore, as shown in Example 1, schizophrenic patients have higher AGEs content than healthy individuals, and the tendency can be evaluated by AGE-Reader that measures the AGEs content in the skin. It has been shown.

Example 3 Analysis of glyoxalase I gene in patients with schizophrenia
(1) The glyoxalase I (GLO-I) gene was analyzed in schizophrenic patients (700) and healthy subjects (600).

  The frameshift mutation in the GLO-I gene (single base insertion between positions 79 and 80 in the base sequence of the coding region of the GLO-I gene (SEQ ID NO: 1)) is Blend Taq (TOYOBO Cat # BTQ- 101i), PCR primer F: 5'-GAGTTTGCCTCCTTTATGCG-3 '(SEQ ID NO: 8) and R: 5'- AACAGATCCCCTCCACACTT-3' (SEQ ID NO: 9) (i) 1 cycle at 94 ° C for 2 minutes (Ii) PCR-direct sequence method in which PCR is performed under the conditions of 94 ° C, 30 seconds-62.5 ° C, 20 seconds-72 ° C, 30 seconds for 40 cycles, and (iii) 72 ° C, 2 minutes for 1 cycle. Identified by In addition, the base substitution abnormality (mutation from alanine to cytosine at position 332 in the base sequence of the coding region of the GLO-I gene (SEQ ID NO: 1)) that causes a mutation (Glu → Ala) of the 111th amino acid of GLO-I is , Using Blend Taq (TOYOBO Cat # BTQ-101S), F: 5'-TCAGAGTGTGTGATTTCGTG-3 '(SEQ ID NO: 10) and R: 5'- CATGGTGAGATGGTAAGTGT-3' (SEQ ID NO: 11) ℃, 2 minutes for 1 cycle, (ii) 94 ° C, 30 seconds-62.5 ° C, 20 seconds-72 ° C, 30 seconds for 40 cycles, (iii) 72 ° C, 2 minutes for 1 cycle Identified by PCR-direct sequence method.

  As a result, a significant difference was observed centering on the SNP in which the 111th Glu of GLO-I amino acid sequence (SEQ ID NO: 2) was substituted with Ala. Four Ala / Ala homozygotes were detected in schizophrenic patients (700 patients), but none were found in healthy persons (600 patients).

(2) Of 4 Ala / Ala homozygous schizophrenia patients, 3 were measured for erythrocyte glyoxalase activity according to the method of Example 1 (1). The three persons have the following medical history and pathology.
(a) Patient B (hospitalized patient): Female, age 50 years, CREAT 0.57, 41 years old. At present, hallucination delusions have disappeared, but negative symptoms (decreased motivation and flattening of emotions) are conspicuous.
(b) Patient C (hospitalized patient): Male, age 66 years, CREAT 0.86, 15-year-old has negative onset symptoms and almost no spontaneous language. All you need to do is shake your head vertically and horizontally.
(c) Patient D (outpatient visit): Male, age 50 years, CREAT 0.72, 19-year-old onset of delusion of "harassment of neighbor" and repeated moving.

  Similarly, the erythrocyte glyoxalase activity of Glu / Ala heterozygotes and Glu / Glu heterozygous schizophrenic patients and healthy subjects was measured. The results are shown in FIG. As can be seen from this result, erythrocyte glyoxalase activity was significantly reduced in patients with schizophrenia with Ala / Ala homozygote. In addition, a GLO-I amino acid sequence (SEQ ID NO: 2) is constructed with a mutant GLO-I and GFP (pAcGFP-GLO1-Ala) in which position 111 is Ala, and normal GLO-I and GFP in which position 111 is Glu. The GLO-I activity of the mutant GLO-I was significantly higher than that of normal GLO-I when the construct (pAcGFP-GLO1-Glu) was expressed in COS cells and the GLO-I activity was measured. (Fig. 10).

