JP2008143848A - Method for predicting diuretic sensitivity in hypertensive patients. - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for selecting and determining a hypertensive patient on whom a diuretic takes effect well based on a gene polymorphism associated with the efficacy of a hypoglycemic agent with the diuretic as an active ingredient, and to provide a reagent and a reagent kit useful for easily conducting the method. <P>SOLUTION: The method comprises using such a criterion that a hypertensive patient in question has the following genotypes (1) to (4) for at least one gene as mentioned below: (1) Glutamate-cysteine ligase (C588T); genotype 1; (2) CD18 (C1323T): genotype 23; (3) Cytochrome P450 2J2*7 (G-50T): genotype 1; and (4) Alcohol dehydrogenase 2 (Arg 47 His (G/A)): genotype 1. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to an antihypertensive agent comprising a diuretic used as an active ingredient for a specific hypertensive patient having an inherent genetic polymorphism. The present invention also relates to a method for detecting the effectiveness of an antihypertensive agent containing a diuretic as an active ingredient for individual hypertensive patients based on genetic information of the hypertensive patient, and a material used for carrying out the method.

  The method of the present invention provides effective and accurate therapeutic guidelines for treating and ameliorating hypertension to hypertensive patients and medical staff, and as a result of being able to select a therapeutic agent suitable for each individual patient, It is useful in that effective and accurate treatment can be performed and the mental, physical and economic burden caused by invalid treatment can be avoided.

  Hypertension and its resulting diseases are the leading cause of death worldwide, and one in four adults, equivalent to about 1 billion people worldwide, suffer from these diseases. It has been broken. In particular, the risk of cardiovascular complications due to hypertension in diabetic patients is two to four times higher than in patients without diabetes, and the International Diabetes Federation advocates the need for strict blood pressure control.

  However, despite various treatments, about 70% of patients with hypertension, and nearly 90% of patients with diabetes and hypertension have not achieved the target blood pressure reduction. Is the actual situation. Although it is recognized that hypertension is a major risk factor for cardiovascular diseases such as stroke and myocardial infarction, it is less likely that blood pressure is not adequately controlled (especially masked hypertension) unknown. In fact, about 90% of hypertensive patients who developed cerebral infarction reported blood pressure treatment before the onset, indicating that this was caused by a lack of antihypertensive treatment. . In addition, clinical trial results have shown that lowering systolic blood pressure by 10 mmHg can reduce the onset of stroke by about 40%, and this also indicates that “10 mmHg higher” is a very significant risk. I understand that.

  In order to prevent such cardiovascular complications caused by high blood pressure, regular examinations are conducted from an early stage so that patients and healthcare professionals have accurate knowledge about hypertension care and provide accurate and effective treatment ( Non-pharmacotherapy, pharmacotherapy) is necessary.

Currently, the following six drugs are widely used for drug treatment of hypertension depending on the mechanism of action:
(A) diuretics,
(B) a calcium antagonist,
(C) an angiotensin receptor antagonist,
(D) angiotensin converting enzyme inhibitor,
(E) β-blocker,
(F) Alpha blocker.

  However, when these antihypertensive drugs (antihypertensive agents) are administered to hypertensive patients, those who are highly effective (good responders), those who are less effective (poor responders), or those who have no effect (non) Patient responses vary, such as responders, and there is a recognized need to accurately select and prescribe drugs that are appropriate for individual patients. However, there is no objective selection criterion in determining the types and dosages of these drugs, and they are highly dependent on the empirical judgment of doctors. Various side effects have been pointed out depending on the type of drug. Therefore, establishment of a system for prescribing an appropriate antihypertensive drug (hypertensive agent) according to the patient is extremely important in medicine. In particular, in recent years, based on the philosophy of Evidence Based Medicine (EBM), in addition to doctors' expertise, experience, and technology, evidence (evidence) on the latest and best medical technology confirmed by scientific methods has also been obtained. In addition, since the most effective and safe medical care is required for individual patients, effective patient groups for the administration of various antihypertensive drugs are based on scientific evidence. It is important to make clear. By doing so, the patient's compliance can be improved, an effective therapeutic effect can be obtained, and the medical expenses can be efficiently operated.

  Recently, it has been reported that onset of hypertension involves not only non-genetic factors such as diet and lifestyle habits including obesity, but also some genetic factors (for example, Non-Patent Documents 1 to 4). ). For example, as genes related to hypertension (hypertension-related genes), beta2 bradykinin receptor (C-58T) (Non-patent document 1), endothelin-1 (Lys198Asn) (non-patent document), antitensinogen (M235T) (Non-patent document 3) ), UCP2 (-866 G / A) (Non-Patent Document 4), etc. have been identified and their polymorphisms [eg SNPs (single nucleotide polymorphisms)] and hypertension have been reported Has been.

In addition, it has been reported that there are differences between individuals and side effects in the expression of drug efficacy due to differences in gene polymorphisms that are directly involved in the mechanism of action of specific drugs (for example, Non-Patent Documents 5-6). However, it is not known that genetic polymorphisms of genes other than hypertension-related genes reported in these reports are selection indicators for evaluating the effectiveness of various antihypertensive agents. Based on this, there is still no idea of selecting an effective antihypertensive agent suitable for the patient.
Wang B, et al. Genetic variation in the promoter region of the beta2 bradykinin receptor gene is associated with essential hypertension in a Chinese Han population.Hypertens Res. 2001 May; 24 (3): 299-302. Jin JJ, et al. Association of endothelin-1 gene variant with hypertension. Hypertension. 2003 Jan; 41 (1): 163-7. Hunt SC, et al. Angiotensinogen genotype, sodium reduction, weight loss, and prevention of hypertension: trials of hypertension prevention, phase II.Hypertension. 1998 Sep; 32 (3): 393-401. Ji Q, et al. A common polymorphism of uncoupling protein 2 gene is associated with hypertension.J Hypertens. 2004 Jan; 22 (1): 97-102. Schelleman H, et al. Drug-gene interaction between the insertion / deletion polymorphism of the angiotensin-converting enzyme gene and antihypertensive therapy. Ann Pharmacother. 2006 Feb; 40 (2): 212-8. Bleumink GS, et al. Mortality in patients with hypertension on angiotensin-I converting enzyme (ACE) -inhibitor treatment is influenced by the ACE insertion / deletion polymorphism.Pharmacogenet Genomics. 2005 Feb; 15 (2): 75-81.

  The present invention provides a gene polymorphism related to the effectiveness of an antihypertensive agent containing a diuretic as an active ingredient, and selects and determines a hypertensive patient in which the diuretic effectively responds based on this information. Objective. Another object of the present invention is to provide a reagent and a reagent kit useful for carrying out the method easily.

  A further object of the present invention is to provide a method for preselecting hypertensive patients in which diuretics are effective and effectively treating or improving the patients.

