Predicting or diagnosing composition for the occurrence or timing of allergic rhinitis, Diagnosing kit, Method for providing information and Screening method for drugs for preventing or treating allergic rhinitis using the same

Abstract

The invention relates to a composition, a kit, and a method for diagnosing allergic rhinitis using the same, wherein COL5A2, CDK2AP2, COQ9, and PYM1 proteins or genes showing differences in expression in blood transcriptome analysis of an early-onset allergic rhinitis group, a late-onset allergic rhinitis group, and a normal group are identified, and the early and late onset of allergic rhinitis can be predicted through Decision tree prediction model analysis using the same, and can be utilized for early diagnosis of the phenotype and prognosis prediction of allergic rhinitis, as well as being useful in the field of future new drug development targets and treatments.

Description

Technology Field

[0001] The invention relates to a composition for predicting or diagnosing whether or when allergic rhinitis occurs, a diagnostic kit, a method for providing information, and a method for selecting a preventive or therapeutic agent for allergic rhinitis. Background Technology

[0002] Rhinitis is caused by chronic or acute inflammation of the nasal mucosa by viruses, bacteria, or irritants. The inflammation typically leads to the production of excessive amounts of mucus, causing a runny nose, nasal congestion, and postnasal drip.

[0003] Allergic rhinitis affects more than 20% of the world's population, and its prevalence is increasing every year. Allergic rhinitis is a proinflammatory immune response to indoor and outdoor allergens, such as dust and pollen. It causes problems in social life, sleep, school, and work, and a patient's quality of life can vary depending on the severity and duration of the condition. Therefore, early diagnosis and prevention of rhinitis are crucial.

[0004] Recent analysis of the natural course of rhinitis has confirmed the existence of various phenotypes, and diagnosing rhinitis is difficult due to factors such as reduced sensitivity and specificity to allergen reactions in subjects. Furthermore, although pediatric allergic rhinitis is one of the most common respiratory diseases in Korean children with a prevalence of 40–50%, it is even more difficult to confirm whether symptoms such as runny nose, nasal congestion, or postnasal drip occurring at a young age are actually rhinitis.

[0005] Therefore, there is a need to develop new prognostic biomarkers based on the phenotype and endotype of allergic rhinitis. The problem to be solved

[0006] One aspect provides a diagnostic composition capable of predicting whether or when allergic rhinitis occurs, comprising a preparation capable of measuring the activity or expression level of one or more proteins or genes selected from the group consisting of COL5A2, CDK2AP2, COQ9 and PYM1.

[0007] Another aspect is to provide a kit comprising a diagnostic composition capable of predicting whether or when allergic rhinitis occurs.

[0008] Another aspect is to provide a method for providing information for predicting whether and when allergic rhinitis occurs, comprising the step of measuring the activity or expression level of COL5A2, CDK2AP2, COQ9, or PYM1 proteins or genes in biological samples isolated from a target individual.

[0009] Another aspect is providing a screening method for the prevention or treatment of allergic rhinitis. means of solving the problem

[0010] One aspect provides a diagnostic composition capable of predicting whether or when allergic rhinitis occurs, comprising a preparation capable of measuring the activity or expression level of one or more proteins or genes selected from the group consisting of COL5A2, CDK2AP2, COQ9, and PYM1.

[0011] The term “diagnosis” in this specification may include determining the susceptibility of a subject to a specific disease or condition, determining whether the subject currently has a specific disease or condition, determining the prognosis of a subject with a specific disease or condition, or therametrics (e.g., monitoring the condition of a subject to provide information on therapeutic efficacy). Specifically, the diagnosis is to predict whether or when allergic rhinitis will occur. The prediction means a prospect or preliminary evaluation regarding the progression of the disease or recovery. Specifically, the prediction is to determine whether or when allergic rhinitis will occur in an individual at risk of developing allergic rhinitis.

[0012] The term "allergic rhinitis" in this specification refers to a proinflammatory immune response to indoor and outdoor allergens, such as dust or pollen, which is caused by chronic or acute inflammation of the nasal mucosa. This inflammation generally causes a runny nose, nasal congestion, and postnasal drip, and leads to the production of an excessive amount of mucus.

[0013] The term "time of onset of allergic rhinitis" in this specification refers to the time when symptoms of allergic rhinitis first appear. The time of onset can be divided into early onset and late onset. Early onset means that symptoms appear before the age of 2-3 and reach a peak at the age of 4-5, and late onset means that symptoms appear after the age of 4-5 and reach a peak at the age of 6-8.

[0014] In this specification, the term normal group (G1) refers to a group of subjects who have not developed allergic rhinitis.

[0015] In this specification, the term early onset allergic rhinitis group (G2) refers to a group in which symptoms of allergic rhinitis began before the age of 2-3 years.

[0016] In this specification, the term allergic rhinitis late erection onset group (G2) refers to a group in which symptoms of allergic rhinitis begin after the age of 4-5.

