TWI392741B - Genetic polymorphism for determining ace activity and blood pressure response to ace inhibitor - Google Patents

Description

Gene polymorphism affecting angiotensin-converting enzyme activity and response to angiotensin-converting enzyme activity inhibitor drug and method thereof

The present invention relates to a gene polymorphism and its use, and more particularly to a gene polymorphism and method for influencing angiotensin-converting enzyme activity and predictive response to angiotensin-converting enzyme activity inhibitor drug response.

In 2007, after the Wellcome Trust Case Control Consortium (WTCCC) published seven complex diseases using the genome-wide association studies (GWAS), The detection method is getting more and more attention. Although GWAS finds a variety of novel loci that may be associated with common diseases in a wide range of samples, most studies have shown that these loci have only mild to moderate associations with these diseases. This phenomenon is most likely due to the heterogeneous nature of most complex diseases. Angiotensin-converting enzyme activity In the upper reaches of hypertension and between disease genes and hypertension, angiotensin-converting enzyme activity is not only highly heritable (0.2-0.7) but also the variability of cases measured at different time points. Very small, so angiotensin-converting enzyme activity is a good mediator phenotype, suitable for gene mapping to indirectly look for genetic variants that affect blood pressure regulation, cardiovascular function, and other potentially relevant biological mechanisms.

Angiotensin converting enzyme (ACE) is a very important enzyme in the renin-angiotensin-aldosterone system and has its electrolyte balance and blood pressure regulation. Importance, the first type of angiotensin I can be converted into a second type of angiotensin II. Type II angiotensin is a potent angiotensin and breaks down bradykinin, a potent vasodilator. At the same time, the second type of angiotensin-converting enzyme acts in other physiological processes through direct pathological effects on the tissue, including vascular remodeling and inflammatory effects, or indirectly through the bioavailability of nitric oxide and its results. Have an impact. Further, the genetic variation of angiotensin-converting enzyme, in particular its 16th intron Alu repeat insertion/deletion diversity (I/D), and the risk of many clinical outcomes Related, including cardiovascular disease, angiotensin-converting enzyme inhibitor (ACE inhibitor referred to herein as ACEI) therapy response, diabetic nephropathy, muscle performance, longevity, and Alzheimer's disease, however, the results are still contradictory . However, other studies have shown that the I/D polymorphism of the D-pair gene increases angiotensin-converting enzyme activity. Therefore, an effective method should be to divide complex diseases into more homogenous endogenous or phenotypes, which have a higher heritability than the original phenotype. This approach, by focusing on a smaller range of diseases, can help identify genes associated with subgroups of previously complex diseases and can be helpful in understanding the cause of the disease.

However, although angiotensin-converting enzyme (ACE) is widely associated with many biological systems, the pharmacological mechanisms involved in the regulation of ACE activity and angiotensin-converting enzyme inhibitor (ACEI) are not clear and therefore cannot be It is used to evaluate angiotensin-converting enzyme-related abnormalities, diseases, or blood pressure response when taking hypertensive drugs.

Accordingly, it is an object of the present invention to provide a method for identifying a high risk that humans develop an angiotensin-converting enzyme-associated disorder (ACE-linked disorder).

Another object of the present invention is to provide a method of predicting a patient's blood pressure response to angiotensin-converting enzyme inhibitor therapy (ACE inhibitor therapy).

The technical means adopted by the present invention to solve the problems of the prior art is to use a whole genome-related research method. The first stage is to use the Illumina Infinium II HumanHap 550 SNP chips whole genome scanning platform for 400 young hypertension cases. Correlation analysis of genome scanning. In the first stage, there are 8 SNPs on the ACE gene associated with angiotensin-converting enzyme activity. These 8 SNPs are located on the ACE gene, from the promoter to the exon24 position, -log The p value ranged from 11.18 to 20.06, with the highest -log p value for SNP rs4343 located on exon 17, near the position of the ACE gene I/D polytype (Intron 16). In addition to the ACE gene, another SNP with a -log p of more than 7 is rs4955828 (-log p value = 7.59) located on the ABO blood group gene promoter.