  The inventors of the present invention have been diligently studying to achieve the above object, and have found that polymorphisms that define drug responsiveness, particularly responsiveness to diuretics, in hypertensive patients, particularly hypertensive patients with diabetes. We succeeded in identifying the genetic polymorphism. Specifically, among the hypertensive patients having diabetes, the present inventors have (1) a patient having C (cytosine) as a homozygote (CC) at position 588 of the Glutamate-cystein ligase gene; (2) CD18 Patients with T (thymine) as a heterozygote (TC or CT) or homozygote (TT) at position 1323 of the gene; (3) G (guanine) at position -50 of the Cytochrome P450 2J2 * 7 gene It was found that patients having a body (GG); and (4) patients whose 47th amino acid of Alcohol dehydrogenase 2 is homozygous of His (histidine) can significantly reduce blood pressure by diuretics.

  From these findings, by examining the genotype of any of the genes (gene polymorphisms) of (1) to (4) above for individual patients with hypertension, The administration effectiveness can be determined, and the drug can be selected and determined as an effective antihypertensive agent for the patient (in other words, for a hypertensive patient, the patient has an antihypertensive effect with a diuretic as an active ingredient) Effective and accurate hypertension treatment by selectively administering the drug to a patient whose administration effectiveness has been determined (administration effective person). As a result, the present invention was completed.

That is, the present invention has the following aspects.
I. Antihypertensive agent with diuretic as active ingredient I-1. A diuretic is an active ingredient, characterized in that it is administered to a hypertensive patient who has been confirmed to have the following genotype for at least one gene of any one of (1) to (4) Antihypertensive agent:
Gene (gene polymorphism) : genotype
(1) Glutamate-cystein ligase (C588T) ： 1
(2) CD18 (C1323T): 23
(3) Cytochrome P450 2J2 * 7 (G-50T): 1
(4) Alcohol dehydrogenase 2 (Arg47His (G / A)): 1.

  I-2. The antihypertensive agent according to I-1, wherein the hypertensive patient is a patient having diabetes.

  I-3. The blood pressure according to I-1 or I-2, wherein the diuretic is at least one drug selected from the group consisting of thiazide diuretics, thiazide analog diuretics, loop diuretics and K-retaining diuretics Depressant.

  I-4. The thiazide diuretics are trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, indapamide, chlorthalidone, and pharmaceutically acceptable salts thereof; thiazide-like diuretics are trypamide, methiclan, mefluside, and pharmaceutically acceptable thereof The blood pressure described in I-3, wherein the loop diuretic is furosemide, and pharmaceutically acceptable salts thereof; the K-retaining diuretic is spironolactone, triamterene, and pharmaceutically acceptable salts thereof. Depressant.

II. A method for detecting the efficacy of antihypertensive drugs containing diuretics as active ingredients in hypertensive patients
II-1. A method for detecting the administration effectiveness of an antihypertensive agent comprising a diuretic as an active ingredient for a hypertensive patient, wherein the hypertensive patient has the following genotype for at least one of the genes (1) to (4): ) As an indicator:
Gene (gene polymorphism) : genotype
(1) Glutamate-cystein ligase (C588T) ： 1
(2) CD18 (C1323T): 23
(3) Cytochrome P450 2J2 * 7 (G-50T): 1
(4) Alcohol dehydrogenase 2 (Arg47His (G / A)): 1.

II-2. The method described in II-1 having the following steps:
(A) detecting a genotype in a genetic polymorphism of at least one gene described in (1) to (4) above for a biological sample of a hypertension patient;
(B) A step of discriminating whether the genotype in the polymorphism of at least one gene described in (1) to (4) is the genotype described in II-1.

II-3. The method according to II-2, further comprising the following step (c):
(c) Based on the result of (b) above, when the genotype in the genetic polymorphism of at least one gene described in (1) to (4) matches the genotype described in II-1 above And a step of determining that administration of an antihypertensive agent containing a diuretic as an active ingredient is effective for the treatment of the hypertensive patient.

  II-4. The method according to any one of II-1 to II-3, wherein the hypertensive patient is a patient having diabetes.

  II-5. The diuretic is at least one drug selected from the group consisting of thiazide diuretics, thiazide-like diuretics, loop diuretics, and K-retaining diuretics, any of II-1 to II-4 How to describe.

  II-6. The thiazide diuretics are trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof; thiazide-like diuretics are indapamide, chlorthalidone, tripamide, methiclan, mefluside, and pharmaceutically acceptable salts thereof The method according to II-5, wherein the loop diuretic is furosemide, and pharmaceutically acceptable salts thereof; the K-retaining diuretic is spironolactone, triamterene, and pharmaceutically acceptable salts thereof. .

III. A tool for detecting the efficacy of antihypertensive drugs containing diuretics as active ingredients in hypertensive patients
III-1. (1) Glutamate-cystein ligase (C588T), (2) CD18 (C1323T), (3) Cytochrome P450 2J2 * 7 (G-50T), and (4) Alcohol dehydrogenase 2 (Arg47His (G / A)) An array for detecting administration efficacy of an antihypertensive agent comprising a diuretic as an active ingredient for a hypertensive patient, comprising a probe for detecting a genetic polymorphism of at least one gene selected from the group.

  III-2. The array according to III-1, wherein the hypertensive patient is a patient having diabetes.

  III-3. The array according to III-1 or III-2, wherein the diuretic is at least one drug selected from the group consisting of thiazide diuretics, thiazide-like diuretics, loop diuretics, and K-retaining diuretics .

  III-4. The thiazide diuretics are trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof; thiazide-like diuretics are indapamide, chlorthalidone, tripamide, methiclan, mefluside, and pharmaceutically acceptable salts thereof The array described in III-3, wherein the loop diuretic is furosemide, and pharmaceutically acceptable salts thereof; the K-retaining diuretic is spironolactone, triamterene, and pharmaceutically acceptable salts thereof. .

  III-5. (1) Glutamate-cystein ligase (C588T), (2) CD18 (C1323T), (3) Cytochrome P450 2J2 * 7 (G-50T), and (4) Alcohol dehydrogenase 2 (Arg47His (G / A)) About a hypertensive patient comprising a primer pair that can specifically amplify a region having a gene polymorphism of at least one gene selected from the group, and / or a nucleic acid probe that specifically hybridizes to the gene or its amplification product A reagent kit for detecting the administration effectiveness of an antihypertensive agent comprising a diuretic as an active ingredient.

  III-6. The reagent kit according to III-5, wherein the hypertensive patient is a patient with diabetes.

  III-7. A reagent kit according to III-5 or III-6, comprising the array according to any one of III-1 to III-4.

  III-8. Any of III-5 to III-7, wherein the diuretic is at least one drug selected from the group consisting of thiazide diuretics, thiazide-like diuretics, loop diuretics, and K-retaining diuretics Reagent kit to be described.