[0017] In this specification, the term "biomarker" or "marker" refers to a substance capable of distinguishing between a normal population and individuals at risk of developing allergic rhinitis to diagnose whether allergic rhinitis has occurred or to predict the timing of its onset. It may include all organic biomolecules, such as polypeptides, proteins, nucleic acids, genes, lipids, glycolipids, glycoproteins, or sugars, that show an increase or decrease in individuals at risk of developing allergic rhinitis. In one embodiment, it may be a gene whose expression level changes in individuals at risk of developing allergic rhinitis, but is not limited thereto.

[0018] The terms "COL5A2 (NCBI GenBank accession number, NM_000393), CDK2AP2 (NM_001271849), COQ9 (NM_020312) and PYM1 (NM_001143853)" in this specification are genes or proteins that show differences in expression in blood transcriptome analysis of a normal group (G1), an early-onset allergic rhinitis group (G2), and a late-onset allergic rhinitis group (G3).

[0019] The above genes may be obtained from known databases such as NCBI GenBank. For example, but not limited to, the above COL5A2 may refer to a nucleic acid containing the nucleotide sequence of NCBI Gene ID: 0000 in humans or a nucleic acid containing the nucleotide sequence of EnsEMBL ID: ENSG00000129988. It also refers to proteins in which the above genes are expressed, namely "COL5A2 (NCBI accession number, NP_000384), CDK2AP2 (NP_001258778), COQ9 (NP_064708), and PYM1 (NP_001137325)".

[0020] In this specification, the term "measurement of gene expression level" refers to a process of confirming the degree of mRNA expression of a marker gene that can predict whether or when allergic rhinitis occurs, and can be performed by measuring the amount of mRNA. Analysis methods for measuring the mRNA level include, but are not limited to, reverse transcription polymerase chain reaction (RT-PCR), competitive reverse transcription polymerase chain reaction (Competitive RT-PCR), real-time reverse transcription polymerase chain reaction (Real-time RT-PCR), RNase protection assay, Northern blotting, or DNA microarray chip (DNA chip).

[0021] The term "preparation for measuring the expression level of a gene" in this specification refers to a molecule that can be used to detect a marker by confirming the expression level of the mRNA or protein of a marker whose expression level increases or decreases depending on whether or when allergic rhinitis occurs.

[0022] In one embodiment, the agent capable of measuring the gene expression level may comprise one or more selected from the group consisting of primers, probes, and antisense nucleotides that specifically bind to the COL5A2, CDK2AP2, COQ9, or PYM1 genes, but is not limited thereto.

[0023] The term “primer” in this specification refers to any combination of primer pairs consisting of forward and reverse primers that recognize a target gene sequence, and preferably refers to a primer pair that provides analytical results having specificity and sensitivity. The primer refers to a single-stranded oligonucleotide capable of acting as a starting point for template-directed DNA synthesis under suitable conditions (i.e., the presence of four different nucleoside triphosphates, polymerase, temperature, buffer, etc.). The suitable length of the primer varies depending on various factors, such as temperature and the application of the primer, but is typically 15 to 30 nucleotides.

[0024] The sequence of the above primer does not need to be completely complementary to a portion of the template sequence; it is sufficient to have sufficient complementarity within the range where it can hybridize with the template and perform its inherent function. Additionally, the above primer can be prepared with a sequence complementary to the mRNA (i.e., cDNA) sequence of the target gene. The design of such a primer can be easily carried out by a person skilled in the art by referring to the cDNA sequence of the target gene.

[0025] The term "probe" in this specification refers to a substance capable of specifically binding to a target substance to be detected within a biological sample, and means a substance capable of specifically confirming the presence of the target substance within the sample through said binding. Specifically, the probe is a substance commonly used in the art and can be produced in the form of an oligonucleotide probe, a single-strand DNA probe, a double-strand DNA probe, an RNA probe, etc. Specifically, it may be a nucleic acid fragment such as RNA or DNA corresponding to a few bases or as long as several hundred bases, and is labeled so as to confirm the presence or absence and expression level of a specific mRNA.

[0026] The above primer or probe can be chemically synthesized using a phosphoramidite solid support synthesis method or other widely known methods. In addition, the above primer or probe can be modified through various methods known in the art. Examples of such modifications include methylation, capping, substitution with one or more homologues of natural nucleotides, and modification between nucleotides, for example, modification to uncharged linkages (e.g., methyl phosphonate, phosphotriester, phosphoroamidate, carbamate, etc.) or charged linkages (e.g., phosphorothioate, phosphorodithioate, etc.).

[0027] The term "antisense oligonucleotide" in this specification refers to DNA or RNA or derivatives thereof comprising a nucleic acid sequence complementary to a specific mRNA sequence, which acts to inhibit the translation of the mRNA into a protein by binding to the complementary sequence within the mRNA. An antisense oligonucleotide sequence refers to a DNA or RNA sequence that is complementary to the mRNA of the said genes and capable of binding to said mRNA. This can inhibit essential activities for the translation, translocation into the cytoplasm, maturation, or any other overall biological function of said gene mRNA. The length of the antisense oligonucleotide may be 6 to 100 bases, preferably 8 to 60 bases, and more preferably 10 to 40 bases. The antisense oligonucleotide may be synthesized in vitro by conventional methods and administered in vivo, or the antisense oligonucleotide may be synthesized in vivo.

