KR20220087271A - A single nucleotide polymorphism marker composition for diagnosing an adverse reactions with angiotensin converting enzyme inhibitor and a method using the same - Google Patents

Abstract

A composition and kit for diagnosing angiotensin converting enzyme inhibitor adverse reactions comprising a single nucleotide polymorphism (SNP) marker, and a method of providing information on the risk of angiotensin converting enzyme inhibitor adverse reactions using the marker. Using a composition for diagnosing angiotensin convertase inhibitor adverse reactions comprising at least one gene single nucleotide polymorphism (SNP) marker selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 according to an aspect Because high-risk groups can be effectively diagnosed or classified, ACEI can be effectively used without adverse reactions to reduce the risk of patients with cardiovascular disease and to provide personalized treatment.

Description

A single nucleotide polymorphism marker composition for diagnosing an adverse reactions with angiotensin converting enzyme inhibitor and a method using the same}

A composition and kit for diagnosing angiotensin converting enzyme inhibitor adverse reactions comprising a single nucleotide polymorphism (SNP) marker, and a method of providing information on the risk of angiotensin converting enzyme inhibitor adverse reactions using the marker.

Angiotensin converting enzyme inhibitor (ACEI) is a first-line treatment for hypertension and congestive heart failure and is known to play a key role in patients with cardiovascular disease. This drug is known to be the most important drug because it lowers blood vessel resistance in the kidneys, increases sodium excretion through urine, lowers blood pressure, and has a cardioprotective effect in the heart remodeling process, and has been proven to improve prognosis.

However, ACEI cannot be used due to adverse reactions or side effects, and it is considered an important problem because the frequency of these side effects is higher in Asians. It is reported in about 20% of countries in East Asia, and in particular, among the above adverse reactions, the most common cough side effect is known to be severe enough that about a quarter of those who experienced it stop taking the drug.

The number of patients with cardiovascular disease is steadily increasing due to the westernization and aging of the distribution of diseases in Korea, but there are no special measures for side effects of ACEI drugs. In particular, ACEI adverse reactions are very common and act as an obstacle to drug maintenance, but studies on the mechanism and prediction are still lacking.

Korean Patent Registration No. 10-1927343 B1

One aspect provides a composition for diagnosing angiotensin converting enzyme inhibitor adverse reactions.

Another aspect provides a kit for diagnosing angiotensin converting enzyme inhibitor adverse reactions.

Another aspect provides information on the risk of angiotensin converting enzyme inhibitor adverse events.

One aspect provides a composition for diagnosing angiotensin converting enzyme inhibitor adverse reactions comprising a polynucleotide detecting a single nucleotide polymorphism (SNP) marker of one or more genes selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 do.

The term “single nucleotide polymorphism (SNP)” refers to a DNA sequence that occurs when a single nucleotide (A, T, C or G) in the genome differs between members of a species or between paired chromosomes of an individual. means the diversity of For example, when three DNA fragments from different individuals (eg, AAGT[A/A]AG, AAGT[A/G]AG, AAGT[G/G]AG) contain differences in a single base, Called two alleles (C or T), in general almost all SNPs have two alleles. Within a population, SNPs can be assigned a minor allele frequency (MAF; the lowest allele frequency at a locus found in a particular population). A single base may be substituted (replaced), removed (deleted), or added (inserted) into the polynucleotide sequence. SNPs can cause translational frame mutations. SNPs are the most common genetic polymorphisms in the human genome, and can cause large differences in each individual depending on the location of the SNPs genetically. For example, when a SNP is present at a protein-encoding position, it may affect the structure of the protein, altering the protein function, and may cause disease. When the SNP is present in a non-coding region that does not encode a protein, that is, if it is present in a promoter or an intron, there is a difference in the expression level of the protein for each, so that the overall activity of the protein can be increased or decreased. In addition, aberrant proteins may be expressed through alternative splicing.

The term “polymorphism” refers to a placement in the sequence of a gene that varies within a population. Polymorphisms consist of different “alleles”. The placement of such polymorphisms can be identified by their position in the gene and the different amino acids or bases found therein. These amino acid variations are the result of two different alleles, two possible variant bases, C and T. Because the genotype consists of two different distinct alleles, any of several possible variants can be observed in any one individual (eg, CC, CT or TT in this example). Individual polymorphisms are also known to those skilled in the art and are used, for example, in the Single Nucleotide Polymorphism Database (dbSNP) of Nucleotide Sequence Variation available on the NCBI website. Identifier ("reference SNP", "refSNP" or "rs#"). The term “genotype” also refers to a specific allele of a particular gene in a cell or tissue sample.