  (3) Ala / Ala homozygous schizophrenic patients (4 patients) and schizophrenic patient A of Example 1 (one allele of the GLO-I gene (the nucleotide sequence of the coding region of the GLO-I gene) FIG. 11 shows the results of measurement of the expression level of GLO-I mRNA for GLO-I deficiency) between position 79 and position 80) due to frame shift caused by point mutation. As can be seen from this result, the expression level of GLO-I mRNA in the frameshift patient A was reduced by 50%. ALO / Ala homozygous schizophrenic patients also had a 20% decrease in GLO-I mRNA expression. This suggests that a 20% decrease in the expression level of GLO-I mRNA (GLO-I activity) may be a risk for schizophrenia, even if it does not decrease to 50%.

Example 4 glyoxalase I gene analysis and biochemical analysis of schizophrenic patients (Summary)
Divide schizophrenia patients by type (frameshift [n = 2], Ala / Ala homozygous [n = 5], Glu / Ala heterozygous [n = 1]), and Glyoxalase I activity for each patient (MUnit / 10 ⁶ RBC), pentosidine activity (pmol / mg protein), blood vitamin B6 content (ng / ml) (pyridoxal, pyridoxamine, pyridoxine), blood vitamin B12 content (pg / ml), blood folate content (Ng / ml), blood homocysteine content (nmol / ml), creatinine content (mg / dl), and eGFR (ml / min / 1.73 m ² ) were measured. Measurement of each activity and content was performed according to Example 1 and a conventional method. As a comparative control, the same measurement was performed on healthy subjects (Glu / Glu homozygous type) [n = 7] who did not suffer from schizophrenia.

  The results are shown in FIG. 12, including each person's presence or absence of diabetes.

  From these results, all types of schizophrenia patients had low GLO-I activity and higher pentosidine levels (AGEs) than healthy individuals. Since both patients have normal creatine and eGFR and no diabetes, such high pentosidine levels cannot be explained by decreased renal function or hyperglycemia, and are thought to be based on decreased GLO-I activity. In particular, MZ-65 patients with high pentosidine levels are severe schizophrenic patients with severe negative symptoms and almost no spontaneous language.

  Among patients with schizophrenia, especially those with frameshifts tend to have lower GLO-I activity, higher pentosidine levels (AGEs), lower pyridoxal levels, and higher homocysteine content than healthy individuals Was noticeable. This suggests a decrease in GLO-I, particularly as the etiology of patients with this type of schizophrenia. In other words, all of the above analysis results are due to a decrease in GLO-I in the body, and due to the decrease in GLO-I in the body, the carbonyl level becomes high, and as a result, the AGEs value (pentosidin content) in the body increases. It was thought that the amount of vitamin B6 (pyridoxal) to erase it was lowered. Moreover, since vitamin B6 is important for the formation of homocysteine, the high homocysteine level is considered to reflect the decrease in vitamin B6. Therefore, schizophrenia can be easily diagnosed by using these biochemical values as indices.

Test Example 1 AGEs Suppression Effect of Edaravone Analogue (1) 1- (5-Hydroxy-3-methyl-1-phenyl-1H-pyrazol-4-yl) -6-methyl-1,3-dihydro-furo [ 3,4-c] Pyridin-7-ol (hereinafter referred to as “TM-2002”) was examined for the effect of inhibiting the production of pentosidine, which is a typical AGE.

  Plasma from a hemodialysis patient before dialysis was collected prior to dialysis with consent and sterilized by filtration. A dimethyl sulfoxide solution (50 μL) of TM-2002 was added to plasma (450 μL) (final concentrations: 0.8, 2.0, 5.0 mM) and incubated at 37 ° C. for 1 week, and the amount of pentosidine produced was measured.

  Pentosidine was measured as follows. To each sample (50 μL) after incubation, an equal volume of 10% trichloroacetic acid was added, followed by centrifugation at 5000 g for 5 minutes. After removing the supernatant, the pellet was washed with 5% trichloroacetic acid (300 μL). The pellet was dried under reduced pressure, and then hydrolyzed at 110 ° C. for 16 hours in a 6N HCl solution (100 μL) under a nitrogen atmosphere. Next, 5N NaOH (100 μL) and 0.5 M phosphate buffer (pH 7.4) (200 μL) were added to the acid hydrolyzate, and then filtered through a 0.5 μm pore filter and diluted with PBS. The concentration of liberated pentosidine was measured using reverse phase HPLC using a fluorescence detector (RF-10A, Shimadzu Corporation) (Miyata, T. et al .; Proc. Natl. Acad. Sci. USA, Vol. 93, p.2353-2358, 1996). The effluent was monitored at an excitation / emission wavelength of 335/385 nm. Synthetic pentosidine was used as a standard. The detection limit for pentosidine was 0.1 pmol / mg protein.