  III-9. The thiazide diuretics are trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof; thiazide-like diuretics are indapamide, chlorthalidone, tripamide, methiclan, mefluside, and pharmaceutically acceptable salts thereof The reagent according to III-8, wherein the loop diuretic is furosemide, and pharmaceutically acceptable salts thereof; the K-retaining diuretic is spironolactone, triamterene, and pharmaceutically acceptable salts thereof kit.

  According to the present invention, for hypertensive patients, particularly hypertensive patients with diabetes, (1) Glutamate-cystein ligase (C588T), (2) CD18 (C1323T), (3) Cytochrome P450 2J2 * 7 (G-50T), Or (4) By examining the genotype of at least one gene of Alcohol dehydrogenase 2 (Arg47His (G / A)), the effectiveness of the hypertensive patient with respect to a diuretic can be evaluated. For patients who are judged to be effective, the antihypertensive agent can be treated accurately and effectively according to the individual patient by administering an antihypertensive agent containing diuretics as the active ingredient. It becomes. On the other hand, while reducing the burden of treatment costs (medical expenses) caused by unnecessary (ineffective) treatment that is not suitable for the patient, physical and mental due to side effects that can occur due to ineffective treatment It is also effective in reducing physical distress and preventing the progression and complications of hypertension due to delayed treatment.

(1) Explanation of terms The abbreviations of base sequences (nucleotide sequences), nucleic acids and the like in this specification are defined by IUPAC-IUB [IUPAc-IUB communication on Biological Nomenclature, Eur. J. Biochem., 138; 9 ( 1984)], “Guidelines for the preparation of specifications including base sequences or amino acid sequences” (edited by the Patent Office) and conventional symbols in the field.

  In the present specification, unless otherwise specified, “gene” refers to double-stranded DNA including human genomic DNA, single-stranded DNA (sense strand), and single-stranded DNA having a sequence complementary to the sense strand ( Antisense strands) and fragments thereof are included, and the term “gene” in the present specification indicates the regulatory region, coding region, exon, and intron without distinction unless otherwise specified. And

  (1) Glutamate-cystein ligase gene, (2) CD18 gene, (3) Cytochrome P450 2J2 * 7 gene, and (4) Alcohol dehydrogenase 2 gene described in this specification (base sequence, chromosome position, etc.) Moreover, the positional information regarding a gene polymorphism is based on the literature shown in FIG.

  In this specification, (1) “Glutamate-cystein ligase (C588T)” means that the Glutamate-cystein ligase gene has a gene polymorphism in which the 588th base of the base sequence is C or T. Similarly, (2) “CD18 (C1323T)” means that the CD18 gene has a gene polymorphism in which the base at position 1323 is C or T: (3) “Cytochrome P450 2J2 * 7 (G -50T) "means that the Cytochrome P450 2J2 * 7 gene has a gene polymorphism in which the -50th base of the nucleotide sequence is G or T: and (4)" Alcohol dehydrogenase 2 (Arg47His (G / A )) "Means that the Alcohol dehydrogenase 2 gene is G (forms codon" CGC ") or A (codon) so that the 47th amino acid of Alcohol dehydrogenase 2 (protein) becomes Arg or His. Each having a genetic polymorphism of (forming “CAC”).

  “Gene polymorphism” means the diversity of genes in which two or more alleles (alleles) exist at one locus in the same population. Specifically, it indicates a mutation of a gene that exists at a certain frequency or more in a certain population. The gene mutation referred to here is not limited to the region transcribed as RNA, but includes mutations in all DNAs that can be identified on the human genome including regulatory regions such as promoters and enhancers. 99.9% of human genomic DNA is common among individuals, and the remaining 0.1% is responsible for such diversity, and is involved in individual differences in susceptibility to specific diseases, responsiveness to drugs and environmental factors. obtain. A genetic polymorphism does not always make a difference in phenotype. SNP (single nucleotide polymorphism) is a kind of gene polymorphism, but the gene polymorphism targeted by the present invention is not limited to this.

  In the genotype shown in the present specification, “1” is a polymorphic homozygote having a base that precedes the substituted bases in alphabetical order (A, C, G, T), and “2” is Hetero, “3” represents a polymorphic homozygote having a base that follows in alphabetical order of the bases among the substituted bases. For example, when the target gene (gene polymorphism) is Glutamate-cystein ligase (C588T), the genotype is genotype 1, CT or TC heterogeneous. The case of type 2 and TT homozygous is called genotype 3. As another example, when the gene (gene polymorphism) is Alcohol dehydrogenase 2 (G / A), when the genotype is homologous to AA, it is heterogeneous to genotype 1, AG or GA Is referred to as genotype 2 and GG homozygous is referred to as genotype 3. “12” represents both genotypes of genotypes 1 and 2, and “23” represents both genotypes of 2 and 3 genotypes.

  Further, in this specification, “nucleotide”, “oligonucleotide” and “polynucleotide” are synonymous with nucleic acid and include both DNA and RNA. These may be double-stranded or single-stranded, and in the case of “nucleotide” having a certain sequence (or “oligonucleotide”, “polynucleotide”), it is complementary to this unless otherwise specified. “Nucleotide” (or “oligonucleotide” and “polynucleotide”) having a typical sequence is also intended to be inclusive. Furthermore, when the “nucleotide” (or “oligonucleotide” and “polynucleotide”) is RNA, the base symbol “T” shown in the sequence listing shall be read as “U”.

  Table 1 shows the classification of hypertension prescribed by the Japanese Society of Hypertension and WHO / ISH.

  Hypertensive patients targeted by the present invention include patients belonging to mild hypertension, moderate hypertension, severe hypertension, and systolic hypertension in the above-mentioned hypertension classification.

The term “diabetes” as used in the present invention includes both type 2 diabetes (non-insulin dependent diabetes) and type 1 diabetes (insulin dependent diabetes). According to the guidelines of the Japan Diabetes Society (1999), if the blood glucose level is 200 mg / dL or higher, the fasting blood glucose level is 126 mg / dL or higher, and the 75 g oral glucose tolerance test 2 hour value is 200 mg / dL or higher It can be judged as diabetes. In addition, if the above criteria are met twice on a different day, or if there is a characteristic symptom of diabetes even after one check, HbA _1c (hemoglobin A _1c ) is 6.5% or more, or diabetic retinopathy If there is, it can be judged as diabetes. The diabetic patient targeted by the present invention includes a patient in a border zone (border type diabetic patient). In addition, when the fasting blood glucose level is between 110 and 125 mg / dl, or when the glucose tolerance test 2 hour value is between 140 and 199 mg / dl, it can be determined as borderline diabetes.