The single nucleotide polymorphism may be included in a gene coding sequence, a non-coding region of a gene, or intergenic regions between genes. SNPs in the gene coding sequence do not necessarily accompany the mutation of the amino acid sequence of the target protein due to the degeneracy of the genetic code. SNPs that form the same polypeptide sequence are said to be synonymous, and SNPs that form different polypeptide sequences are said to be non-synonymous.

The non-synonymous SNP may be missense or nonsense, and the missense mutation may result in another amino acid and the nonsense mutation may form an immature stop codon. SNPs that are not present in protein-coding sites can cause gene silencing, transcription factor binding, or non-coding RNA sequences.

The term “diagnosis” means ascertaining the presence or inventive potential of a pathological condition. Among them, one aspect is particularly useful for diagnosing angiotensin converting enzyme inhibitor adverse reactions. Since the SNP of one or more genes selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 of an aspect functions as such an indicator, it can be used as an angiotensin converting enzyme inhibitor adverse reaction diagnostic factor. That is, measurement of polymorphisms in these genes can be used as a useful indicator (diagnostic marker) for diagnosing or judging whether an individual is capable of exhibiting an angiotensin converting enzyme inhibitor adverse reaction.

The at least one gene single nucleotide polymorphism (SNP) marker selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 is rs2252726, rs8176786, rs10766756, rs2252726, rs8176746, Rs4149056, rs8137619, rs8176720, rs817674749, rs383067747, rs383067763 , may include one or more single nucleotide polymorphism (SNP) markers selected from the group consisting of rs10154841 and rs2774602. Additionally, the single nucleotide polymorphism marker may include those present in the genes of RBFOX3 and BDKRB2, for example, rs2061538 or rs5224 may be included as single nucleotide polymorphism markers, respectively.

The NELL1 gene is known to exist on chromosome 11, and its GenBank Accession Gene ID is 4745. The ABO gene is known to exist on chromosome 9, and its GenBank Accession Gene ID is 28. The PDE11A gene is known to exist on chromosome 2, and its GenBank Accession Gene ID is 50940. The CACNA1D gene is known to exist on chromosome 3, and its GenBank Accession Gene ID is 776. The SLCO1B gene is known to exist on chromosome 12, and its GenBank Accession Gene ID is 10599. The RBFOX3 gene is known to exist on chromosome 17, and its GenBank Accession Gene ID is 146713. The BDKRB2 gene is known to exist on chromosome 14, and its GenBank Accession Gene ID is 624.

A person skilled in the art will be able to easily identify the position and sequence of the mutation using the registration number. The specific sequence corresponding to the number registered in the UCSC genome browser or GenBank may change slightly over time. It will be apparent to those skilled in the art that the scope of the present invention also extends to such altered sequences.

The single nucleotide polymorphism marker is the rs8176786 single nucleotide polymorphism site in the NELL1 gene where the 20,959,394th C/T base of human chromosome 11 exists or the rs10766756 single nucleotide polymorphism in the NELL1 gene where the 21,009,736th T/C base of the human chromosome 11 exists. It may include a region. In addition, the single nucleotide polymorphism marker may include the rs2252726 single nucleotide polymorphism site in the PDE11A gene in which the 178,844,286 C/T bases of human chromosome 2 exist.

In addition, the angiotensin converting enzyme inhibitor adverse reaction (ACE Inhibitor Intolerance) refers to side effects that can be exhibited by the subject receiving the drug when the angiotensin converting enzyme inhibitor is administered, for example, the adverse reaction is an angiotensin converting enzyme inhibitor. It may be selected from the group consisting of cough, hypotension, somnolence, dyspnea, chest pain, edema, creatine elevation, and headache.

The composition of one aspect may include a primer, a probe, or an antisense nucleotide capable of detecting a mononucleotide polymorphism marker of the genes of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1. In one embodiment, the primer or its pair, probe or antisense nucleotide may be included, and each of the primer pair, probe or antisense nucleotide is specific for a single polymorphic marker of the genes of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1. can be positively combined.

The primer may be characterized by amplifying the sequence so that the amplified PCR product can obtain a comparison sequence of an optimal size to effectively diagnose and predict glioma using a next-generation sequencer (NGS).

The primer, probe or antisense nucleotide may be a polynucleotide labeled with a detectable label. The detectable label is a label material capable of generating a detectable signal, and may be a label material capable of generating a detectable signal including a fluorescent material, for example, substances such as Cy3 and Cy5. The detectable label may confirm the hybridization result of the nucleic acid.