  The inhibitory effect was evaluated by comparing with a positive control (pyridoxamine (Sigma)) reacted in the same manner as Compound TM-2002. In addition, the inhibitory effect was similarly investigated about aminoguanidine, olmesartan, and edaravone. The results (pentosidine amount nmol / ml) are shown in FIG. 13 (in the figure, “control” means a negative control using only a solvent. The same applies hereinafter). From this result, it is understood that TM-2002 significantly suppresses the production of pentosidine as compared with the positive control pyridoxamine.

  (2) The pentosidine production inhibitory activity of other edaravone analogs (compound (1) or (2)) was examined in the same manner as in (1) above, except that incubating with BSA and arabinose instead of patient plasma. The results were as shown in the following table. Note that “−” indicates that the test was not performed.

  As can be seen from these results, all of the test compounds (1) to (78) exhibited a pentosidine production inhibitory action.

Test Example 2 AGEs production inhibitory effect of phenylene derivative (I) In the present invention, the following test compounds belonging to the phenylene derivative represented by formula (I) were examined for AGEs production inhibitory action.
(1) 3- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -N-pentanoylamino] benzoic acid
(2) 3- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -N-acetylamino] benzoic acid
(3) 3- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -N-propanoylamino] benzoic acid
(4) 3- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -N-butanoylamino] benzoic acid
(5) 3- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -N-octanoylamino] benzoic acid
(6) 3- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -N-pentanoylamino] benzoic acid (5-methyl-2-oxo-1, 3-Dioxolen-4-yl) methyl
(7) 3- [N-[[4- (1H-tetrazol-5-yl) phenyl] methyl] -N-pentanoylamino] benzoic acid
(8) 2- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -N-pentanoylamino] benzoic acid
(9) 4- [N-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyll] methyl] -N-pentanoylamino] benzoic acid
(10) 2-Oxo-3-propyl-1-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -1,3,4-trihydroquinoline-7-carboxylic acid
(11) 2-oxo-3-propyl 1-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -1,3,4-trihydroquinoline-6-carboxylic acid
(12) 2-Oxo-3-propyl-1-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -1,3,4-trihydroquinoline-7-carboxylic acid (5-Methyl-2-oxo-1,3-dioxolen-4-yl) methyl
(13) 1-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -1,3,4-trihydroquinolin-2-one
(14) 3-Oxo-2-propyl-4-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -2H-benzo [e] 1,4-oxazine-6 carboxylic acid
(15) 3-oxo-2-propyl-4- [4- [4- (1H-tetrazol-5-yl) phenyl] methyl] 2-H-benzo [e] 1,4-oxazine-6-carboxylic acid
(16) 3-oxo-2-propyl-4-[[4- [2- (1H-tetrazol-5-yl) phenyl] phenyl] methyl] -2H-benzo [e] 1,4-thiazine-6 carboxylic acid
(17) 3-oxo-2-propyl-4-[[4- (1H-tetrazol-5-yl) phenyl] methyl] 2-H-benzo [e] 1,4-thiazine-6-carboxylic acid
(18) 5- [4-[(2-Ethyl-5,7-dimethylimidazo [4,5-b] pyridin-3-yl) methyl] phenyl] -1,3-thiazolidine-2,4-dione
(19) 5- [4-[(5,7-Dimethyl-2-propylimidazo [4,5-b] pyridin-3-yl) methyl] phenyl] -1,3-thiazolidine-2,4-dione
(20) 3- [N- [4- (2,4-Dioxothiazolidin-5-yl) benzyl] -N-pentanoylamino] benzoic acid.