According to the results of the whole genome correlation study above, the SNP with a -log p value of more than 7 entered the second phase of the confirmed study, because there is a strong linkage disequilibrium (LD) phenomenon between SNPs in the ACE gene, so Only one of the most significant SNPs (rs4343) was selected in the ACE gene. In addition to SNP rs495828 (SNPs affecting type A blood group expression) on the ABO gene, SNP rs8176746 (SNP affecting type B blood group expression; -log p value = 5.59) The ABO gene exon 7 was also selected for the second phase of the study. The second phase confirmed the results of the first phase with an additional 623 hypertension study cases, which were still associated with angiotensin-converting enzyme activity in the second phase of the study. The p value of SNP rs4343 is 3.02x10 ^-25 , the p value of SNP rs495828 is 3.54x10 ^-8 , and the p value of SNP rs8176746 is 9.30x10 ^-5 .

Then, in the independent young family study, hypertension and normal blood pressure were used to verify the correlation between ABO genotype/blood type and ACE activity, and the genetic scores of ABO gene and SNP on ACE gene. It is related to the systolic blood pressure response after taking an angiotensin-converting enzyme inhibitor. In addition, in the follow-up study of 3,350 people in Zhudong and Puzi, the genetic score was also related to the risk of stroke. As the genetic score increased, the risk ratio of stroke occurred. (Risk ratio = 1.47, p = 0.0086), the incidence of stroke also increases as the genetic score increases.

It was confirmed by the technical means employed in the present invention that angiotensin-converting enzyme activity and ACE gene are associated with the ABO gene or blood type in 1023 cases of hypertension and 282 normal blood pressure persons, respectively. The present invention is the first to demonstrate that the ABO genotype plays an important role in the regulation of angiotensin-converting enzyme activity. These results may help us explain the past discussion of angiotensin-converting enzyme activity or ACE I/D polymorphism and blood pressure regulation. And the relationship between cardiovascular disease and research results. At the same time, we found that people with different ACE activity genetic scores have different responses to ACEI drugs. If this information is further confirmed by clinical trials, it can be applied to fine-tune the use of hypertension drugs or heart disease prevention strategies. For example, consider selective An angiotensin-converting enzyme inhibitor drug is administered to an individual having a high angiotensin converting enzyme activity genotype. Through the disclosure of the present invention, it is shown how to use the quantitative trait locus discovered by the whole genome correlation study to further apply to pharmacogenomics, which can be used in the future to correct the physician's habit of prescribing drugs. At the same time, the use of increased genetic scores of ACE activity may increase the risk of stroke, and the future can be applied medically to assess the risk aspects of stroke.

The above description will be more clearly illustrated by the following examples and the accompanying drawings, which are to be construed as being limited by the scope of the invention.

definition

The terminology used in the specification refers to the general meaning in the field. For the purposes of the following discussion in this specification, some terms are indicated in a special font format for convenience, for example, in italics and/or parentheses. The use of these font formats does not affect the scope and meaning of the term itself. Whether or not marked in a special font format, the scope and meaning of the term itself are the same. Therefore, the use of any equivalent language or synonym is not intended to change its meaning. The use of one or more synonyms does not exclude the use of other synonyms. The use of any terms in the embodiments of the present invention is merely illustrative and is not intended to limit the scope and meaning. Likewise, the scope of the invention is not limited only by the embodiments that are present.

Unless specifically defined, all technical and scientific terms appearing herein have the usual meaning as recognized by those of ordinary skill in the art.

The terms "about" and "substantially" as used in the present invention mean within 20%, preferably within 10%, and more preferably within 5%. The numbers provided herein are approximate and, if not explicitly indicated, are intended to have an approximate or approximate meaning.

Example

The apparatus, the device, the method, the related results, and the like according to the embodiments of the present invention described below are for illustrative purposes only and are not intended to limit the scope of the present invention. The names in the examples or their sub-names are for convenience of reading and are not intended to limit the scope of the invention. Further, the theory disclosed herein, whether or not it is inaccurate, is not intended to limit the scope of the invention.