The term “primer” refers to a single-stranded oligonucleotide capable of serving as an initiation point in the polymerization of nucleotides by a polymerase. For example, the primer can serve as an initiation point for template-directed DNA synthesis under suitable conditions, i.e., in the presence of four different nucleoside triphosphates and a polymerase, at a suitable temperature and in a suitable buffer. It may be a single-stranded oligonucleotide. A suitable length of a primer may depend on a variety of factors, such as temperature and the use of the primer. The primer may have a length of 5 to 100 nt, 5 to 70 nt, 10 to 50 nt, or 15 to 30 nt. For example, the shorter the length of the primer, the more stable a hybridization complex can be formed with the template at a low annealing temperature.

The primer may further include a nucleotide analogue such as phosphorothioate or alkylphosphorothioate, a peptide nucleic acid, or an intercalating agent. In addition, it may further include a labeling material that emits fluorescence, phosphorescence, or radioactivity. The fluorescent label material may be VIC, NED, FAM, PET, or a combination thereof. The labeling material may be labeled at the 5' end of the polynucleotide. In addition, the radiolabelled material may be incorporated into the amplification product through a PCR reaction using a polymerase chain reaction (PCR, hereinafter referred to as 'PCR') reaction solution to which a radioisotope such as 32P or 35S is added. have.

The term “probe” refers to a polynucleotide capable of sequence-specific binding to a complementary polynucleotide strand. The probe may have a length of 5 to 100 nt, 10 to 90 nt, 15 to 80 nt, 20 to 70 nt, or 30 to 50 nt. The probe may be used in a hybridization method, for example, a method using a microarray, Southern blotting, dynamic allele-specific hybridization, and a DNA chip. Microarray is used in a sense known in the art, and for example, a probe or a group of probes may be immobilized on a plurality of separated regions on a substrate. The substrate is a suitable rigid or semi-rigid support, including, for example, membranes, filters, chips, slides, wafers, fibers, magnetic or non-magnetic beads, gels, tubing, plates, polymers, microparticles and capillaries. can do. The probe or a probe complementary thereto can be used in a method capable of hybridizing with a nucleic acid obtained from an individual and measuring the degree of hybridization obtained therefrom.

The probe may be single-stranded, preferably an oligodeoxyribonucleotide of 10 to 30 consecutive nucleotides including a nucleotide complementary to the SNP or SNP. In addition, the probes complementary to SNPs may include native dNMPs (ie, dAMP, dGMP, dCMP and dTMP), nucleotide analogues or derivatives. In addition, the probe may also include ribonucleotides. For example, the probe may be a backbone modified nucleotide, such as a peptide nucleic acid (PNA) (M. Egholm et al., Nature, 365:566-568 (1993)), phosphorothioate DNA, phosphorodithioate DNA, Phosphoroamidate DNA, amide-linked DNA, MMI-linked DNA, 2'-O-methyl RNA, alpha-DNA and methylphosphonate DNA, sugar modified nucleotides such as 2'-O-methyl RNA, 2' -Fluoro RNA, 2'-amino RNA, 2'-O-alkyl DNA, 2'-O-allyl DNA, 2'-O-alkynyl DNA, hexose DNA, pyranosyl RNA and anhydrohexitol DNA , and nucleotides with base modifications such as C-5 substituted pyrimidines (substituents are fluoro-, bromo-, chloro-, iodo-, methyl-, ethyl-, vinyl-, formyl-, ethyl- , propynyl-, alkynyl-, thiazolyl-, imidazoryl-, pyridyl- including), 7-deazapurine having a C-7 substituent (substituents are fluoro-, bromo-, chloro-, io do-, methyl-, ethyl-, vinyl-, formyl-, alkynyl-, alkenyl-, thiazolyl-, imidazoryl-, pyridyl-), inosine and diaminopurine.

Another aspect is an angiotensin converting enzyme inhibitor adverse reaction comprising a primer, probe or antisense nucleotide that specifically binds to one or more gene single nucleotide polymorphism (SNP) marker loci selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 kits for diagnosing

The kit includes a single nucleotide polymorphism (SNP) probe that specifically binds to or amplified one or more genes selected from the group consisting of ABO, PDE11A, CACNA1D and SLCO1B1 for the diagnosis of angiotensin converting enzyme inhibitor adverse reactions. In addition to the primers present, one or more other component compositions, solutions, or devices suitable for the assay method may be included.