[0028] In one embodiment, the agent for measuring the activity level of the protein may include one or more selected from the group consisting of antibodies, oligopeptides, ligands, PNA (peptide nucleic acid), and aptamers that specifically recognize the protein, but is not limited thereto.

[0029] The term "antibody" in this specification refers to a substance that specifically binds to an antigen to induce an antigen-antibody reaction. The antibody may comprise not only a complete form having two full-length light chains and two full-length heavy chains, but also functional fragments of the antibody molecule. A functional fragment of the antibody molecule refers to a fragment possessing at least an antigen-binding function and may be, for example, Fab, F(ab'), F(ab')2, or Fv. Furthermore, the antibody includes polyclonal antibodies, monoclonal antibodies, and recombinant antibodies. Additionally, the antibody may be easily manufactured using techniques widely known in the art. The polyclonal antibody may be produced from any animal, such as a goat, rabbit, sheep, monkey, horse, pig, cattle, or dog. The above monoclonal antibody can be prepared using the hybridoma method (see Kohler and Milstein (1976) European Journal of Immunology 6:511-519), which is widely known in the art, or phage antibody library technology (see Clackson et al, Nature, 352:624-628, 1991; Marks et al, J. Mol. Biol., 222:58,1-597, 1991).

[0030] The term "PNA (Peptide Nucleic Acid)" in this specification refers to an artificially synthesized polymer similar to DNA or RNA. Specifically, unlike DNA, which has a phosphate-ribose sugar backbone, PNA has a repeating N-(2-aminoethyl)-glycine backbone linked by peptide bonds, which can significantly increase its binding strength and stability to DNA or RNA.

[0031] The term "aptamer" in this specification means a nucleic acid (DNA, RNA) or peptide that can affinityally and specifically bind to a target molecule.

[0032] The method for measuring the activity level of the above protein may be to measure using any one selected from the group consisting of Western blotting, enzyme linked immunosorbent assay (ELISA), radioimmunoassay (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunohistochemistry, immunoprecipitation assay, complement fixation assay, flow cytometry (FACS), and protein chips.

[0034] Another aspect provides a diagnostic kit for allergic rhinitis comprising a composition capable of predicting whether or when allergic rhinitis occurs.

[0035] The above-mentioned allergic rhinitis, measurement of gene expression levels, and measurement of protein activity levels are as described above.

[0036] The above kit may be an RT-PCR (Reverse transcription polymerase chain reaction) kit, a DNA chip kit, an ELISA (Enzymelinked immunosorbent assay) kit, a protein chip kit, a rapid kit, or an MRM (Multiple reaction monitoring) kit, but is not limited thereto. Additionally, the above kit may be configured to include one or more other components, solutions, or devices suitable for the analysis method.

[0037] For example, the RT-PCR kit may include a pair of primers specific to the marker gene, and may also include a test tube or other suitable container, reaction buffer (with varying pH and magnesium concentration), deoxynucleotides (dNTPs), enzymes such as Taq-polymerase and reverse transcriptase, DNAse, RNAse inhibitor DEPC-water, sterile water, etc. The DNA chip kit may include a substrate to which cDNA or oligonucleotides corresponding to a gene or its fragment are attached, and reagents, preparations, enzymes, etc., for producing a fluorescent probe, and the substrate may include cDNA or oligonucleotides corresponding to a control gene or its fragment. The ELISA kit includes an antibody specific to the protein. The antibody is an antibody with high specificity and affinity for the marker protein and minimal cross-reactivity with other proteins, and is a monoclonal antibody, a polyclonal antibody, or a recombinant antibody. Other ELISA kits may include reagents capable of detecting conjugated antibodies, such as labeled secondary antibodies, chromophores, enzymes (e.g., conjugated with antibodies), their substrates, or other substances capable of binding to antibodies. The protein chip kit may be a kit in which one or more antibodies against a marker are arranged at a fixed location on a substrate and immobilized at a high density. The protein chip may be used to isolate proteins from a biological sample, hybridize the isolated proteins with the protein chip to form an antigen-antibody complex, and read the complex to confirm the presence or expression level of the protein. In this specification, the term "antigen-antibody complex" refers to a combination of a corresponding protein antigen in a biological sample and an antibody that recognizes it.The above antigen-antibody complex may be detected using methods known in the art, such as spectroscopic, photochemical, biochemical, immunochemical, electrochemical, spectrophotometric, chemical, and other methods. The rapid kit may include an antibody specific to the protein and / or an antibody specific to a control protein. Additionally, the rapid kit may include other materials capable of detecting the bound antibody, such as a reagent, for example, a nitrocellulose membrane with the specific antibody and secondary antibody immobilized thereon, a membrane bound to an antibody-bound bead, an absorbent pad, and a sample pad.

[0038] In one embodiment, the kit may be an integrated magneto-electrochemical sensor kit, but is not limited thereto. The integrated magneto-electrochemical sensor refers to a system that detects an electrical signal by enzymatic signal amplification using magnetic beads. Specifically, it may refer to a system that attaches a substance capable of detecting a target substance (e.g., an antibody) to magnetic beads and detects an electrical signal by enzymatic signal amplification generated by a chromogenic substance capable of detecting the target substance (e.g., horseradish peroxidase (HRP), alkaline phosphatase (ALP), α-D-galactosidase (α-Gal).