The present invention utilizes the Whole Genome Related Research (GWAS) method to perform a whole gene for 400 young-onset hypertension (YOH) cases using 560,168 SNP genotypes derived from Illumina Infinium II HumanHap 550 SNP wafers. Correlation analysis of body scans, identification of quantitative trait loci (QTLs) associated with ACE activity, and evaluation of the efficacy of genome-wide correlation studies (GWAS) for mapping genetic maps of high genetic traits. Obviously, in addition to the ACE gene, the ABO gene is also significantly associated with ACE activity. The association of ABO blood group with ACE activity was also tested here, and comparing the different number of ACE activities with/without ACE inhibitor (ACEI) increased the blood pressure trend of the dual gene (as with the substitute for drug response). The genetic information obtained on ACE activity can be applied to the clinical classification and treatment of ACE-related diseases after further repeated experiments.

method

Quantitative ACE activity traits using the Whole Genome Related Study (GWAS)

Two-stage GWAS was used to study the association between genes/locus affecting ACE activity. Genetic studies were conducted in a total of 1023 cases of young hypertension (AS-YOH). Includes 400 individuals in the first phase and 623 individuals in the second phase. The criteria for selection of these hypertensive individuals include the following six points: (1) systolic blood pressure (SBP) > 140 mmHg and / or diastolic blood pressure (DBP) > 90 mmHg more than two months, or taking antihypertensive drugs, Two consecutive measurements of SBP/DBP>120/80 mmHg in two months, (2) ages between 20 and 51 years, the first diagnosis of hypertension, (3) no second factor caused high Blood pressure, including, for example, chronic kidney disease, renal artery stenosis, primary aldosteronism, aortic stenosis, thyroid abnormalities, Cushing's disease, and pheochromocytoma, and confirmed by extended clinical tests, including, for example, blood Biochemical tests, renal function tests, endocrine tests, and abdominal ultrasound, (4) fasting blood glucose values below 126 mg/dl, (5) body mass index (BMI) below 35 kg/m ² , and (6) self-reported The Han Chinese ethnic group has more than two generations. Reference can be made to the reference: "A genome-wide association study identifies new loci for ACE 15 activity: potential implications for response to ACE inhibitor" Pharmacogenomics J. 2010 Jan 12, to make the invention more complete.

The collection of data is done in accordance with standard procedures. The blood pressure and pulse measurements are based on the standard procedures established by the Taiwan Nutritional Health Change Survey. Serum ACE activity (IU/L, nmole/min/mL) was measured from heparin-treated plasma taken from fasting blood samples (10-12 hours unfed) using spectrometry (ACEcolor, Fujirebio Inc) . The CV intra assay results were 4.0% and were calculated from replicate samples. In addition, the information is based on socio-demographic factors, smoking and eating habits, past medication history, and current drug use profiles, using interview interviews. This multi-center study was approved by the Academia Sinica Medical Research Ethics Committee. At the initial clinical meeting, the consent of all subjects was obtained.

Independent young hypertension family study to reproduce the relationship between ABO blood group and ACE activity

The data were collected from 464 hypertensive individuals with a genetic study of young hypertension. This family study has been approved by the Medical Research Ethics Committee of the Institute of Biomedical Sciences of the Academia Sinica. And all subjects have been approved by the subject.

The hyperbone source (proband) in the study came from four regional hospitals: the Health Department Lizhudong Hospital, the Health Department Li Taoyuan Hospital, Fuxin Hospital and Minsheng Hospital. The criteria for admission to hypertensive patients were essentially the same as for younger hypertensive families except for the age at which the first diagnosis of hypertension was less than 40 years old. Most individuals in this study provided self-reported blood type information. For those who did not provide information, the blood type was measured by a blood type test kit (Gamma Biologicals, Houston, Tx, USA). According to the previous literature (Blumenfeld et al), SNP rs498528 is a good substitute for the A antigen dual gene, while SNP rs8176746 is a good substitute for the B antigen dual gene. The two SNPs are used herein to make the self-described blood type effective. The A and B antigen dual gene lines showed high consistency with A blood type and B blood type (Chisq p-value was 6.85×10 ^-113 and 3.66×10 ^{-57 , respectively} ). Methods for measuring serum ACE activity are as described in the previous literature (Kasahara Y et al).

Correlation between ACE activity and SNP in patients with normal blood pressure

In order to investigate the association between ACE activity and SNPs of ACE and ABO genes in patients with normal blood pressure, a normal blood pressure was selected from the YOH family. The experiment was repeated with a total of 282 normal blood pressure.

Genotype analysis

For GWAS, 400 white blood cell DNA samples were genotyped by deCODE genetics (Reykjavik, Iceland) using Illumina Infinium II HumanHap550 SNP chips (Illumina, San Diego, CA, USA) and the WTCCC standard was used to control its quality, only These individuals are excluded when more than 3% of the required data is lost. When the Hardy Weinberg equilibrium (p<1x10 ^-7 ) is violated, the SNPs have a call rate of <95%, and a few minor allele frequencies <1% exclude the SNPs. The S-value of the two SNPs is greater than 7, and one SNP is a substitute for the B antigen-paired gene, and a first-stage non-synonymous polymorphisms of the exon 7 of the ABO gene ( -log p=5.59), genotype analysis was performed for the second-stage validation study and the reproducibility study using the Sequenom platform (Sequenom MassARRAY System) of the Academia Sinica Genotype Identification Center.