  50 μl of a dimethyl sulfoxide solution of a test compound (final concentration 5 mM) was added to 450 μl of protein (renal failure patient serum from which informed consent was obtained), and the resulting mixture was incubated at 37 ° C. for 1 week. Next, the amount of pentosidine, which is one of the AGEs produced, was measured as follows.

  First, in order to release the pentosidine produced in the protein, 50 μl of 10% trichloroacetic acid was added to 50 μl of the sample after the reaction, and the protein was precipitated by centrifugation and collected. The recovered protein was washed with 300 μl of 5% trichloroacetic acid and dried, and then 100 μl of 6N hydrochloric acid was added, followed by hydrolysis at 110 ° C. for 16 hours. A gradient method using HPLC (ODS C18, 4.6 × 250 mm, 335 nm, 385 nm) with a fluorescence detector and using 0.1% trifluoroacetic acid-added distilled water / 0.08% trifluoroacetic acid-added 80% acetonitrile as the mobile phase (Miyata, T et al., J. Am. Soc., Nephro 1., 7, 1198-1206, 1996; Miyata) , T., et al., Proc. Natl. Acad. Sci., USA, 93 2353-2358, 1996).

  In order to examine the AGEs production inhibitory effect, the ratio of the above-mentioned pentosidine production amount when the pentosidine production amount by the control is 100% is calculated as the pentosidine production rate (%), and from this value, the pentosidine production inhibition rate (AGEs) The production inhibition rate (%) was determined. The results are shown in Table 15. As can be seen from these results, all of the test compounds (1) to (20) exhibited a pentosidine production inhibitory action.

  Pentosidine is one of the AGEs structures, and the phenylene derivative (I) of the present invention suppresses the production of pentosidine, and thus has been found to have an AGEs production inhibitory effect. Thus, the phenylene derivative (I) provided by the present invention has an AGEs production inhibitory action and can be effectively used for the treatment or improvement of schizophrenia.

Test Example 3 AGEs Production Inhibitory Effect of Phenylene Derivative (II) The following test compounds belonging to the phenylene derivative represented by the general formula (II) in the present invention were examined for AGEs production inhibitory action by the same method as in Test Example 1.
(II-1) 3- {N- (3-phenylpropionyl) -N- [2 ′-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-6) 3- {N-pentyl-N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-7) 1- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -2-methyl-1,2,3,4-tetrahydroquinoline-7-carboxylic acid,
(II-8) 5- [4- (N-methylsulfonyl-N-phenylaminomethyl) phenyl] -1,3-thiazolidine-2,4-dione,
(II-9) 5- [4- (N-butylsulfonyl-N-phenylaminomethyl) phenyl] -1,3-thiazolidine-2,4-dione,
(II-10) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (methylsulfonyl) amino} benzoic acid,
(II-11) 3- {N-butylsulfonyl-N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-15) 5- {4- [2- (2-pyridyl) benzimidazol-1-ylmethyl] phenyl} -1,3-thiazolidine-2,4-dione,
(II-16) 5- [4- (2-phenylimidazol-1-ylmethyl) phenyl] -1,3-thiazolidine-2,4-dione,
(II-17) 1- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-3-propyl-1,2-dihydroquinoline-7-carboxylic acid,
(II-18) 3- {N-cyclohexanecarbonyl-N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-19) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (pyridine-3-carbonyl) amino} benzoic acid,
(II-20) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N- (3-phenylpropionyl) amino} benzoic acid,
(II-21) 3- {N- (biphenyl-4-carbonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-22) 3- {N- (phenylsulfonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-23) 3- {N- (4-methylphenylsulfonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl) amino) benzoic acid,
(II-24) 3- {N- (biphenyl-4-sulfonyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-25) 3- {N- (2-naphthylsulfol) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid,
(II-27) 1- [4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-3-propyl-2,3-1H-indole-6-carboxylic acid ,
(II-28) 1- [4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-3-propyl-2,3,4,5-tetrahydro-1H- Benzo [b] azepine-8-carboxylic acid,
(II-29) 3,3-Dimethyl-1- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -2-oxo-1,2,3,4-tetrahydroquinoline -7-carboxylic acid,
(II-30) 1 ′-[4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -2′-oxo-1 ′, 4′-dihydro-2′H-spiro [ Cyclopropane-1,3′-quinoline] -7′-carboxylic acid,
(II-32) 3- {N-phenethyl N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-33) 3- {N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] -N-[(5-methyl-2-phenyl 1,3-oxazole- 4-yl) acetyl] amino} benzoic acid,
(II-36) 3- {N- (3-cyclohexylpropanoyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-42) 3- {N- (2-oxo-2-phenylethyl) -N- [2 '-(1H-tetrazol-5-yl) biphenyl-4-ylmethyl] amino} benzoic acid,
(II-63) 3- {N- [4- (2,4-Dioxo-1,3-thiazolidin-5-yl) benzyl] -N- [4- (2-methylthiazol-4-yl) benzenesulfonyl Amino} benzoic acid,
(II-68) 3- {N- (biphenyl-4-ylmethyl) -N- [4- (2,4-dioxo-1,3-thiazolidin-5-yl) benzyl] amino} benzoic acid.