[0039] In one embodiment, the kit may include a computer with a built-in formulation, device, and algorithm for measuring the level of allergic rhinitis gene expression, and may relate to a kit that associates the result of measuring the level of the marker with allergic rhinitis through the algorithm.

[0041] Another aspect provides a method for providing information for predicting whether and when allergic rhinitis occurs, comprising the step of measuring the activity or expression level of COL5A2, CDK2AP2, COQ9, or PYM1 proteins or genes in a biological sample isolated from a target individual.

[0042] The measurement of the level of activity or expression of the above allergic rhinitis, protein, or gene is as described above.

[0043] The term "target subject" in this specification means a subject for whom the occurrence of allergic rhinitis is uncertain.

[0044] In this specification, the term "biological sample" may be a sample, cultured cell, or feces isolated from an individual having or at risk of having an allergic rhinitis-related disease. Specifically, the biological samples include bronchial biopsy, lung biopsy, nasal cavity biopsy, sinus biopsy, sputum, bronchoalveolar lavage fluid (BALF), nasal lavage fluid, nasal swap, nasal aspirate, nasopharyngeal swab, nasopharyngeal aspirate, nasopharyngeal lavage fluid, whole blood, plasma, serum, tears, mucus, breath, urine, semen, saliva, peritoneal washings, ascites, cystic fluid, meningeal fluid, amniotic fluid, glandular fluid, and lymph. It may be fluid), pleural fluid, nipple aspirate, synovial fluid, joint aspirate, organ secretions, cell, cell extract, cerebrospinal fluid, or a combination thereof, and preferably may be selected from the group consisting of blood, serum and plasma, but is not limited thereto.

[0045] In one embodiment, the method for providing information for predicting whether and when allergic rhinitis occurs may further include a step of determining that the subject is an early-onset allergic rhinitis group (G2) if the activity or expression level of COL5A2 is higher than that of a normal group (G1) and the activity or expression level of COQ9 is lower than that of a control group.

[0046] In addition, in one embodiment, the method for providing information for predicting whether and when allergic rhinitis occurs may further include a step of determining that the subject is an allergic rhinitis late-onset group (G3) if the activity or expression level of COL5A2 is higher than that of a normal group (G1) and the activity or expression level of COQ9 is higher than that of a control group.

[0047] In addition, in one embodiment, the method may further include a step of determining that the subject is a normal group or an early-onset allergic rhinitis group (G2) if the activity or expression level of COL5A2 is lower than that of a normal group (G1) and the activity or expression level of CDK2AP2 is lower than that of a control group.

[0048] In addition, in one embodiment, the method may further include a step of determining that the subject is a normal group (G1) or an early-onset allergic rhinitis group (G2) if the activity or expression level of COL5A2 is lower than that of the normal group (G1), the activity or expression level of CDK2AP2 is higher than that of the normal group, and the activity or expression level of PYM1 is lower than that of the normal group (G1).

[0049] The above normal group (G1), early-onset allergic rhinitis group (G2), and late-onset allergic rhinitis group (G3) are as described above.

[0050] In this specification, the term “increase in level of gene expression” means that the concentration of mRNA in a sample of an individual has increased significantly to a measurable degree compared to a normal group (G1), and may mean, for example, an increase of 1.1 times or more. For example, it may mean an increase of 1.1 to 2.5 times, specifically, 1.1 times, 1.2 times, 1.3 times, 1.4 times, 1.5 times, 1.6 times, 1.7 times, 1.8 times, 1.9 times, 2.0 times, 2.1 times, 2.2 times, 2.3 times, 2.4 times, or 2.5 times.

[0051] In this specification, the term “reduction in the level of gene expression” means that the concentration of mRNA in a sample of an individual is significantly reduced to a measurable level compared to a normal group (G1), and may mean, for example, a reduction of 0.1 times or less. For example, it may mean a reduction of 0.1 times to 0.9 times, specifically, a reduction of 0.1 times, 0.2 times, 0.3 times, 0.4 times, 0.5 times, 0.6 times, 0.7 times, 0.8 times, or 0.9 times.

[0052] In this specification, the term “increase in level of protein activity” means that the activity of the protein in the sample of an individual has increased significantly enough to be measurable compared to the normal group (G1), and may mean, for example, an increase of 1.1 times or more. For example, it may mean an increase of 1.1 times to 2.5 times, specifically, 1.1 times, 1.2 times, 1.3 times, 1.4 times, 1.5 times, 1.6 times, 1.7 times, 1.8 times, 1.9 times, 2.0 times, 2.1 times, 2.2 times, 2.3 times, 2.4 times, or 2.5 times.

[0053] In this specification, the term “reduction in the level of protein activity” means that the activity of the protein in the sample of an individual is significantly reduced to a measurable level compared to the normal group (G1), and may mean, for example, a reduction of 0.1 times or less. For example, it may mean a reduction of 0.1 times to 0.9 times, specifically, a reduction of 0.1 times, 0.2 times, 0.3 times, 0.4 times, 0.5 times, 0.6 times, 0.7 times, 0.8 times, or 0.9 times.