See Figure 1 for a brief step flow. The steps include: Step 101: Proliferating a specific SNP sequence from the genomic DNA with the first primer (forward) and the second primer (reverse), respectively. Step 102: Mixing with the SNP sequence using a primer (UEP_SEQ) to perform a primer extension. Step 103: Detection of the SNP dual genotype (EXT1_SEQ and EXT2_SEQ) by genotyping instrument.

Statistical Analysis

The ACE activity distribution is corrected by performing a square root operation on the original values. Here, the general linear model (GLM) is used to establish the correlation between ACE activity and genetic identification data, and the sex (0, 1), age and ACEI in the first, second and third stages and in the combined analysis ( 0, 1) Perform calibration. For the multiple testing corrections in the first stage, the genome-wide significance level is set to be less than 1x10 ^-7 .

In order to estimate the degree of genetic correlation explained by the selected SNPs, a stepwise linear multiple regression model (Multiple Regression Model) was performed on the data combined in the first and second phases. Add a SNP to the model only when the p-value is less than 0.1. To determine the number of dual genes or genotypes for each individual's ACE activity, by binding from SNP rs4343 (1 point for each G dual gene), SNP rs495828 (1 point for CC genotype), and SNP rs8176746 (CA or The AA genotype was given 1 point) genotype data (Table 4). GLM with medication status (1,0)* genetic score was used to compare blood pressure trends by increasing the number of dual genes/genotypes by ACE activity between those who took ACEI alone and those who did not take any antihypertensive drugs ( The blood pressure value is the dependent variable).

To repeat the association between ACE activity and ABO blood type, a regression analysis of the relationship between ACE activity and ABO phenotype was performed using the independent hypertension family study. Here, the blood type is set to the O type as a control group. In the regression model, age, gender (0, 1), ACEI (0, 1), etc. were taken as covariates, and the family ID was also corrected by the Generalized estimating equation (GEE). The same covariate was also used in the ACE activity-SNP association study of normal blood pressure individuals, since each family included only one body and the family ID was not corrected.

result

Research sample characteristics

There were no significant differences between the individuals in the first and second phases, including gender distribution, mean ACE activity, systolic blood pressure (SBP), and ACEI drug use ratio (see Table 1). In particular, there was a significant difference in mean age, body mass index (BMI), and diastolic blood pressure (DBP) values, with very small differences in BMI and DBP values.

Data are expressed as mean ± standard deviation

Abbreviation: NS (not significant)

^a t-test is used to compare the initial and second stage averages (significant level is set to p=0.05)

GWAS results

In order to reduce the age, gender, and additional confluing effects of taking ACEI, the factors in the linear regression analysis of quantitative trait loci (QTLs) affecting ACE activity are retrieved. The results are shown in Figure 2. As expected, the 8 SNPs with the highest correlation among the ACE gene (located between the promoter and exon 24) were obtained, and the -log p value ranged from 11.18 to 20.06. 2 picture). It is important to note that the SNP rs4343 located in exon 17 has the highest -log p value. It was found here that SNP rs495828 located in the ABO gene also has a significant correlation with ACE activity (-log p>7; Figure 2).

Based on the above GWAS results, a repetitive experiment was performed here using a SNP of -log p>7. Here, the most significant SNPs in the same linkage disequilibrium domain (LD block) (SNP rs4343 of the ACE gene and SNP rs495828 of the ABO gene) or SNPs located in the coding region (SNP rs8176746 of the ABO gene) were selected. Using an additional 623 hypertensive patients from the AS-YOH family, a total of 3 candidate SNPs were selected for the second phase for confirmation studies. These three SNPs were confirmed to be significantly associated with ACE activity, and the results are shown in Table 2: SNP rs4343 of ACE gene (p value = 3.02 x 10 ^-25 ), SNP rs495828 of ABO gene promoter (p value = 3.54) x 10 ^-8 ) and SNP rs8176746 of exon 7 of the ABO gene (p value = 9.30 x 10 ^-5 ). After excluding individuals taking ACEI, these three SNPs still had the same degree of significant association with ACE activity (data not shown). Furthermore, these SNPs and ACE activities have the same degree of significant correlation in small-scale replicate experiments (282 normal blood pressure individuals selected from the YOH family study).