  The results are shown in Table 16. As can be seen from the results, all of the test compounds (21) to (20) exhibited a pentosidine production inhibitory action.

  From this result, it was found that the phenylene derivative (II) of the present invention suppresses the production of pentosidine, and thus has an AGEs production inhibitory effect. Thus, the phenylene derivative (II) provided by the present invention has an AGEs production inhibitory action and can be effectively used for the treatment or improvement of schizophrenia.

Formulation Example 1 Capsule Test Compound ^* 10 mg
Lactose 100 mg
Corn starch 58 mg
Magnesium stearate 2 mg
180 mg total
* Compound (1), compound (2), phenylene derivative (I) or (II).

Mix the powder of each component shown above well and pass through a 60 mesh sieve (mesh standard is Tyler standard). 180 mg of the resulting powder is weighed out and filled into gelatin capsules (No. 3) to prepare a therapeutic or ameliorating agent for schizophrenia in capsule form.
Formulation Example 2 Tablet Test Compound ^* 10 mg
Lactose 85 mg
Corn starch 34 mg
Microcrystalline cellulose 10 mg
Magnesium stearate 1 mg
150 mg total
* Compound (1), compound (2), phenylene derivative (I) or (II).

The powder of each component shown above is mixed well and compressed into tablets each weighing 150 mg. If necessary, these tablets may be coated with sugar or film. Thus, a tablet form treatment or amelioration agent for schizophrenia is prepared.
Formulation Example 3 Granule test compound ^* 10 mg
Lactose 839 mg
Corn starch 150 mg
Hydroxypropylcellulose 1 mg
1000 mg total
* Compound (1), compound (2), phenylene derivative (I) or (II).

  A powder of each component shown above is mixed well, moistened with pure water, atomized with a basket granulator, and dried to prepare a therapeutic or ameliorating agent for schizophrenia in the form of granules.

SEQ ID NO: 4 is 1-200 upstream of adenine at position 201 of the base sequence (SEQ ID NO: 1) of the glyoxylase I (GLO-I) gene (corresponding to position 79 of the base sequence of the coding region (SEQ ID NO: 2)) The base sequence of the region is shown.
SEQ ID NO: 5 shows the base sequence of the 203-502 region downstream of cytosine at position 202 (corresponding to position 80 of the base sequence of the coding region (SEQ ID NO: 2)) in the base sequence of the GLO-I gene (SEQ ID NO: 1) .
SEQ ID NO: 6 shows the nucleotide sequence of the 154-453 region upstream of adenine at position 454 of the nucleotide sequence (SEQ ID NO: 1) of the GLO-I gene (corresponding to position 332 of the nucleotide sequence of the coding region (SEQ ID NO: 2)) .
SEQ ID NO: 7 shows the nucleotide sequence of the 455-754 region downstream of adenine at position 454 of the nucleotide sequence (SEQ ID NO: 1) of the GLO-I gene (corresponding to position 332 of the nucleotide sequence of the coding region (SEQ ID NO: 2)) .
SEQ ID NO: 8 shows the base sequence of the forward primer used for the PCT-direct sequence for detecting insertion mutation between 79 and 80 in the base sequence (SEQ ID NO: 2) of the coding region of GLO-I gene.
SEQ ID NO: 9 shows the base sequence of the reverse primer used for the PCT-direct sequence for detecting insertion mutation between 79-80 in the base sequence (SEQ ID NO: 2) of the coding region of GLO-I gene.
SEQ ID NO: 10 shows the base sequence of the forward primer used for the PCT-direct sequence for detecting the base substitution mutation at position 332 in the base sequence of the coding region of the GLO-I gene (SEQ ID NO: 2).
SEQ ID NO: 11 shows the base sequence of the reverse primer used for the PCT-direct sequence for detecting the base substitution mutation at position 332 in the base sequence (SEQ ID NO: 2) of the coding region of the GLO-I gene.