[0054] In one embodiment, the method may additionally use clinical information other than the marker of the individual in addition to the results of measuring and analyzing the level of the gene to provide information for predicting whether or when allergic rhinitis occurs. Such clinical information includes, for example, the patient's age, gender, weight, dietary habits, body mass, ultrasound, computed tomography (CT), magnetic resonance imaging (MRI), angiography, etc.

[0055] Another aspect provides a screening method for an allergic rhinitis prevention or treatment, comprising the steps of: treating a separated biological sample with a candidate substance; and selecting the candidate substance as an allergic rhinitis prevention or treatment if the expression level of the COL5A2, CDK2AP2, COQ9, or PYM1 gene in the biological sample increases or decreases compared to a control group not treated with the candidate substance.

[0056] The above-mentioned allergic rhinitis, measurement of gene expression levels, and measurement of protein activity levels are as described above.

[0057] In this specification, the term "candidate substance" may be an individual nucleic acid, protein, other extract or natural product, or compound, etc., that is presumed to have the potential to prevent or treat allergic rhinitis according to a conventional selection method or is selected randomly. In this specification, the term "control group" refers to isolated cells, isolated tissues, or animals other than humans that have not been treated with the candidate substance, and which are isolated cells, isolated tissues, or animals other than humans that are in parallel with the group treated with the candidate substance. Effects of the invention

[0058] According to a composition, kit, and diagnostic method for allergic rhinitis based on a specific pattern, COL5A2, CDK2AP2, COQ9, and PYM1 proteins or genes showing differences in expression in blood transcriptome analysis of an early-onset allergic rhinitis group, a late-onset allergic rhinitis group, and a normal group can be identified, and the occurrence or timing of allergic rhinitis can be predicted through analysis of a Decision tree prediction model using the same. Brief explanation of the drawing

[0059] Figure 1 shows a group-by-group comparative analysis performed on the entire blood transcriptome using numeric regression. P This is a schematic diagram showing the results of identifying 181, 687, 349, and 463 genes with highly significant differences in expression of < 0.001 for each group comparison, and 14 candidate genes selected for Decision tree prediction model analysis. Figure 2 is a graph showing the results of a prediction model analysis on whether or when allergic rhinitis occurs using a Decision tree program, utilizing genes (COL5A2, CDK2AP2, COQ9, PYM1) that showed differences in expression in the blood transcriptome analysis of a normal group (G1), an early-onset allergic rhinitis group (G2), and a late-onset allergic rhinitis group (G3). Specific details for implementing the invention

[0060] The following examples will be explained in more detail. However, these examples are for illustrative purposes only and the scope of the present invention is not limited to these examples.

[0061] Example 1. Characteristics of the subject

[0062] The demographic and clinical characteristics of the subjects participating in the experiment are as follows.

[0063] Specifically, after a detailed examination by a respiratory allergist, the examination details were recorded in a symptom sheet. The presence of an allergic disease was defined through a physical examination and questions regarding symptoms, diagnosis, and treatment related to the disease over the past 12 months. Allergic rhinitis symptoms were defined as having been diagnosed with allergic rhinitis within the past 12 months and having experienced at least two symptoms (clear runny nose, nasal obstruction, sneezing, or itching).

[0064] Twelve people were selected for the normal group (G1; Healthy controls), 12 for the early onset allergic rhinitis group (G2), and 12 for the late onset allergic rhinitis group (G3).

[0066] Example 2. Confirmation of allergy symptoms for each subject group

[0067] To identify allergic symptoms in the early-onset group (G2), late-onset group (G3), and normal group (G1) of pediatric allergic rhinitis, tests were conducted on the allergens to which they were most sensitive.

[0068] Specifically, European and American house dust mite antigens, two types of mold antigens (Altenaria, Aspergillus), three types of pollen antigens (Birch, Alder, Oak), dog hair antigens, cat hair antigens, mugwort antigens, Japanese knotweed antigens, weed antigens, cockroach antigens, etc. were tested.

[0070] Example 3. Confirmation of changes in expression of differentially expressed genes (DEGs)

[0071] 3.1 Identification of Differentially Expressed Genes (DEGs)

[0072] To identify genes showing differences in expression during blood transcriptome analysis of the early-onset (G2), late-onset (G3), and normal (G1) groups of pediatric allergic rhinitis, the mRNA levels of genes for each group were measured. Specifically, cDNA was synthesized using the GeneChip WT Amplification Kit (Affymetrix, Santa Clara, CA) from total RNA extracted using the PAXgene Blood RNA kit (PreAnalytiX) from blood tissues stored in dedicated PAXgene Blood RNA Tubes (PreAnalytiX, Hombrechtikon, Switzerland) containing RNA preservatives, and then hybridized with the GeneChip® Human Gene 2.0 ST Array (Affymetrix) according to the manufacturer's instructions. After scanning the array using the GCS3000 scanner (Affymetrix), analyses such as data quality checks, background correction, and purification were performed using the Affymetrix GeneChip Command Console software.

[0073] For the entire transcriptome, group-specific comparative analysis using numeric regression (1. early onset group vs. late onset group, 2. normal group vs. total rhinitis group, 3. normal group vs. early onset group, 4. normal group vs. late onset group) was performed. P We identified 181, 687, 349, and 463 genes, respectively, that showed a significant difference in expression of < 0.001, and the results are shown in Figure 1.