In one aspect of the kit, an antigen can be attached to the surface of a support such as glass or nitrocellulose, and the microarray manufacturing technique is described, for example, in Schena et al., 1996, Proc Natl Acad Sci USA. 93(20):10614-9; Schena et al., 1995, Science 270(5235):467-70; and U.S. Pat. Nos. 5,599,695, 5,556,752 or 5,631,734. Fluorescence intensity can be measured using a scanning confocal microscope, for example, Affymetrix, Inc. or from Agilent Technologies, Inc. and the like.

The kit may be a kit for predicting genetic mutation of a target gene. Kit is used in the sense known in the art. The kit may include, for example, the polynucleotide as described above and items necessary for its specific use. It may include a reagent necessary for a method of its use together with the polynucleotide as described above.

The kit of one aspect may include a reagent for performing an amplification reaction, and may include a heat-resistant DNA polymerase, dNTPs, a buffer, and the like. In addition, the kit of one aspect may further comprise a user's guide describing optimal conditions for conducting the reaction. A handbook is a printout explaining how to use the kit, eg, how to prepare a PCR buffer, and suggested reaction conditions. Instructions include a brochure in the form of a pamphlet or leaflet, a label affixed to the kit, and instructions on the surface of the package containing the kit. In addition, the guide includes information published or provided through electronic media such as the Internet.

Another aspect includes the steps of 1) obtaining a nucleic acid sample from a biological sample of an individual; 2) amplifying the polymorphic site of one or more gene single nucleotide polymorphism (SNP) markers selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 from the obtained biological sample or hybridizing with a probe; and 3) determining the genotype of the amplified or hybridized polymorphic site and, if the genotype corresponds to a risk allel, determining an angiotensin converting enzyme inhibitor adverse reaction risk risk How to provide information about.

The term “sample” includes blood and other liquid samples of biological origin, biopsy specimens, solid tissue samples such as tissue cultures, or cells derived therefrom, for example consisting of the patient's blood, plasma, serum, urine and saliva. It may be one or more selected from the group. More specifically, it may be selected from the group consisting of blood, serum, plasma, lymph, cerebrospinal fluid, ascites, urine and tissue biopsy, and the sample may be pre-treated before use for detection. For example, it may include filtration, distillation, extraction, concentration, inactivation of interfering components, addition of reagents, and the like. In addition, nucleic acids and proteins can be isolated from the sample and used for detection.

The step of obtaining a nucleic acid sample from the biological sample may be performed by a conventional DNA isolation method. For example, the target nucleic acid is subjected to polymerase chain reaction (PCR), ligase chain reaction (LCR), transcription amplification, or realtime-nucleic acid sequence based amplification (realtime-nucleic acid). It can be obtained by amplifying it through sequence based amplification: NASBA) and purifying it.

One or more gene single nucleotide polymorphism (SNP) markers selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 of step 2) are rs2252726, rs8176786, rs10766756, rs2252726, rs8176746, rs4149056, rs817672019, rs8176720, rs8176749 , rs77063376, rs3830637, rs10154841 and rs2774602 may be at least one selected from the group consisting of, and may additionally include rs2061538 or rs5224 group.

The step of determining the risk group in step 3) is when the genotype of the rs2252726 single nucleotide polymorphism site is “T”, when the genotype of the rs8176786 single nucleotide polymorphism site is “T”, the genotype of the rs10766756 single nucleotide polymorphism site is When “C”, when the genotype of the rs2252726 single-nucleotide polymorphism site is “T”, when the genotype of the rs8176746 single-nucleotide polymorphism site is “T”, when the genotype of the rs4149056 single-nucleotide polymorphism site is “C” , when the genotype of the rs8176719 single nucleotide polymorphism site is “C”, when the genotype of the rs8176749 single nucleotide polymorphism site is “T”, when the genotype of the rs8176747 single nucleotide polymorphism site is “G”, the rs8176720 single nucleotide When the genotype of the polymorphic site is “C”, when the genotype of the rs77063376 single nucleotide polymorphism site is “A”, when the genotype of the rs3830637 single nucleotide polymorphism site is “GGA”, the genotype of the rs10154841 single nucleotide polymorphism site is In the case of “T”, if the genotype of the rs2774602 single nucleotide polymorphism site is “T”, it may be determined that the risk of an angiotensin converting enzyme inhibitor adverse reaction is high, and additionally, the genotype of the rs2061538 single nucleotide polymorphism site is “ In the case of "A", if the genotype of the rs5224 single nucleotide polymorphism site is "A", the risk of angiotensin converting enzyme inhibitor adverse reactions is high, and the individual is judged as a risk group for angiotensin converting enzyme inhibitor adverse reactions. .