^a dominant dual gene / recessive dual gene.

^{b The} percentage of recessive dual gene frequencies.

^c . Statistic of generalized linear pattern (GLM) correlation between genotype and ACE activity after adjusting for age, gender (0,1) and taking ACEI (0,1) in GWAS including 400 individuals result.

^d In the second phase of the validation study, the statistical results of GLM detection association between genotype and ACE activity after adjusting for age, gender (0, 1), and taking ACEI (0, 1).

^e Statistical significance of GLM-based associations between genotype and ACE activity in age- and gender-relevant studies in a third-stage validation study involving 282 subjects with normal blood pressure.

^f Obtain R ² by combining forward stepwise regression with the first and second stage samples.

By ACE Gene and ABO Changes in ACE activity as interpreted by genes

In the first and second phase analyses combined with 1023 hypertensive patients, approximately 23.9% of the changes in ACE activity were attributed to the following three SNPs (rs4343, rs485828, and rs8176746). The SNP rs4343 in the ACE gene represents a total change of 16.2%, whereas the SNP rs495828 and SNP rs8176746 in the ABO gene calculated an additional 4.9% and 2.8% of the total change, respectively (see Table 2).

Confirmation of the association between ABO blood group and ACE activity by independent samples

Because of the strong correlation between ABO genotype and ACE activity, it is related to whether the ABO blood group and ACE activity are related to individuals in the YOH family study. The results are shown in Table 3, showing that individuals with type A blood group have the lowest ACE activity, while individuals with type B blood type have the highest ACE activity. Individuals with type B antigen had higher mean ACE activity compared to individuals with type O blood group (p = 2.3 x ^{10 -10} ), while individuals with type A blood group had significantly lower mean ACE activity (p = 1.5). X10 ^-8 ). The AB blood type individuals were between Type A and Type B individuals, but were significantly different from Type O blood group individuals (Table 3).

^{The a} P value is obtained from the generalized estimation function (GEE) and corrected by EGE for age, gender (0, 1) and family ID.

ACE activity and ACE and ABO Dose-response relationship between genotypes

Referring to Table 4, the average ACE activity of individuals with ACE- rs4343 as the G-pair gene was increased in units of 3.5 IU per pair of even gene repeats. ABO- rs8176746 showed the same additional effect, which increased in units of 1.4-2.3 IU per A pair of gene repeats. In the individual, ABO- rs495828 is a CC genotype with an average ACE activity higher than that of a CA or AA genotype of about 3 IU per pair of genes.

Increased differential blood pressure trends in dual genes/genotypes by ACE activity in individuals taking ACEI alone and in untreated individuals

Referring to Figure 3, the difference in mean systolic/diastolic (SBP/DBP) differences between the number of ACE activities and the number of dual genes was observed between individuals taking antihypertensive drugs and those not taking them, using generalized linear mode (GLM) for comparison. The state of administration (0, 1) and the increase in ACE activity between the dual genes (0-4) (the one taking more than one type of hypertension drug will be excluded).

The mean amount of systolic blood pressure (SBP) and diastolic blood pressure (DBP) (p = 0.8875 and 0.9511, respectively) was not significant between unaffected ACE activity-increased dual-gene between individuals taking ACEI and those who did not. There is a difference in the ground. The average ACE activity of individuals taking ACEI alone was significantly lower than that of individuals who did not take anti-hypertensive drugs (p value = 1.98 x 10 ^-9 ). The ACE activity of each individual is increased by a dual gene or genotype and is calculated by including ACE- rs4343, ABO- rs495828, and ABO- rs8176746. When ACE- rs4343 is a G-pair gene, ABO- rs495828 is a CC genotype, or ABO- rs8176746 is a CA or AA genotype, a score of 1 is given. At this time, the average ACE activity was found associated with the above-described alleles / genotypes linearly increased (Table 4; p trend value = 3.47x10 ^-53). The mean ACE activity and SBP/DBP level between individuals who took ACEI alone and those who did not take anti-hypertensive drugs increased with increasing ACE activity for even genes or genotypes (Figure 3). According to the medication (0,1)* genetic score interaction items, the p values were 0.0448 (ACE activity), 0.0295 (SBP), and 0.1241 (DBP), respectively.