(2) Method for determining administration efficacy of antihypertensive agent containing diuretic as active ingredient The method for determining the administration effectiveness of antihypertensive agent containing diuretic as an active ingredient according to the present invention has hypertension patients, particularly diabetes. Whether blood pressure can be lowered by administering antihypertensive agents containing diuretics as an active ingredient to the patient according to the genetic polymorphism (genotype-considering genotype) of hypertensive patients It is a method of determining whether or not. A method for determining the effectiveness of administration of a hypotensive agent containing a diuretic as an active ingredient according to the embodiment of the present invention will be described with reference to the flowchart shown in FIG. Each processing described below includes a CPU, a memory, a recording device (for example, a hard disk), an input device (for example, a keyboard, a mouse, a flexible disk drive, a CD-ROM drive), a display device (for example, a liquid crystal display), a communication device ( For example, the description will be made assuming that a computer equipped with a network board) is used. That is, the processing target data is input via the input device and the communication device and recorded in the recording device, and the CPU executes processing on the data read from the recording device using the memory as a work area. The intermediate result of the process and the final result are recorded in a predetermined area of the recording unit as necessary.

  First, in step S1, an input of genetic information of a hypertension patient is accepted and the input data is recorded in a recording device. Here, the genetic information includes genotype information, for example, a pair of text data representing the SNP name and text data representing the genotype. The gene information may be input as a code. For example, a code (text data) assigned to each SNP name and each genotype may be input. The genetic information may be input via a keyboard, may be input via a read drive from a state recorded on a recording medium, or may be input via a communication line. Furthermore, the genetic information may be input by being selected from a plurality of candidates displayed on the display device, or may be input using other known methods.

  In step S2, a table of genetic polymorphism information is read from the recording unit. An example of a table recorded in advance in the recording device is shown in FIG. As will be described later, this is a table of genetic polymorphism information indicating the administration effectiveness of a blood pressure lowering agent containing a diuretic as an active ingredient for a hypertensive patient, particularly a hypertensive patient having diabetes. In the table of FIG. 3, the name of the SNP (SNP NAME) and the genotype (GENOTYPE) (according to the numerical notation shown in the explanation of the above terms) are described. In this case, the name (SNP NAME) and genotype (GENOTYPE) of the SNP correspond to the gene polymorphism information. In other words, it is assumed that the data shown in FIG. 3 and the data indicating the corresponding relationship are recorded as text data, for example, in the recording device.

  In step S3, it is determined whether the genetic polymorphism information in the table read in step S2 is included in the patient genetic information input in step S1, and the result is temporarily stored in a predetermined area of the memory. Record. For example, in the case of FIG. 3, “Glutamate-cysteine ligase (C588T)” and “1” in the first line are read, the SNP name is “Glutamate-cysteine ligase (C588T)”, and the genotype is “1”. It is determined whether the polymorphism information is included in the patient's genetic information. If it is included, for example, “1” is set in a flag corresponding to the genetic polymorphism information in the first row (assumed that the initial value is set to “0” on the memory). If not included, do nothing for the flag and read “CD18 (C1323T)” and “23” in the second line, and check whether they are included in the patient's genetic information. If it is determined, if it is included, “1” is set to a flag corresponding to the genetic polymorphism information in the second row (assumed that the initial value is set to “0” on the memory). If not included, the following genetic polymorphism information is similarly processed. In this manner, whether or not the four gene polymorphism information in the table of FIG. 3 is included in the patient's gene information is temporarily stored as data set in the four flags on the memory, for example. .

  Here, it may be determined whether or not all genetic polymorphism information in the table is included in the patient genetic information. However, if at least one genetic polymorphism information is included in the patient genetic information. For example, you may transfer to step S4, without processing regarding the remaining gene polymorphism information.

  In step S4, according to the determination result in step S3, whether or not a blood pressure lowering agent containing diuretic as an active ingredient is effective for the patient is displayed (for example, displayed on a display device). For example, if at least one of the above four flag data is “1”, it indicates that it is valid, and if all the flag data is “0”, it indicates that it is not valid.

  By the above processing, according to genetic information possessed by a specific hypertensive patient, the effectiveness of administering a blood pressure lowering agent containing a diuretic as an active ingredient to the patient, that is, lowering the blood pressure of the patient. Whether or not is possible. Therefore, tailor-made treatment for each patient is possible.

  The table used for the determination is not limited to the table shown in FIG. 3, and may be any table including information on the SNP name (SNP NAME) and genotype (GENOTYPE) shown in FIG. 3.

  Further, although the embodiment of the present invention has been described as processing performed by a computer using electronic data corresponding to FIG. 3 according to the flowchart shown in FIG. 2, the present invention is not limited to this. It suffices to use the table shown in FIG. 3 or a table containing genetic polymorphism information equivalent to this (both are not limited to electronic data), and doctors and others can add the table shown in FIG. It is also possible to determine whether the name (SNP NAME) and genotype (GENOTYPE) of the described SNP are included, and to determine the effectiveness of an antihypertensive agent containing a diuretic as an active ingredient for the patient it can.

(3) Antihypertensive agent having administration efficacy according to Genotype The table shown in FIG. 3 is a table showing gene polymorphism information used for determining the administration efficacy of an antihypertensive agent containing diuretic as an active ingredient. is there. That is, the antihypertensive agent containing a diuretic as an active ingredient is effective in lowering the blood pressure of a hypertensive patient having diabetes having at least one of the gene polymorphisms described in the table of FIG.

  More specifically, the table shown in FIG. 3 is obtained by statistically processing a patient's blood pressure value for a group of diabetic patients to whom a hypotensive agent containing a diuretic as an active ingredient is administered. . First, a cross-sectional survey was conducted on 3939 patients with consent from among diabetic patients who visited 8 institutions nationwide including Osaka University between January 2005 and May 2006. The survey items were the degree of diabetic complications, and environmental and genetic factors that could be related to diabetic complications. Then, using the data obtained by the cross-sectional survey, statistical analysis was performed on the administration status and gene polymorphisms of antihypertensive agents containing diuretics as active ingredients. More specifically, the systolic blood pressure (also referred to as SBP) and the diastolic blood pressure for a group of 151 diabetic hypertensive patients to whom a hypotensive agent containing a diuretic as an active ingredient was administered. (Diastolic Blood Pressure: also referred to as DBP) was evaluated statistically. As a result, as will be described later, it was found that patients having a specific genotype in a specific gene significantly decreased SBP and DBP compared to patients having a different genotype. From this, the table of FIG. 3 was obtained.

  Using the above data, patients are classified into two sets according to whether or not each gene polymorphism information in the table shown in FIG. 3 is included in the patient's genetic information, and p <0.05 ( The results of t-test at a significance level of 5% are shown in FIGS. 4 and 5 show the results regarding SBP and DBP, respectively.