In the method, the step of analyzing the genotype is a method of determining a nucleotide or base sequence, and sequencing, hybridization by microarray, allele specific PCR, dynamic allele-specific hybridization hybridization), PCR extension analysis, PCR-SSCP (PCR-single strand conformation polymorphism), and TaqMan method may be performed by one or more techniques selected from the group consisting of. In the step of analyzing the genotype, the composition or kit may be used.

The sequencing analysis may use a conventional method for sequencing, and may be performed using an automated gene analyzer. Allele-specific PCR refers to a PCR method of amplifying a DNA fragment in which the single nucleotide mutation is located with a primer set including a primer designed with the 3' end of the base in which the single nucleotide mutation is located. In PCR extension analysis, first, a DNA fragment containing a base in which a single nucleotide mutation is located is amplified with a pair of primers, and then all nucleotides added to the reaction are inactivated by dephosphorylation, and here an extension primer specific for the single nucleotide mutation, A primer extension reaction is performed by adding a dNTP mixture, dideoxynucleotide, reaction buffer and DNA polymerase. The TaqMan method involves designing and manufacturing primers and TaqMan probes to amplify a desired DNA fragment, labeling probes of different alleles with FAM and VIC (Applied Biosystems), and amplifying and analyzing them.

The subject of the method may be a mammal, including a human, specifically, an East Asian, more specifically, a Korean.

Using a composition for diagnosing angiotensin convertase inhibitor adverse reactions comprising at least one gene single nucleotide polymorphism (SNP) marker selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 according to an aspect Because high-risk groups can be effectively diagnosed or classified, ACEI can be effectively used without adverse reactions to reduce the risk of patients with cardiovascular disease and to provide personalized treatment.

1 is a diagram illustrating a result of performing target NGS data analysis on a subject sample using a designed target panel.

Hereinafter, the present invention will be described in more detail through examples. However, these Experimental Examples and Examples are for illustrative purposes of the present invention, and the scope of the present invention is not limited to these Experimental Examples and Examples.

Example 1. Identification of adverse reaction-related gene mutations according to the use of angiotensin converting enzyme inhibitor (ACEI) through NGS analysis of candidate genes

1.1 Selection of candidate genes through GWAS analysis

Among the retrospective cohort for cardiovascular disease established at Yonsei University Severance Hospital, the group with adverse events (143 patients) through the analysis of adverse reaction clinical information according to the use of ACEI among subjects who had GWAS data among subjects who had experience using ACEI ) and the group without adverse reactions (85 persons). At this time, the group with the adverse reaction included all the groups that primarily showed other adverse reactions including cough, such as hypotension, somnolence, dyspnea, chest pain, creatine elevation, and headache.

For these subjects, full-length gene analysis was performed after data QC using the CEL file, which is the Axiom Genome-Wide ASI Array chip raw data file. There was no subject excluded from sample QC, and association analysis was performed on 668,091 marker positions after marker QC for a total of 904,333 marker positions. Criteria used for marker QC were 1) p-value ≥ 1E-07 of Hardy-Weinberg Equilibrium (HWE) and 2) case > 0.95 and control > 0.95 of call rate, and last as MAF (Minor Allele Frequency): case > 0.01 or control > 0.01.

For association analysis, ACEI adverse reaction-related gene mutations were screened through logistic regression analysis based on an additive model using PLINK, and related new genetic mutations were secured. A total of five candidate genes related to ACEI adverse reactions were identified as CRIM1 (Cysteine Rich Transmembrane BMP Regulator 1), NELL1 (NEL-like protein 1), and CACNA1D ( Calcium channel voltage-dependent L type alpha 1D subunit), VOPP1 and MYBPC1 were selected.

1.2 Selection of candidate genes through bioinformatics analysis

Genes known to be involved in the ACEI drug metabolism pathway based on bioinformatics analysis, in particular, mutations related to the renin-angiotensin system and bradykinin pathway were selected as additional candidate genes.