[0074] As shown in Figure 1, 14 important candidate genes that overlapped in comparisons with other groups including the normal group were selected while distinguishing between the early-onset group and the late-onset group.

[0075] For 14 selected candidate genes, a decision tree based on gene expression between pediatric allergic rhinitis phenotypes was performed using the conditional inference tree (Ctree) function of R language 3.6.0 (http: / / www.r-project.org).

[0076] Based on the above results, the COL5A2, CDK2AP2, COQ9, and PYM1 genes, which showed differences in expression in the blood transcriptome analysis of the early-onset, late-onset, and normal groups of pediatric allergic rhinitis, were selected.

[0077] 3.2 Confirmation of changes in COL5A2 expression

[0078] Blood transcriptomes of the early-onset allergic rhinitis group, late-onset allergic rhinitis group, and normal group were analyzed according to the method of Example 3.1, and changes in COL5A2 gene expression were confirmed. Specifically, for COL5A2 mentioned in the present invention, gene expression was measured using 31 probes used in the GeneChip® Human Gene 2.0 ST Array. The information on the probes for measuring COL5A2 gene expression is shown in Table 1.

[0079] Probe_id Probe_sequence Sequence number 2444768 AAGGACCGTCCATCGAATTTCTCAC 1 417593 CCGTGCGAACGGGTAGTATCTAGAA 2 2073982 AGGCCATATAGCAAGAAGTTCTGTG 3 220082 CGTTTGGGTAGGTCACATGGTGCAT 4 2523823 ATGTCGACGAGAACCCCTATAGTAC 5 535891 CGGGTCAACCAGGAAGTCCATTTCT 6 799696 GAGGTTACCAGGACATCCCCTTGGA 7 2391235 TTGCTGGCCCGTCTCTCCTTAACAA 8 567560 GAGGTCCAGAAGCACCCCTGGGACC 9 2460217 TAAAGGACCAAGACGCCCGTCTCAA 10 2468562 ACAAGGACCTGATTTTCCACCAGCT 11 2272560 CAAGAGCTCCACTTTTACCCGGTTG 12 2593136 GGAGCTCCAAATCAACCGGGAGGAC 13 2557752 CAGGCCGTCCAGGATGACCTCTTTT 14 2200019 GTCCAGAAGTTCCATACGGCCCTCT 15 1072589 TCGTCAACCGGGCAATCCTGGATCT 16 2034958 GAGGTCCGAGGTATCCTTAGTCTCC 17 1208455 CCACGAGGACCGTTAGCACCAAAAG 18 2366563 CGGTGTACCATAAGTCCCAGGCTAT 19 771243 AGAGGTCCTGGAGTCCCATCGTGAC 20 2305857 CAGGTCAACCGAGAGGTCCAGAAGG 21 451440 CAGGAGAACCCTATGGTCCGAGAAG 22 2258053 GGGTGACCAGGTTACCCACGGTACC 23 56652 GTGTTTCCAGAACTTCCGGGATTTC 24 2174682 CCCGAGGACCAGAAGGTCCAGACTT 25 794977 CAGGCTAACCAAGTGCACCTGGTCT 26 753002 AACCCTCAGTTCATCCTGATTACGG 27 2159269 CCCTCTTGGTCCTAATCACGGACAA 28 1914577 ACCTCGGTAAGAGACACTGTTCTAT 29 988701 TTTGACCCGCCTTCGTTCTGGAGAG 30 33353 AACGAACCGTCTGTGACCTACCAAT 31

[0080] 3.3 Confirmation of changes in CDK2AP2 expression

[0081] Blood transcriptomes of the early-onset allergic rhinitis group, late-onset allergic rhinitis group, and normal group were analyzed according to the method of Example 3.1, and changes in the expression of the CDK2AP2 gene were confirmed. Specifically, for CDK2AP2 mentioned in the present invention, gene expression was measured using 19 probes used in the GeneChip® Human Gene 2.0 ST Array. The information on the probes for measuring CDK2AP2 gene expression is shown in Table 2.

[0082] Probe_id Probe_sequence Sequence number 2204086 CCGGAGTCGCAGACCTGGATAGGCC 32 219279 CAGACCTGGATAGGCCGGTGACGTC 33 270309 GATCAGTCTCTCACGGACCGTCTCT 34 177874 GAGTCCTTTGGCTGTGACCTGATAG 35 1449890 CGGATCTCCTACAGGTCATAATGAT 36 2077190 AAGTTGTCCCGTTGGAACTCCTAAT 37 2508914 GACAGTCAGTATCTCCTCTCACCCGT 38 45310 CTGGCGGAAGGTACCGATGCACGT 39 462661 CCGCGAGGAAAGTCTGGCGACAAAT 40 1911838 ATTGCTGAAACCTGGCGGGAAGGTAC 41 1334166 GGGCCAGGACCCGAACCTTTTCGAC 42 1278732 AGTCGTACCTTACTTCGTAGGAAAC 43 2037878 ACTGGAGAATGAGCCGGACAACGA 44 1247259 GGACTCAAGACGTTATGACCCGGAC 45 654701 GAGCTTCTCTTCTGCCGGACTCAAG 46 1304507 AGCTCCAGAGCTTCTCTTCTGCGG 47 2543333 TAGAAGCGCGTGGTTAGCCCTCACT 48 108379 GTCCAGAGCCGGTTATTCCTCCGAG 49 1977478 AGGCCGGTCTGGAGATAAATGGTCC 50