  4 and 5, SNP NAME is the SNP name, n1 is the number of patients in the set having the SNP of the genotype GENOTYPE (1) (hereinafter referred to as the first set), and mean1 and SD1 are the first set, respectively. Mean value and standard deviation. n2 is the number of patients in the set (hereinafter referred to as the second set) that does not have the SNP of the genotype GENOTYPE (1) (that is, the SNP of the GENOTYPE (2)), and mean2 and SD2 are the second set, respectively. Mean value and standard deviation. Genotypes GENOTYPE (1) and GENOTYPE (2) are represented by the numbers defined above. T-test P value is the p value of the t test.

  In FIG. 4, since all t values of the t test are less than 0.05, it can be seen that there is a significant difference between the first and second sets. When comparing the average values of the SBPs of the first and second sets, the first set is in the range of 131.4 to 134.0, the second set is in the range of 137.7 to 151.0, The average value of one set is smaller than the average value of the second set. This means that the antihypertensive agent whose active ingredient is the administered diuretic worked effectively for the first set of patients. For the second group of patients, this means that the antihypertensive agent containing the administered diuretic as an active ingredient did not work so effectively.

  Regarding FIG. 5, the same thing as FIG. 4 can be seen. That is, in FIG. 5, since all t values of the t test are less than 0.05, it can be seen that there is a significant difference between the first and second sets. Then, comparing the average values of the DBPs of the first and second sets, the first set is in the range of 72.7 to 74.9, the second set is in the range of 77.6 to 87.2, The average value of one set is smaller than the average value of the second set. This means that the antihypertensive agent whose active ingredient is the administered diuretic worked effectively for the first set of patients. For the second group of patients, this means that the antihypertensive agent containing the administered diuretic as an active ingredient did not work so effectively.

  From the above, it can be said that an antihypertensive agent containing this diuretic as an active ingredient works effectively as an antihypertensive agent for a hypertensive patient having diabetes having the SNP of the genotype GENOTYPE (1) shown in FIG. . Conversely, the antihypertensive agent containing this diuretic as an active ingredient is not so effective as a hypotensive agent for hypertensive patients with diabetes who do not have the SNP of the genotype GENOTYPE (1) shown in FIG. I can say that.

  Here, the diuretics are thiazide diuretics (trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof), thiazide-like diuretics (indapamide, chlorthalidone, tripamide) as described later. , Methiclan, mefluside, and pharmaceutically acceptable salts thereof), loop diuretics (furosemide, and pharmaceutically acceptable salts thereof), or K-retaining diuretics (spironolactone, triamterene, and pharmaceutical agents thereof) Acceptable salt).

(4) Antihypertensive agent comprising diuretic as an active ingredient The antihypertensive agent provided by the present invention has at least one gene (gene polymorphism) of the following (1) to (4) having the following genotype ( genotype) is a drug that is effective for hypertensive patients and contains diuretics as active ingredients. The drug is used for hypertensive patients in which at least one of the following genes (1) to (4) is confirmed to have the following genotype, particularly hypertensive patients with diabetes It is characterized by being administered.

  The hypertensive patient to be treated by the antihypertensive agent of the present invention may be any person having at least one of the above genotypes among the above (1) to (4), but any two or more of these genes A person having the above genotype may be used.

  The diuretics as active ingredients of the antihypertensive agent of the present invention are roughly classified into (a) thiazide diuretics, (b) thiazide-like diuretics, (c) loop diuretics, and (d) K-retaining diuretics. It can be classified into four types of drugs.

The (a) thiazide diuretics and the (b) thiazide-like diuretics are diuretics that mainly act on the distal tubule, and exert a blood pressure lowering action in association with the excretion action of Na ⁺ . (a) As thiazide diuretics, hydrochlorothiazide (trade name “Dicrotride” (Manyu Pharmaceutical), etc.), trichlormethiazide (trade name “Fluitran” (Shiono Pharmaceutical), etc.), bench Examples thereof include benzylhydrochlorothiazide (trade name “Behide” (Kyorin Pharmaceutical Co., Ltd.)). (B) Thiazide-like diuretics include meticrane (trade name “Aresten” (Nippon Shinyaku), etc.), indapamide (trade name “Natricks” (Kyoto Yakuhin), “tenaxyl” ( Aripresa), tripamide (trade name “Normonal” (Eisai), etc.), chlortalidone (trade name “Hygroton” (Novartis Pharma), etc.), mefruside (trade name “Bikaron” (Mitsubishi) Welpharma) etc.].

  (c) The loop diuretic is a diuretic that acts on the henile ascending limb medulla. Unlike the thiazide diuretics, the loop diuretics have the advantage that they can be used even during kidney damage because they do not decrease renal blood flow and glomerular filtration rate. Examples of such loop diuretics include furosemide (trade names “Lashix”, “eutensin” (Aventis Pharma), etc.).

(d) K-retaining diuretics are diuretics that act primarily on distal tubules. Such K-retaining diuretics include spironolactone (trade name “Arducton A” (Pfizer), etc.) having an action of promoting Na ⁺ water excretion of distal tubules and suppressing K excretion by aldosterone receptor antagonistic action. And triamterene (trade name “Triterene” (Kyoto Yakuhin Co., Ltd.) etc.) that exerts K-retaining action by acting directly on the distal tubule and cortical tubule regardless of aldosterone.

  The antihypertensive agent of the present invention only needs to contain these diuretics in an effective amount that exerts an antihypertensive action, and as long as it contains other components such as pharmaceutically acceptable carriers and additives. You may do it. The antihypertensive agent of the present invention can be usually administered orally or parenterally (for example, intravenous administration), and in a form suitable for their administration [for example, tablets, pills, powders, solutions, suspensions] Agents, emulsions, granules, capsules, and the like; intravenous injections such as liquids, or injections].

  Examples of the pharmaceutically acceptable carrier that can be blended with the antihypertensive agent include a wide variety of carriers that are commonly used in the art depending on the various administration forms. For example, fillers, extenders, binders, wetting agents, disintegrants, surfactants, lubricants, buffers, chelating agents, pH adjusters, surfactants, isotonic agents, soothing agents, etc. Can be mentioned. Moreover, a stabilizer, a disinfectant, a coloring agent, a preservative, a fragrance, a flavoring agent, a sweetener, and the like can be blended with the antihypertensive agent of the present invention as necessary.

  The antihypertensive agent of the present invention can also be prepared as a sustained-release preparation or a sustained-release preparation containing a diuretic as an active ingredient according to a known method.

  The amount of diuretic to be contained in the antihypertensive agent of the present invention is not particularly limited and is appropriately selected from a wide range, but it is usually preferably about 1 to 70% by weight.