As a result of a large-scale full-length genome analysis of ACEI intolerance conducted by the European PREDICTION-ADR consortium on 972 patients and 5,161 patient controls, 8 SNPs from 4 gene regions were found to be statistically significant. Specifically, RBFOX3 (RNA Binding Protein Fox-1 Homolog 3), GABRG2 (Gamma-aminobutyric acid receptor subunit gamma-2), SH2B1 (sarcoma (Src) homology 2 (SH2) B adapter protein 1), MBOAT1 (membrane bound O- It is an acyltransferase domain containing 1) gene, and it was confirmed that the gene showing the strongest correlation was the RBFOX3 gene.

As a result of the whole genome analysis study of cough induction related to ACEI use in Swedish subjects, it was confirmed that CLASP1, PDE11A, KCNMB2, TGFA, SLC38A6 and MMP16 genes were statistically related to cough induction, and the use of ACEI conducted in the eMERGE network As a result of the full-length gene analysis related to cough induction, the KCNIP4 gene was found to be related to the induction of cough according to the use of ACEI.

Example 2. Confirmation of final genes related to adverse reactions according to the use of angiotensin converting enzyme inhibitors (ACEI) through NGS analysis of candidate genes

2.1 Design of the final target panel for NGS analysis

Through the full-length genome analysis and bioinformatics study of Example 1, 34 genes and 14 marker mutation sites, which are candidate gene sites related to adverse reactions according to the use of angiotensin converting enzyme inhibitor (ACEI), were finally selected, and the results are shown in Table 1 shown in Then, a target capture panel based on the hybridization method was finally designed.

[Table 1]

2.2 Target panel capture and NGS data analysis

Target NGS analysis was performed on a total of 337 subject samples using the target panel designed in Example 2.1, and the results are shown in FIG. 1 . As confirmed in FIG. 1 , the average depth of all the checked samples was 300X or more, and it was possible to obtain a depth of at least 20x or more in an average of 99.79% of the area. In addition, a genotype call was performed for the gene mutation location for each sample using the GATK calling program for a region of 20X or more, and a total of 405 mutations were identified.

2.3 Analysis of associations related to adverse events according to the use of ACEI - with respect to all adverse events

For the mutations identified in Example 2.2, association analysis related to adverse reactions according to the use of ACEI was performed. Of the 337 patients, 213 patients experienced ACEI adverse reactions, coughing 112 patients, hypotension 28 patients, somnolence 21 patients, dyspnea 11 patients, chest pain 6 patients, edema 5 patients, creatine elevation 5 patients, and headache. There were 5 patients, and 124 patients in the group that did not experience adverse reactions were found. Thereafter, the association analysis was performed with respect to the adverse reaction experience group and the unexperienced group, and the results are shown in Table 2.

[Table 2]

Association analysis results for the ACEI adverse reaction experience group (n=213) and the unexperienced group (n=124)

As confirmed in Table 2 above, as a result of the association analysis for all adverse reactions caused by ACEI, the NELL1 intron gene mutation rs10766756, which was confirmed to be meaningful in the Korean GWAS analysis, was significantly associated with the ACEI adverse reaction in the target sequencing analysis. This was confirmed. Additionally, the NELL1 exonic mutation rs8176786 (NELL1 p.Arg382Trp) was also confirmed to be a mutation highly related to ACEI adverse events, and the MBOAT1 gene mutation marker rs10946364 showed a value of 0.04 in the TT vs TA&AA analysis, which is the genotyping analysis of all adverse reaction experience groups. It was found that the correlation was high.

In addition, a case in which the occurrence of an adverse reaction occurred earlier than the median value of 5 months was classified as an extreme case. Further, the results are shown in Table 3.

[Table 3]

Association analysis results for the group experiencing severe ACEI adverse reactions (n=118) and the unexperienced group (n=124)

As confirmed in Table 3, it was confirmed that the NELL 1 gene mutation, Rs10766756, showed a significant difference even in the group experiencing severe ACEI adverse reactions. In addition, it was confirmed that the PDE11A intron mutation Rs2252726 and the RBFOX3 intron mutation Rs2061538 have a high correlation.

2.4 Analysis of associations related to adverse events according to the use of ACEI - Each adverse reaction to ACEI in connection with

Among the ACEI adverse reaction experience group, the most common adverse reaction, cough adverse reaction experience group, was 112 people, followed by hypotension experience group 28 people, dizziness experience group 21 people, dyspnea experience group 11 They appeared in the order of persons, and correlation analysis was performed on 124 persons in the adverse reaction experience group and the US experience group, respectively.