^a When ACE- rs4343 is a G-pair gene, ABO- rs495828 is a CC genotype or ABO- rs8176746 is a CA or AA genotype, a score of 1 is given.

^{b The} overall p-value was calculated by GLM and corrected for age, sex (0, 1) and ACEI (0, 1).

Example 1

Using the above results, a method for identifying humans with a high risk of developing an ACE-linked disorder can be developed, the steps of which include:

(a) providing a human nucleic acid sample;

(b) genotypic analysis of the ACE activity of the nucleic acid sample by increasing the ACE activity, wherein the ACE activity is increased by the single gene polymorphism SEQ ID NO: 1 (SNP rs4343 of the ACE gene), SEQ ID NO: 2 (SNP rs495828 of ABO gene) and SEQ ID NO: 3 (SNP rs8176746 of ABO gene);

(c) specifying that SEQ ID NO: 1 is a G-pair gene, SEQ ID NO: 2 is a CC genotype, or SEQ ID NO: 3 is a CA or AA genotype of 1 (score);

(d) summing the scores of SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3 to obtain a genetic score ranging from 1 to 4;

The genetic score is an indicator of the high risk of developing angiotensin-converting enzyme-associated abnormalities.

In the present embodiment, the angiotensin-converting enzyme-associated abnormality may be heart disease or hypertension, and the human system may be an individual who needs to prevent heart disease or who needs antihypertensive treatment.

Example 2

In addition, using the above results, a method for predicting a patient's blood pressure response to angiotensin-converting enzyme inhibitor therapy (ACE inhibitor therapy) can be developed accordingly, the steps of which include:

(a) providing a nucleic acid sample of a patient;

(b) genotypic analysis of the ACE activity of the nucleic acid sample by increasing the ACE activity, wherein the ACE activity is increased by the single gene polymorphism SEQ ID NO: 1 (SNP rs4343 of the ACE gene), SEQ ID NO: 2 (SNP rs495828 of ABO gene) and SEQ ID NO: 3 (SNP rs8176746 of ABO gene);

(c) specifying that SEQ ID NO: 1 is a G-pair gene, SEQ ID NO: 2 is a CC genotype, or SEQ ID NO: 3 is a CA or AA genotype of 1 (score);

(d) summing the scores of SEQ ID NO: 1, SEQ ID NO: 2 and SEQ ID NO: 3 to obtain a genetic score ranging from 1 to 4;

The genetic score is an indicator of the patient's blood pressure response to angiotensin-converting enzyme inhibitor treatment.

In this embodiment, the patient system can be an individual in need of prevention of heart disease or in need of antihypertensive treatment.

discuss

The present invention utilizes a two-stage GWAS to identify genes/locus that affect ACE activity. In the process, SNPs in three loci were identified, which are located in two genes, including one SNP located in the ACE structural gene and two SNPs in the ABO gene. Although Caucasians' ACE activity is known to be associated with ABO blood group (non- ABO genotype), two SNPs located in the ABO gene have not previously been found to be directly associated with ACE activity, and are further demonstrated in individuals. In contrast, different numbers of ACE activities increase the dual gene, a differential blood pressure response that may be associated with ACEI.

In the present invention, the discovery of the ABO gene on chromosome 9 is 1 point of the peak of the junction at chromosome 149.4 cM confirmed by the genome-wide linkage family survey (genome-wide linkage family survey) (centi-Morgan, cM) within the length. This is a unique example of two independent study designs, one for individual case-control studies and the other for family-based genetic analysis, showing proximity or location at ABO. Genes and the same novel loci of the ACE gene. The ABO gene encodes a glycosyltransfererase responsible for the transfer of a monosaccharide to a cell-surface H antigen. It consists of three main types: A, B and O. Each has its own specificity and activity. Saccharification of the H antigen of type A or B in the ABO cohort by A-type glycosyltransferase (for N-acetylgalactosamine) and B-type glycosyltransferase (for D-galactose) catalytic. O-dual genes do not produce active enzymes. ABO organizes blood group phenotypes and is found to be associated with infectious diseases, various cancers, blood pressure, and vascular diseases. More specifically, O-type tissue blood types have a low risk association with myocardial infarction, peripheral vascular disease, heart disease, and venous thromboembolism. These findings are only partially explained, for example, the O-form has a smaller amount of factor VIIIc (factor VIIIc) and Willebrand factor than other blood types, and the exact mechanism is still unknown.