  The dosage and administration form of the antihypertensive agent of the present invention are not particularly limited, and for example, a method according to the type of diuretic compounded as an active ingredient, various preparation forms, patient age, sex and other conditions, and the degree of disease. Be administered. For example, hydrochlorothiazide is about 10 to 100 mg at a time, once or several times a day; trichloromethiazide is about 1 to 8 mg a day, divided into one or more times; methiclan is about 150 mg once a day Or multiple times; indapamide is divided into 1 to 2 mg daily or divided into multiple doses; trypamide is divided into 15 mg daily or divided into multiple doses; chlorthalidone is divided into 10 to 100 mg daily or once Divide (daily or every other day administration); Mefluside is divided into 25 to 50 mg per day in one or more; Fursemide is divided into 40 to 80 mg per day in one or more; Spironolactone is 50 to 100 mg per day The degree is divided into one or more; it is recommended that triamterene be administered at a dose of about 90 to 200 mg per day in one or more divided doses.

(5) Method for detecting administration efficacy of antihypertensive agent containing diuretic as active ingredient for hypertensive patient The present invention relates to a method for detecting administration efficacy of antihypertensive agent containing diuretic as an active ingredient for hypertensive patient I will provide a.

  The method can be carried out by detecting a genotype in at least one gene polymorphism of any one of the following (1) to (4) for a biological sample of a hypertensive patient.

  Of the genes shown in the above (1) to (4), (1) Glutamate-cystein ligase gene will be used as an example to explain the present invention in more detail. By detecting the genotype of position 588 [Glutamate-cystein ligase (C588T)], CC homozygote (genotype: 1), CT or TC heterozygote (genotype: 2), or TT homozygote This is a method for identifying different zygotes (genotype: 3), and based on the description in Table 3 above, when it is a CC homozygote (genotype: 1), it is a diuretic for the treatment of hypertensive patients. It can be determined that the administration of the drug is effective. Conversely, when Glutamate-cystein ligase (C588T) is not a CC homozygote (genotype: 1), it can be determined that administration of a diuretic is not effective for the treatment of the hypertensive patient. Similarly, when the genes of (2) to (4) are also targeted, the effectiveness of administration of diuretics for hypertensive patients based on the genotype described in Table 3 above (ineffectiveness of administration from the opposite viewpoint) Can be detected.

  The genes (gene polymorphisms) shown in Table 3 are gene markers that indicate the effectiveness of diuretics (sensitivity to diuretics) for hypertensive patients, particularly hypertensive patients with diabetes. The gene marker can be suitably used to determine whether administration of a diuretic is effective for hypertension treatment (hypertensive treatment) for hypertensive patients, particularly hypertensive patients with diabetes. If the gene (gene polymorphism) of the target patient matches the genotype shown in Table 3, the patient is responsive (sensitive) to the diuretic, that is, the diuretic is in the patient's It means to respond to antihypertensive treatment.

  Specifically, genotype detection is carried out by performing PCR in the region containing the genetic polymorphism of at least one gene described in (1) to (4) with respect to the genomic DNA of the subject, and using the SSCP method. (Ii) The method of detecting restriction enzymes on PCR products by performing PCR in the region containing the genetic polymorphism of at least one gene described in (1) to (4) for the subject genomic DNA (Iii) a method of directly sequencing the PCR product and determining the sequence; (iv) targeting at least one of the genes described in (1) to (4) using the genomic DNA of the subject. ASO (allele specific oligonucleotide) method using oligonucleotides that hybridize to the region containing the gene polymorphism as a probe and hybridizing with individual DNA, (v) (1) to (4) The genetic polymorphism of at least one gene described in) An oligonucleotide that hybridizes to a region comprising using as a probe, and a method of detecting in a mass spectrometer or the like, can be carried out by using a known method.

More specifically, the method of the present invention can be carried out by performing the following steps (a) and (b):
(A) a step of detecting a genotype in a genetic polymorphism of at least one gene described in (1) to (4) for a biological sample of a hypertensive patient;
(B) A step of discriminating whether the genotype in the genetic polymorphism of at least one gene described in (1) to (4) is the genotype described in Table 3 above.

  The step (a) can be performed specifically on genomic DNA extracted from a biological sample of a hypertensive patient.

  By these steps, when the genotype in the polymorphism of at least one gene described in (1) to (4) matches the genotype described in Table 3 above, hypertension provided with the genomic DNA sample It can be determined that administration of diuretics is effective for treating hypertension in patients. Conversely, if they do not match the genotypes listed in Table 3, diuretics are not effective for this treatment and can give the option of considering administration of other antihypertensive agents.

Such a determination step is not an essential step of the method of the present invention, but such a determination step can optionally be included as the following step (c):
(c) Based on the result of (b) above, when the genotype in the genetic polymorphism of at least one gene described in (1) to (4) matches the genotype described in Table 3, Determining that the administration of diuretics is effective for the treatment of hypertension, and conversely, if the genotypes listed in Table 3 do not match, determining that the administration of diuretics is not effective for the treatment of hypertensive patients .

  That is, according to the method of the present invention, it is possible to determine whether or not the diuretic is effective for administration to hypertensive patients. These determinations can be made even by a specialist prior to administration by determining whether the polymorphisms of the above-mentioned (1) to (4) genes in the hypertensive patient are the genotypes shown in Table 3 or not. Extremely difficult decisions can be made automatically / mechanically.

  The above steps (a) and (b) are known methods [eg Bruce, et al., Geneme Analysis / A laboratory Manual (vol. 4), Cold Spring Harbor Laboratory, NY., (1999)]. Can be used.

Specific examples of the biological sample to be processed in step (a) include genomic DNA as described above. Such genomic DNA can be any cell (including cultured cells, excluding germ cells), tissue (including cultured tissue), or body fluid (eg, blood, saliva, lymph, airway mucosa, semen, isolated from hypertensive patients. , Sweat, urine, etc.) can be prepared as a material. Preferably it is blood or urine. 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.

  In step (b), the base located at the gene polymorphic site of at least one gene of (1) to (4) is identified from the extract containing human genomic DNA obtained as described above. As a method for identifying the target base, in addition to the method of directly determining the gene sequence of the corresponding region, when the polymorphic sequence is a restriction enzyme recognition site, the genotype is utilized by utilizing the difference in the restriction enzyme cleavage pattern. (Hereinafter referred to as RFLP), a method based on hybridization using a polymorphic specific probe (for example, a specific probe is pasted on a chip, glass slide, nylon membrane, and hybridization to those probes) By detecting the difference in intensity, the type of polymorphism is determined, or the efficiency of specific probe hybridization is determined by detecting the amount of probe that polymerase degrades during template double-strand amplification. , The fluorescence emitted by a certain type of double-strand-specific fluorescent dye can be detected by following the temperature change. Detecting the temperature difference in strand melting and identifying the polymorphism by this method, attaching complementary sequences to both ends of the polymorphic site-specific oligo probe, and forming the secondary structure within the self molecule according to the temperature Or a method for identifying a genotype using a difference between hybridization to a target region and the like. 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) TaqMan-PCR method [Genet. Anal., 14, 143-149 (1999); J. Clin. Micro biol., 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 (Matrix Assisted Laser Desorption-time of Flight / Mass Spectrometry) method [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・ Padlock Probe method [Nat. Genet., 3, p225-232 (1998); Gene medicine, 4, p50-51 (2000)], (q) UCAN method [see Takara Shuzo Co., Ltd. home page (http://www.takara.co.jp)], (r) DNA chip or DNA microarray ("SNP gene polymorphism strategy" Kenichi Matsubara, Yoshiyuki Tsuji, Nakayama Bookstore, p128-135), (s) ECA method [Anal. Chem., 72, 1334-1341, (2000)].