[0083] 3.4

[0084] Blood transcriptomes of the early-onset allergic rhinitis group, late-onset allergic rhinitis group, and normal group were analyzed according to the method of Example 3.1, and changes in the expression of the COQ9 gene were confirmed. Specifically, for COQ9 mentioned in the present invention, gene expression was measured using 26 probes used in the GeneChip® Human Gene 2.0 ST Array. The information on the probes for measuring COQ9 gene expression is shown in Table 3.

[0085] Probe_id Probe_sequence குற்றியு 1573620 GCACCCGCTGCACGGGCGAAGGTTT 51 865902 ACCAGGGCATCGATGGCCAGCGCAG 52 2079033 CTCCGAGGACGTCGACGCTACGGAC 53 9223 CCTCCGAGGACGTCGACGTACGGA 54 175807 GCGCACGGAAGGTACGAAGTCGAAGCGACA 55 478285 CCCGTCTTTTTGGACTAGGTCTCAG 56 1685493 CCCGATTCCAGAAGTCTACTCGTCT 57 985210 GTCGTCGGTCGTACAAGCCCTTCCT 58 2406177 GGGCCGAGTGTGCACACGATCTTCT 59 1907010 GACGTAAAACACTGGGTCACGTTAT 60 216087 TTCTCCTTCTGTCTGGTCAAGGACT 61 91548 GGTCTGACTCTTACGACTAGGGTAT 62 1019845 ACTAGGGTATGTAACTCGTGACCG 63 1221427 TGTACACCGTAATGCGACCCCTGGT 64 657228 CTGTACACCGTAATGCGACCCCTGG 65 644987 ACCGTAATGCGACCCCTGGTCAGGT 66 458013 GACCACTACTACGTCCTGAGGAGAG 67 2289253 CTGTGAACCGCGAAGGACCTTTTTGG 68 712505 AAATTGACCATGTGGGCGGCTCGGT 69 989927 TTCCTGAGTACCACGTCGTCACTG 70 1280131 CATTTCAGGTGTCCTCTCCGTGACC 71 1143639 CTGTCCAGATTTGGTCGCAGCCACT 72 1075284 ACTGTCCAGATTTGGTCGCAGCCAC 73 2310417 CCACGGTAGGTGTATTGGACCACAA 74 698911 ACCCGTGGAACTAGTACAGAATTGG 75 1451625 ACCCGACGAAGAAGTCAAGGATTAT 76

[0086] 3.5 PYM1 발현 변화 확인

[0087] Blood transcriptomes of the early-onset allergic rhinitis group, late-onset allergic rhinitis group, and normal group were analyzed according to the method of Example 3.1, and changes in the expression of the PYM1 gene were confirmed. Specifically, for PYM1 mentioned in the present invention, gene expression was measured using 29 probes used in the GeneChip® Human Gene 2.0 ST Array. The information on the probes for measuring PYM1 gene expression is shown in Table 4.

[0088] Probe_id Probe_sequence 서열번호 568195 GGGTCCATGGAAACGACGACTAAAC 77 1966937 GAACCAAGGACAAGCGCCACAAATA 78 172500 ACCTGACGTCTTGTTTGGCACCCCG 79 1096826 CGGTGGTGACTCTTTCGGTTCTTCT 80 630419 CCGGAGAGGTTCTGTCGGTTTGCAT 81 2097704 GTTGTGTCGCTGGACTGCCCTGGAC 82 279795 GTTCATATAGCGCAGTTGTGTCGCT 83 465988 CTTCGAAGACGAGACGGAAAGTAAT 84 2376114 ACACTAATGGTACACGGGATGGCAT 85 1355940 AAGGTGTGTCCTACCGGTAAGTTCT 86 233746 AAGAAAAACTGAAGTGGGAACCTAT 87 611981 AACTGAAGTGGGAACCTATGGTAGT 88 1179406 AAACTGAAGTGGGAACCTATGGTAG 89 1034550 TACCGCTGAGGGATACAATGACTGC 90 1525242 AGGGATACAATGACTGCTCTGGCCG 91 864858 ACCGCTGAGGGATACAATGACTGCT 92 1825593 GAGTGCGGCCTTCACCAAACGCAAA 93 267732 CCGAACGATGCTAGCCCTGAGGTGA 94 582715 CCACGTGAGAGGGAAATTCTGATTC 95 1017429 GACAAAATGTCCGTCGTTCGCGAGT 96 56852 CTGACACGGAGCCTTGAAGAGAACC 97 88994 TTGGCAGGGATCTTTGTGAACACGC 98 2368692 GTTCGGCGAGACTCTAGACCCACGT 99 2160226 CGCTCTTCGTTGATCCCGGGAGTAG 100 685704 CCGCTCTTCGTTGATCCCGGGAGTA 101 2589482 ACCCGCTCTTCGTTGATCCCGGGAG 102 1052976 GGATAGTTCCACTTTTCACTCTAAG 103 1208394 CCTTCCGAGTAAAGAGGATAGTTCC 104 8300 AGAAGGAGTCCTTCCGAGTAAAGAG 105