The results of the present invention show that bleeding patterns may affect blood pressure and cardiovascular disease by affecting ACE activity. Of great importance is the role of sugar in the ACE configuration and function. Since the ACE molecule is a carrier of the ABO antigen, the oligosaccharide group of the ABO antigen is covalently bonded to the ACE molecule, which may affect the water solubility of ACE and its sensitivity to proteases, resulting in different degrees of activation of the activated ACE enzyme.

In the early 1980s, the Veterans Affairs Cooperative Trial pointed out that different races have different responses to antihypertensive medications. The results of the present invention can aid in its understanding of pharmacogenetics. Although pharmacological genetic associations between ACE I/D genotypes and blood pressure responses have not been demonstrated in large-scale clinical trial studies, by virtue of the present invention, relativity has a majority of ACE activities that increase dual genes/genotypes and Individuals taking ACEI have lower mean blood pressure values than those who did not. Conversely, individuals who do not have such dual genes have a higher blood pressure value than those who do not. It has been shown that the blood pressure response to ACEI may be associated with serum ACE activity, while the ACE activity is determined by multiple genetic loci. If the ACE- rs4343 genotype was used alone, no modification effect on the ACEI blood pressure response in the GenHAT study was found here (data not shown). Since ACEI shows its effect of lowering blood pressure by reducing ACE activity, it is reasonable to observe that ACEI is more effective for individuals with high ACE activity and may be detrimental to individuals with low ACE activity. If the present invention can be confirmed by clinical experiments, including the expression of ACE activity in the ACE , ABO gene, the performance of the dual gene/genotype, or ACE activity may be a reliable predictor of SBP response to ACEI, which can help determine whether the patient should ACEI was chosen as an antihypertensive drug.

There is currently a lot of information on blood pressure regulation, but only a small percentage of the changes affecting blood pressure are attributed to known candidate genes. So far, the high-impact genes for finding high blood pressure have not been very satisfactory. Even the recent impressive large-scale whole-body scan study (WTCCC) has not made a big breakthrough. Hypertension is the only one. No disease with a related SNP (-log p value over 7) was found. The Family Blood Pressure Program in the United States attempted to verify that the WTCCC found a maximum of six SNPs with a -log p value that was unsuccessful, and that subsequent validation studies in ethnic groups in Asia did not yield positive results. Undoubtedly, the factors affecting blood pressure regulation are related to many phenomena, including the multiple roles of genes with low penetration rate, the interaction of genes and genes, ethnic differences, phenotypic heterogeneity, and interaction between genes and the environment. And some environmental factors that are not controlled. These factors add up to the difficulty of finding the genes responsible for hypertension. Unless a very large sample is used, it is difficult to find the causative gene of hypertension, but the gene is often found to be small in large samples. The GWAS study of ACE activity in the present invention is based on the latter method and has been shown to help reduce the scale of hypertension heterogeneity.

The results of the research of the present invention are not all without drawbacks. First, the study of the present invention was performed in individuals with hypertension, although it has been described by a small-scale confirmation study in normal blood pressure individuals. There is still doubt if the results of the present invention are applied to the normal population before further experiments are repeated. Second, the use of hypertension drugs is based on self-reporting in individual cases. This may lead to classification errors and an underestimation of the actual strength of the genotype-drug response association. Third, the phenomenon of differential blood pressure in individuals with high ACE activity and in individuals without this genotype still needs further study, which must be specifically implemented through clinical trial data.

The results of the present invention demonstrate that ACE activity is significantly associated with genetic variation in ACE and ABO loci in hypertensive individuals. In view of our knowledge, the present invention is the first attempt to demonstrate that the ABO gene may play an important role in regulating ACE activity. These results can help explain some of the contradictory results regarding the relationship between ACE activity, blood pressure regulation, and cardiovascular disease. If the results of the present invention can be specifically achieved through clinical trials, they can be applied to fine-tuning strategies for hypertension administration in the future, such as direct administration of ACEI to those individuals who are genetically sensitive to high ACE activity. The results of the present invention provide an example of how GWAS can be used to identify quantitative features, ACE activity, and pharmacogenomics, and can be applied to customized trials of hypertension treatment and the achievement of errors.