  The above are typical gene polymorphism detection methods, but the method of the present invention is not limited to these, and other known or future developed gene polymorphism detection methods can be widely used. Moreover, when detecting the gene polymorphism of the present invention, these gene polymorphism detection methods may be used singly or in combination of two or more.

  In addition, when a mutation occurs in the amino acid to be encoded based on the difference between the gene polymorphisms (1) to (4), the method of the present invention is not limited to the method of directly detecting and identifying the genotype of the gene, A method of detecting the amino acid sequence of a protein that is an expression product of the gene and identifying the presence or absence of an amino acid mutation can also be employed.

  As described below, for example, probes and primers for detecting and distinguishing gene polymorphisms of the above-mentioned (1) to (4) genes, for each hypertensive patient, whether the patient is a diuretic administration effective person. It can be used as a tool for selecting whether or not.

(6) For hypertensive patients, a tool for detecting the effectiveness of administration of an antihypertensive agent containing a diuretic as an active ingredient (6-1) Probe, array having the probe Including the target gene polymorphism and the base For detection of oligo or polynucleotide, the gene polymorphism [(1) Glutamate-cystein ligase (C588T), (2) in the base sequence of the gene (1) to (4) described in Table 3 above is used. Specific for oligos or polynucleotides containing CD18 (C1323T), (3) Cytochrome P450 2J2 * 7 (G-50T), or (4) Alcohol dehydrogenase 2 (Arg47His (G / A))), or their complements Oligos or polynucleotides that can hybridize to and identify the base of the polymorphic site of the gene are used.

  Such an oligo or polynucleotide has a length of 16 to 500 bases, preferably 20 to 200 bases, more preferably 20 to 50 bases including the gene polymorphism in the base sequence of the genes (1) to (4). Oligo having the above base length (16 to 500 base length, preferably 20 to 200 base length, more preferably 20 to 50 base length) so as to specifically hybridize with a continuous gene region (or its complementary region) Or it is designed as a polynucleotide.

  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 preferably has a base sequence complementary to the base sequence of the gene region containing the gene polymorphism to be detected, but if such specific hybridization is possible, It need not be completely complementary.

  The oligo or polynucleotide is used as a “probe” for detecting the gene polymorphism of the genes (1) to (4) and determining the effectiveness of the diuretic for hypertensive patients, in other words, for diuretics in hypertensive patients. Designed as a “probe” for detecting dosing effectiveness. In addition, these oligos or polynucleotides can be prepared according to a conventional method by a commercially available nucleotide synthesizer, for example, based on the base sequence of the region containing the gene polymorphisms of (1) to (4) genes.

  More preferably, the probe is labeled with a radioactive substance, a fluorescent substance, a chemiluminescent substance, an enzyme, or the like so that an oligo or polynucleotide containing a gene polymorphism of (1) to (4) gene can be detected ( Later).

  In addition, the probe (oligo or polynucleotide) can be used by being immobilized on an arbitrary solid phase, and the probe targeted by the present invention includes the immobilized probe (for example, a gene chip on which the probe is immobilized, a cDNA microarray, Oligo DNA arrays, membrane filters, etc.) are also included. The probe can be preferably used as a DNA chip or DNA array for detecting the effectiveness of diuretic administration (or for determining the sensitivity of diuretics).

  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). For example, in the case of 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, in the region containing the gene polymorphism. A complementary oligonucleotide having a length of about 20 bases 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, 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.

(6-2) Primer The present invention also relates to the gene polymorphism of the gene [(1) Glutamate-cystein ligase (C588T), (2) Used as a primer to specifically amplify a sequence region containing CD18 (C1323T), (3) Cytochrome P450 2J2 * 7 (G-50T), or (4) Alcohol dehydrogenase 2 (Arg47His (G / A))] Provided oligonucleotides. Such a primer (oligonucleotide) specifically hybridizes to a part of a continuous oligo or polynucleotide containing nucleotides of each gene polymorphism site in the base sequence of the genes (1) to (4), It 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 according to the detection method used, but is generally 15 to 1000 bases, preferably 20 to 500 bases, more preferably 20 to 200 bases It is.

  In addition, these oligonucleotides can be prepared in accordance with a conventional method using, for example, a commercially available nucleotide synthesizer, based on the base sequence including the gene polymorphisms of the genes (1) to (4).

(6-3) Labeled substance The probe or primer of the present invention is added with an appropriate labeling substance for detecting a gene polymorphism such as a fluorescent dye, an enzyme, a protein, a radioisotope, a chemiluminescent substance, biotin and the like. Is 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 polymorphism 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.

  The probe described above (which may be labeled or immobilized) or primer (which may be labeled) is used to determine whether diuretics are effective in treating hypertensive patients. In other words, it can be used as a reagent for detecting (selecting) the effective diuretic administration person and the ineffective administration person for hypertensive patients.

(6-4) Reagent Kit for Detecting Effectiveness of Administration of Diuretics The present invention also detects the administration effectiveness of antihypertensive agents containing diuretics as active ingredients for hypertensive patients, particularly hypertensive patients with diabetes. It provides as a reagent kit for doing.

  Specifically, the reagent kit of the present invention comprises a primer pair capable of specifically amplifying a region having a gene polymorphism of at least one gene of (1) to (4) described in Table 3, and / or It contains a nucleic acid probe that specifically hybridizes to a gene or its amplification product. These may be labeled or immobilized on a solid phase.

  In addition to the probe or primer described above, the kit of the present invention includes other reagents and instruments necessary for carrying out the method of the present invention, such as a hybridization reagent, a probe label, a label detection agent, and a buffer as necessary. Etc. may be included as appropriate. Furthermore, the reagent kit of the present invention may include a document explaining a method for using a reagent such as a primer or a probe, and a method (determination criteria) for determining the administration effectiveness of a diuretic using the reagent kit.

  Hereinafter, the present invention will be described more specifically by way of examples of the present invention. A gene of a hypertensive patient having diabetes was analyzed, a patient having any of the gene (SNP) and genotype shown in FIG. 3 was selected, and an antihypertensive agent containing a diuretic as an active ingredient was administered. Further, patients having the gene (SNP) shown in FIG. 3 but having a genotype different from that shown in FIG. 3 were selected and administered with the same agent.