Among them, the median cough occurrence was 3 months in the cough adverse reaction experience group, and patients who developed an adverse reaction earlier than 3 months were classified as severe cases. Accordingly, the results of analyzing the association between the cough adverse reaction experience group (n=112 people) and the non-experience group (n=124 people) are shown in Table 4. In addition, correlation analysis between the severe cough adverse reaction experience group (n=58 people) and the unexperienced group (n=124 people) was additionally conducted, and the results are shown in Table 5.

[Table 4]

Association analysis results for the ACEI cough adverse reaction experience group (n=112 people) and the non-experience group (n=124 people)

[Table 5]

Results of association analysis between the ACEI severe cough adverse reaction group (n=58) and the unexperienced group (n=124)

As can be seen in Tables 4 and 5 above, it was confirmed that the NELL 1 gene mutation was statistically significant in the analysis of the cough adverse reaction and its severe experience group, and the NELL1 intron gene mutation Rs10766756 and the NELL1 exonic mutation Rs8176786 (NELL1 p.Arg382Trp) was also confirmed to be a mutation highly related to ACEI cough adverse reaction. In addition, Rs5224, a mutation in the BDKRB2 gene, which is a Bradykinin receptor that regulates kinin level, was also found to be meaningful in genotyping analysis, and this mutation was also confirmed to be meaningful in all ACEI adverse reactions and severe experience group analysis (synonymous) appeared to be mutated. In addition, it was confirmed that the PDE11A intron gene mutation, Rs2252726, was highly correlated with the ACEI cough adverse reaction in the genetic analysis.

Table 6 shows the results of analyzing the association between the hypotensive adverse reaction experience group (n=28 patients) and the non-experienced group (n=124 patients) in the hypotensive adverse reaction experience group.

[Table 6]

Association analysis results for the ACEI hypotensive adverse reaction experience group (n=28 people) and the unexperienced group (n=124 people)

As confirmed in Table 6 above, as a result of analyzing the association with the hypotensive adverse reaction-related gene mutation, it was confirmed that the ABO gene mutation and the mutation in the coding region of the PDE11A gene were mainly related. Specifically, it was confirmed that the NELL1 intron gene mutations Rs10766756 and Rs8176719, Rs77038916, Rs8176749, Rs8176747, Rs8176720, and Rs77063376 mutations represent significant differences.

Table 7 shows the results of analyzing the association between the drowsiness experience group (n=21 people) and the US group (n=124 people) in the case of the drowsiness experience group.

[Table 7]

Association analysis results for the ACEI sleepy adverse reaction experience group (n=21) and the non-experienced group (n=124)

As confirmed in Table 7, it was confirmed that the sleepiness adverse reaction was related to various genetic mutations. Specifically, it was confirmed that the Rs8176719, Rs77063376, Rs3830637, Rs10154841, and Rs2774602 mutations represent a significant difference.

In the case of the respiratory distress adverse reaction experience group, the results of analyzing the association between the respiratory distress adverse reaction experience group (n=11 people) and the unexperienced group (n=124 people) are shown in Table 8.

[Table 8]

Association analysis results for the ACEI dyspnea adverse reaction experience group (n=11 people) and the non-experience group (n=124 people)

As confirmed in Table 8, it was confirmed that the PDE11A gene mutation, Rs3830637, exhibited a high degree of correlation as a result of analysis of the association with respiratory distress adverse reaction.

Example 3. Confirmation of reproducibility of association through identification of genetic variants

Four mutations (Rs2252726, Rs8176746, Rs8176786, Rs10766756) and sleepiness abnormality were found to be specific and highly correlated in Koreans, centering on gene mutations related to all adverse reactions and cough adverse reactions among the SNPs identified in the association study of Example 2 The mutation found in the reaction but the SLCO1B1 Rs4149056 gene mutation, which was found to be a functional genetic mutation, was included in the reproducibility study, and secondary validation was performed. The five SNPs are shown in Table 9.

[Table 9]

For the reproducibility study, 142 patients (including 28 coughs) and 109 control patients were selected as the independent patient group for the reproducibility study. Reproducibility analysis was performed on adverse events and cough in the independent patient group, respectively, and the independent patient group and target sequencing patient group were combined. Two analyzes were also performed in one patient group. In this process, patients who experienced adverse reactions after 60 months were excluded from the analysis.

[Table 10]

[Table 11]

As can be seen in Table 11 above, it can be confirmed that Rs8176786, a mutation of NELL1 p.Arg382Trp, Rs10766756, a mutation of NELL1 intron, and Rs2252726, a mutation of PDE11A intron are significant mutations related to ACEI adverse reactions, especially coughing adverse reactions. could Accordingly, if the high-risk group for ACEI adverse reactions is classified according to the presence of risk alleles of the three mutations, ACEI can be used safely and with high patient satisfaction, enabling customized treatment for patients with cardiovascular disease.