It can be seen from the above examples that the gene polymorphism and the method for influencing the angiotensin-converting enzyme activity and the reaction of the angiotensin-converting enzyme activity inhibitor drug provided by the present invention have industrial use value, but the above description It is to be understood that the preferred embodiments of the present invention are susceptible to those skilled in the art, and that various modifications may be made without departing from the spirit and scope of the invention.

The drawings are used to illustrate the principles of the invention, and the same reference numerals are used to indicate the same or similar elements in the various figures:

Figure 1 is a flow chart showing the genotype analysis method;

Figure 2 shows the results of correlation analysis between whole genome scanning correlation and ACE activity using Illumina Infinium II HumanHap 550 SNPs. The closest gene line is noted above, and the significance level is set to -log p =7 (gray dotted line) After adjusting for age, gender (0,1) and ACEI (0,1) and other factors, the correlation between SNP and ACE activity was determined by GLM, and the same linkage disequilibrium domain was selected. Repetitive studies of significant SNPs or SNPs in the coding regions of individual genes;

Figure 3 shows the difference in the average SBP/DBP between the antihypertensive and non-administered individuals with different ACE activities increasing the number of dual genes, using GLM for comparative drug status (0, 1) and ACE activity to increase the dual gene ( Interaction between 0-4) (one who takes more than one drug will be excluded).

Claims (10)

A method for identifying a human having a high risk of developing an ACE-linked disorder, the steps comprising: (a) providing a human nucleic acid sample; (b) increasing the ACE activity of the nucleic acid sample by dual The gene is genotyped, wherein the ACE activity is increased by the single gene polymorphism SEQ ID NO: 1 (SNP rs4343 of ACE gene), SEQ ID NO: 2 (SNP rs495828 of ABO gene) and SEQ ID NO: 3 (SNP rs8176746 of ABO gene); (c) Designation that SEQ ID NO: 1 is a G-pair gene, SEQ ID NO: 2 is a CC genotype, or SEQ ID NO: 3 is a CA or AA genotype of 1 point ( Score); and (d) summing the scores of SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 to obtain a genetic score ranging from 1 to 4; wherein the genetic score is developed An indicator of the high risk of angiotensin-converting enzyme-associated abnormalities. The method of claim 1, wherein the ACE-linked disorder is a heart disease. The method of claim 1, wherein the ACE-linked disorder is hypertension. The method of claim 1, wherein the human system is in need of prevention of heart disease. The method of claim 1, wherein the human system requires antihypertensive treatment. A method for predicting a patient's blood pressure response to angiotensin-converting enzyme inhibitor therapy (ACE inhibitor therapy), the steps comprising: (a) providing a nucleic acid sample of a patient; and (b) increasing the ACE activity of the nucleic acid sample by adding a dual gene Genotyping analysis, wherein the ACE activity is increased by the single gene polymorphism SEQ ID NO: 1 (SNP rs4343 of the ACE gene), SEQ ID NO: 2 (SNP rs495828 of the ABO gene), and SEQ ID NO: 3 (SNP rs8176746 of ABO gene); (c) SEQ ID NO: 1 is a G-pair gene, SEQ ID NO: 2 is a CC genotype or SEQ ID NO: 3 is a CA or AA genotype is 1 (score) And (d) summing the scores of SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 to obtain a genetic score ranging from 1 to 4; wherein the genetic score is for the patient to the blood vessel An indicator of the blood pressure response of a vasopressin-converting enzyme inhibitor. The method of claim 6, wherein the patient is in need of antihypertensive therapy. The method of claim 6, wherein the patient is in need of prevention of heart disease. The use of a single-isolated polynucleotide for the identification of a human having a high risk of developing an angiotensin-converting enzyme-related disorder (ACE-linked disorder) or predicting a patient's treatment for angiotensin-converting enzyme inhibitor (ACE inhibitor therapy) The blood pressure response, wherein the isolated polynucleotide comprises the nucleotide sequence of SEQ ID NO: 2 (SNP rs495828 of the ABO gene) and SEQ ID NO: 3 (SNP rs8176746 of the ABO gene). A use of a single-dual gene for identifying a high risk of developing an ACE-linked disorder in humans, or predicting an ACE inhibitor therapy for a patient The blood pressure response, wherein the single-dual gene line consists of the following groups: SNP of the ACE gene rs4343 is a G-pair gene, the SNP of the ABO gene is a CC genotype, and the SNP rs8176746 of the ABO gene is a CA or AA genotype.