[0089] 실시예 4. 조건부 추론 트리분석 및 통계분석

[0090] The presence of genes according to the timing of allergic rhinitis onset was analyzed using the conditional inference tree function. Specifically, recursive splitting was used to determine the gene expression cutoff between atopic dermatitis phenotypes using the conditional inference tree (Ctree) function of the R package. The model parameters were set to minsplit = 2, minbucket = 1, mincriterion = 0.95, testtype = "Teststatistic", and maxdepth = 4. For Ctree analysis, genes with a value less than 0.05 at each branch point with the smallest P- were selected as internal nodes. The Ctree model was constructed using the "PARTY" program developed in R language 3.6.0 (http: / / www.r-project.org). The results of the gene expression analysis according to the timing of allergic rhinitis onset using the above conditional inference tree function are shown in Figure 2.

[0091] Figure 2 is a graph showing the results of a prediction model analysis on whether or when allergic rhinitis occurs using a Decision tree program, utilizing genes (COL5A2, CDK2AP2, COQ9, PYM1) that showed differences in expression in the blood transcriptome analysis of a normal group (G1), an early-onset allergic rhinitis group (G2), and a late-onset allergic rhinitis group (G3).

[0092] The above results indicate that a combination of four candidate genes (COL5A2, CDK2AP2, COQ9, PYM1) that showed differences in expression in blood transcriptome analysis of a normal group (G1), an early-onset allergic rhinitis group (G2), and a late-onset allergic rhinitis group (G3) is a useful biomarker that can predict whether or when allergic rhinitis occurs.

Claims

Claim 1 A diagnostic composition capable of predicting whether or when allergic rhinitis occurs, comprising a preparation capable of measuring the activity or expression level of one or more proteins or genes selected from the group consisting of COL5A2, CDK2AP2, COQ9, and PYM1. Claim 2 The composition of claim 1, wherein the agent capable of measuring the gene expression level comprises one or more selected from the group consisting of primers, probes, and antisense nucleotides that specifically bind to the COL5A2, CDK2AP2, COQ9, or PYM1 genes. Claim 3 A composition according to claim 1, wherein the agent for measuring the activity level of the protein comprises one or more selected from the group consisting of antibodies that specifically recognize the protein, oligopeptides, ligands, PNA (peptide nucleic acid), and aptamers. Claim 4 A diagnostic kit capable of predicting whether or when allergic rhinitis occurs, comprising any one of the compositions of claims 1 to 3. Claim 5 A method for providing information for predicting the occurrence and timing of allergic rhinitis, comprising the step of measuring the activity or expression level of COL5A2, CDK2AP2, COQ9, or PYM1 proteins or genes in a biological sample isolated from a target individual. Claim 6 A method according to claim 5, further comprising the step of determining that the subject is an early-onset allergic rhinitis group if the activity or expression level of COL5A2 is higher than that of a normal group and the activity or expression level of COQ9 is lower than that of a control group. Claim 7 A method according to claim 5, further comprising the step of determining that the subject is a late-onset allergic rhinitis group if the activity or expression level of COL5A2 is higher than that of a normal group and the activity or expression level of COQ9 is higher than that of a control group. Claim 8 A method according to claim 5, further comprising the step of determining that the subject is a normal group or an early-onset allergic rhinitis group if the activity or expression level of COL5A2 is lower than that of a normal group and the activity or expression level of CDK2AP2 is lower than that of a control group. Claim 9 A method according to claim 5, further comprising the step of determining that the subject is a normal group or an early-onset allergic rhinitis group if the activity or expression level of COL5A2 is lower than that of a normal group, the activity or expression level of CDK2AP2 is higher than that of a normal group, and the activity or expression level of PYM1 is lower than that of a normal group. Claim 10 A method according to claim 5, further comprising the step of determining that the subject is a late-onset allergic rhinitis group if the activity or expression level of COL5A2 is lower than that of the normal group, the activity or expression level of CDK2AP2 is higher than that of the normal group, and the activity or expression level of PYM1 is higher than that of the normal group. Claim 11 The method of claim 5, wherein the biological sample is selected from the group consisting of blood, serum and plasma. Claim 12 The method of claim 5, wherein the step of measuring the gene expression level is to use a method selected from the group consisting of reverse transcription polymerase chain reaction (RT-PCR), competitive reverse transcription polymerase chain reaction (Competitive RT-PCR), real-time reverse transcription polymerase chain reaction (Real-time RT-PCR), RNase protection assay (RPA), Northern blotting, and DNA microarray chip. Claim 13 The method of claim 5, wherein the step of measuring the protein activity level is to use a method selected from the group consisting of Western blotting, enzyme linked immunosorbent assay (ELISA), radioimmunodiffusion (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunohistochemistry, immunoprecipitation, complement fixation assay, flow cytometry (FACS), and protein chips. Claim 14 delete

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