Claims (11)

A composition for diagnosing an angiotensin converting enzyme inhibitor adverse reaction, comprising a polynucleotide detecting a single nucleotide polymorphism (SNP) marker of one or more genes selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1. The method according to claim 1, wherein the single nucleotide polymorphism (SNP) marker is one or more selected from the group consisting of rs2252726, rs8176786, rs10766756, Rs2252726, rs8176746, rs4149056, rs8176719, rs8176749, rs8176747, rs8176720, rs77063376, rs3830637, rs10154841 and rs phosphorus composition. The composition for diagnosing angiotensin converting enzyme inhibitor adverse reaction according to claim 1, wherein the polynucleotide comprises a primer, a probe, or an antisense nucleotide. The method according to claim 2, wherein the single nucleotide polymorphism marker is the rs8176786 single nucleotide polymorphism site in the NELL1 gene in which the 20,959,394th C/T base of human chromosome 11 exists or the NELL1 gene in which the 21,009,736th T/C base of human chromosome 11 exists. Angiotensin converting enzyme inhibitor adverse reaction diagnostic composition comprising a single nucleotide polymorphism site within rs10766756. The composition for diagnosing an angiotensin converting enzyme inhibitor adverse reaction according to claim 2, wherein the single nucleotide polymorphism marker comprises the rs2252726 single nucleotide polymorphism site in the PDE11A gene where the 178,844,286th C/T base of human chromosome 2 exists. The composition for diagnosing angiotensin converting enzyme inhibitor adverse reaction according to claim 1, wherein the angiotensin converting enzyme inhibitor adverse reaction is selected from the group consisting of cough, hypotension, somnolence, dyspnea, chest pain, edema, creatine elevation and headache. To diagnose an angiotensin converting enzyme inhibitor adverse reaction comprising a primer, probe or antisense nucleotide that specifically binds to a single nucleotide polymorphism (SNP) marker locus of one or more genes selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 kit for. 1) obtaining a nucleic acid sample from a biological sample of an individual;
2) amplifying the polymorphic site of one or more gene single nucleotide polymorphism (SNP) markers selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 from the obtained biological sample or hybridizing with a probe; and
3) Check the genotype of the amplified or hybridized polymorphic site to determine the risk of angiotensin-converting enzyme inhibitor adverse reactions, including the step of determining the angiotensin-converting enzyme inhibitor adverse reaction risk group if the genotype corresponds to a risk allele How to provide information about. The method according to claim 8, wherein the biological sample is one or more selected from the group consisting of blood, plasma, serum, urine and saliva. The method according to claim 8, wherein the at least one gene single nucleotide polymorphism (SNP) marker selected from the group consisting of NELL1, ABO, PDE11A, CACNA1D and SLCO1B1 in step 2) is rs2252726, rs8176786, rs10766756, rs2252726, rs8176746, rs4149056, rs8176719, rs8176749, rs8176747, rs8176720, rs77063376, rs3830637, rs10154841 and rs2774602 method that is at least one selected from the group consisting of. The method of claim 8, wherein the step of determining the risk group of step 3)
When the genotype of the Rs2252726 single nucleotide polymorphism site is “T”,
When the genotype of the Rs8176786 single nucleotide polymorphism site is “T”,
When the genotype of the Rs10766756 single nucleotide polymorphism site is “C”,
When the genotype of the Rs2252726 single nucleotide polymorphism site is “T”,
When the genotype of the Rs8176746 single nucleotide polymorphism site is “T”,
When the genotype of the Rs4149056 single nucleotide polymorphism site is “C”,
When the genotype of the Rs8176719 single nucleotide polymorphism site is “C”,
When the genotype of the Rs8176749 single nucleotide polymorphism site is “T”,
When the genotype of the Rs8176747 single nucleotide polymorphism site is “G”,
When the genotype of the Rs8176720 single nucleotide polymorphism site is “C”,
When the genotype of the Rs77063376 single nucleotide polymorphism site is “A”,
When the genotype of the Rs3830637 single nucleotide polymorphism site is “GGA”,
When the genotype of the Rs10154841 single nucleotide polymorphism site is “T”, or
When the genotype of the Rs2774602 single nucleotide polymorphism site is “T”, the method is to determine the individual as a risk group for angiotensin converting enzyme inhibitor adverse reactions.

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