Claims (22)

For the biological sample of the subject,
(1) Amount of protein modification product (carbonyl protein modification product) of a carbonyl compound in a biological sample,
(2) expression level or activity of glyoxylase I in a biological sample,
(3) a genetic abnormality of the glyoxylase I gene, and
(4) measuring at least one selected from the group consisting of the amount of pyridoxal in the biological sample,
Testing method for schizophrenia. The test method for schizophrenia according to claim 1, comprising the following steps (A) and (B):
(A) For a biological sample of a subject, (1) the amount of a carbonyl compound protein modification product (carbonyl protein modification product) in the biological sample, (2) the expression level or activity of glyoxylase I in the biological sample, (3) a genetic abnormality of the glyoxylase I gene, and (4) measuring at least one selected from the group consisting of the amount of pyridoxal in the biological sample, and
(B) A step of comparing the measured value in the subject obtained in the step (A) with the measured value (control value) in a healthy subject corresponding to the measured value. When each measurement value measured with respect to the biological sample of the subject falls under the following, it is determined that the subject is suffering from schizophrenia or has a high risk of developing it in the future. Testing method for schizophrenia:
(1) The amount of the protein modification product (carbonyl protein modification product) of the carbonyl compound in the biological sample of the subject is greater than the amount in the biological sample of the healthy subject.
(2) The expression level or activity of glyoxylase I in the biological sample of the subject is lower than the expression level or activity in the biological sample of a healthy person,
(3) The glyoxylase I gene in the subject has an abnormality that causes a decrease in glyoxylase I activity
(4) The amount of pyridoxal in the subject's biological sample is less than that in the healthy subject's biological sample.   The method for examining schizophrenia according to any one of claims 1 to 3, further comprising the step of (5) measuring the amount of homocysteine in the biological sample in addition to the measurement of the amount of pyridoxal in the biological sample. .   5. The subject according to any one of claims 1 to 4, wherein the subject has symptoms or findings characteristic of schizophrenia as defined in diagnostic criteria for schizophrenia, or is suspected of having schizophrenia according to the criteria. The test method for schizophrenia to be described.   The test method for schizophrenia according to any one of claims 1 to 5, wherein the subject is a person who does not have renal dysfunction, diabetes or inflammation.   (3) An adenine insertion between adenine at position 79 and cytosine at position 80 in the base sequence of the coding region of the glyoxylase I gene shown in SEQ ID NO: 1 (SEQ ID NO: 2) The method for examining schizophrenia according to any one of claims 1 to 6, which is a frame shift abnormality caused by the above.   (3) The gene abnormality of the glyoxylase I gene is a base substitution mutation that causes a mutation of the 111st amino acid (Glu → Ala) in the amino acid sequence of glyoxylase I shown in SEQ ID NO: 3. The inspection method of schizophrenia described in any one of.   A step of measuring at least one selected from the group consisting of (1), (2) and (4) above for a subject having a glyoxylase I gene abnormality leading to a decrease in glyoxylase I activity A test method for schizophrenia according to any one of claims 1 to 6.   The above-mentioned glyoxylase I gene abnormality is a frameshift abnormality caused by adenine insertion between the 79th adenine and the 80th cytosine in the base sequence of the coding region of the glyoxylase I gene shown in SEQ ID NO: 1 (SEQ ID NO: 2). Or the method for examining schizophrenia according to claim 9, which is a base substitution mutation that causes a mutation of the 111st amino acid (Glu → Ala) in the amino acid sequence of glyoxylase I shown in SEQ ID NO: 3.   