  6 and 7 show the correspondence between gene polymorphisms and SBP and DBP. The meaning and notation of SNP NAME and GENOTYPE are the same as above. The values shown in DATA1 to DATA3 are measured values of SBP or DBP of each patient having a gene polymorphism represented by SNP NAME and GENOTYPE. AVERAGE is an average value of DATA1 to DATA3 of each row. The “-” cell means that the corresponding data does not exist, and the AVERAGE of the row is an average value of DATA1 and DATA2. In addition, two rows of data are shown for each SNP NAME, the upper row shows data relating to a patient having the genotype shown in FIG. 3, and the lower row shows genes different from the genotype shown in FIG. Data for patients with type are shown.

  As can be seen from FIG. 6, the measured value of SBP is relatively low for patients having either the gene (SNP) or genotype shown in FIG. On the other hand, for patients having the gene (SNP) shown in FIG. 3, but whose genotype is different from that shown in FIG. 3, the measured value of SBP is relatively high. Similarly, as can be seen from FIG. 7, the measured value of DBP is relatively low for the patient having either the gene (SNP) or genotype shown in FIG. 3, and has the gene (SNP) shown in FIG. However, for patients whose genotype is different from that shown in FIG. 3, the measured DBP is relatively high. This means that for patients having either the gene (SNP) or genotype shown in FIG. 3, administration of a blood pressure lowering agent containing a diuretic as an active ingredient is effective, whereas FIG. 3 shows For patients who have a gene (SNP), but whose genotype is different from that shown in FIG. Thus, depending on whether or not a hypertensive patient with diabetes has any of the gene (SNP) and genotype shown in FIG. 3, the administration effect of an antihypertensive agent containing a diuretic as an active ingredient is apparent. There was a difference.

Representative literature disclosing genetic information on (1) Glutamate-cystein ligase gene, (2) CD18 gene, (3) Cytochrome P450 2J2 * 7 gene, and (4) Alcohol dehydrogenase 2 gene, and positional information on gene polymorphism Indicates. It is a flowchart which shows the determination method of the administration effectiveness of the antihypertensive agent which uses the diuretic which concerns on embodiment of this invention as an active ingredient. It is a table | surface which shows the gene polymorphism information utilized for judgment of the administration effectiveness of the blood pressure lowering agent which uses a diuretic as an active ingredient. With regard to systolic blood pressure (SBP) measured by administering antihypertensive agents containing diuretics to hypertensive patients with diabetes, patients are classified into two groups according to the presence or absence of genetic polymorphism, and t-test It is a table | surface which shows the result of having performed. With regard to diastolic blood pressure (DBP) measured by administering antihypertensive agents containing diuretics to hypertensive patients with diabetes, patients are classified into two sets according to the presence or absence of genetic polymorphism, and t-test It is a table | surface which shows the result of having performed. It is a figure which shows the Example of this invention, and has shown the response | compatibility with a gene polymorphism and SBP about the hypertensive patient who has diabetes administered the antihypertensive agent which uses a diuretic as an active ingredient. It is a figure which shows the Example of this invention, and has shown the response | compatibility with a gene polymorphism and DBP about the hypertensive patient who has diabetes administered the antihypertensive agent which uses a diuretic as an active ingredient.

Claims (9)

Effective diuretics characterized by being administered to hypertensive patients with diabetes mellitus confirmed to have the following genotype for at least one gene of any one of (1) to (4) Antihypertensive agent as ingredient:
Gene (gene polymorphism) : genotype
(1) Glutamate-cystein ligase (C588T) ： 1
(2) CD18 (C1323T): 23
(3) Cytochrome P450 2J2 * 7 (G-50T): 1
(4) Alcohol dehydrogenase 2 (Arg47His (G / A)): 1   The diuretic is a thiazide diuretic consisting of trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof; A thiazide-like diuretic consisting of a salt; a loop diuretic consisting of furosemide and its pharmaceutically acceptable salt; and a K-retaining diuretic consisting of spironolactone, triamterene and a pharmaceutically acceptable salt thereof The antihypertensive agent according to claim 1, which is at least one drug selected from the group consisting of: A method for detecting the administration effectiveness of an antihypertensive agent comprising a diuretic as an active ingredient for a hypertensive patient having diabetes, wherein the hypertensive patient has the following gene for at least one of the genes (1) to (4): A method that uses a genotype as an indicator:
Gene (gene polymorphism) : genotype
(1) Glutamate-cystein ligase (C588T) ： 1
(2) CD18 (C1323T): 23
(3) Cytochrome P450 2J2 * 7 (G-50T): 1
(4) Alcohol dehydrogenase 2 (Arg47His (G / A)): 1   The diuretic is a thiazide diuretic consisting of trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof; A thiazide-like diuretic consisting of a salt; a loop diuretic consisting of furosemide and its pharmaceutically acceptable salt; and a K-retaining diuretic consisting of spironolactone, triamterene and a pharmaceutically acceptable salt thereof 4. The method of claim 3, wherein the method is at least one drug selected from the group consisting of:   (1) Glutamate-cystein ligase (C588T), (2) CD18 (C1323T), (3) Cytochrome P450 2J2 * 7 (G-50T), and (4) Alcohol dehydrogenase 2 (Arg47His (G / A)) An array for detecting the administration efficacy of a hypotensive agent comprising a diuretic as an active ingredient for a hypertensive patient having diabetes, comprising a probe for detecting a genetic polymorphism of at least one gene selected from the group.   The diuretic is a thiazide diuretic consisting of trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof; A thiazide-like diuretic consisting of a salt; a loop diuretic consisting of furosemide and its pharmaceutically acceptable salt; and a K-retaining diuretic consisting of spironolactone, triamterene and a pharmaceutically acceptable salt thereof 6. The array of claim 5, wherein the array is at least one drug selected from the group consisting of:   (1) Glutamate-cystein ligase (C588T), (2) CD18 (C1323T), (3) Cytochrome P450 2J2 * 7 (G-50T), and (4) Alcohol dehydrogenase 2 (Arg47His (G / A)) Having a diabetes comprising a primer pair capable of specifically amplifying a region having a gene polymorphism of at least one gene selected from the group, and / or a nucleic acid probe specifically hybridizing to the gene or its amplification product A reagent kit for detecting the administration effectiveness of an antihypertensive agent containing a diuretic as an active ingredient for hypertensive patients.   The reagent kit according to claim 7, comprising the array according to claim 5.   The diuretic is a thiazide diuretic consisting of trichlormethiazide, hydrochlorothiazide, benchylhydrochlorothiazide, and pharmaceutically acceptable salts thereof; A thiazide-like diuretic consisting of a salt; a loop diuretic consisting of furosemide and its pharmaceutically acceptable salt; and a K-retaining diuretic consisting of spironolactone, triamterene and a pharmaceutically acceptable salt thereof The reagent kit according to claim 7 or 8, which is at least one drug selected from the group consisting of:

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