The test method for schizophrenia according to any one of claims 1 to 10, comprising a step of measuring the amount of (1) a protein modification product (carbonyl protein modification product) of a carbonyl compound in a subject, A method characterized by using an immunological technique using an anti-carbonyl protein-modified antibody, high performance liquid chromatography, gas chromatography / mass spectrometry, or AGE-Reader method as a method for measuring the modified product.   The method for examining schizophrenia according to any one of claims 1 to 11, wherein the modified carbonyl protein is pentosidine. In the glyoxylase I gene shown in SEQ ID NO: 1, a continuous oligo or polynucleotide having a length of 16 bases or more containing adenine at position 79 and cytosine at position 80 in the base sequence of the coding region (SEQ ID NO: 2), or position 332 Consecutive oligos or polynucleotides having a length of 16 bases or more including the bases of the above (provided that these oligonucleotides or polynucleotides are RNA, the base symbol “t” in the sequence listing should be read as “u”) And 15-35 nucleotides of oligonucleotide used to specifically amplify the oligo or polynucleotide, or a label thereof,
For the subject, in the base sequence of the coding region of the glyoxylase I gene shown in SEQ ID NO: 1 (SEQ ID NO: 2), insertion of adenine between adenine at position 79 and cytosine at position 80, or from adenine at position 332 to cytosine A primer used to identify mutations and test for schizophrenia. In the glyoxylase I gene shown in SEQ ID NO: 1, a continuous oligo or polynucleotide having a length of 16 bases or more containing adenine at position 79 and cytosine at position 80 in the base sequence of the coding region (SEQ ID NO: 2), or position 332 A continuous oligo or polynucleotide having a base length of 16 bases or more (provided that these oligonucleotides or polynucleotides are RNA, the base symbol “t” in the sequence listing should be read as “u”) And an oligonucleotide having a length of 16 to 500 bases used for specifically detecting the oligo or polynucleotide, or a labeled product thereof,
For subjects, adenine insertion between position 79 and position 80 cytosine in the nucleotide sequence of the glyoxylase I gene sequence shown in SEQ ID NO: 1 (SEQ ID NO: 2) or mutation from position 332 adenine to cytosine Probe used to identify and test for schizophrenia.   A test reagent for schizophrenia comprising the primer according to claim 13 and / or the probe according to claim 14, or a kit containing the reagent.   (2) A reagent used in the examination of schizophrenia having a step of measuring the expression level or activity of glyoxylase I in a biological sample of a subject, and at least an anti-glyoxylase I antibody or glyoxylase I reaction A test reagent containing a substrate.   (1) A reagent used for testing schizophrenia having a step of measuring the amount of a protein modification product (carbonyl protein modification product) of a carbonyl compound in a biological sample of a subject, (a) carbonyl protein modification at a known concentration Or (b) an anti-carbonyl protein-modified antibody that may be labeled.   The test reagent according to claim 17, wherein the modified carbonyl protein is pentosidine.   (4) a reagent for use in a test for schizophrenia comprising a step of measuring the amount of pyridoxal in a biological sample of a subject, wherein (a) at least one of a known concentration of pyridoxal and (b) a pyridoxal measurement reagent Containing a test reagent.   An AGE-Reader device for use in diagnosing schizophrenia, comprising means for measuring the amount of a protein modification product (carbonyl protein modification product) of a carbonyl compound in the skin of a subject.   The AGE-Reader device according to claim 20, to which a document describing that it can be used for diagnosis of schizophrenia is attached.   Use of an AGE-Reader device for diagnosing schizophrenia having means for measuring the amount of a protein modification product (carbonyl protein modification product) of a carbonyl compound in the skin of a subject.

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