KR20220109049A - Biomarker for diagnosis or prognosis of ischemic disease - Google Patents

Abstract

The present invention relates to a biomarker for diagnosing ischemic disease or diagnosing prognosis of ischemic disease, a kit for diagnosing ischemic disease or diagnosing prognosis of ischemic disease by using the same, and a method for diagnosing ischemic disease or diagnosing prognosis of ischemic disease by using the same. More specifically, the present invention relates to at least one biomarker selected from a group of consisting of ADGRG3, AATK, ALOX5, CD177, CREB5, ICAM3, LSM3, and S100A6, a kit for diagnosing ischemic disease or diagnosing prognosis of ischemic disease, which includes a material bound to the biomarker, and a method for providing information on diagnosis of ischemic disease or diagnosis of prognosis of ischemic disease by measuring an expression level of the biomarker in a subject.

Description

Biomarker for diagnosis or prognosis of ischemic disease

The present invention relates to a biomarker for ischemic disease diagnosis or prognosis diagnosis, a kit for ischemic disease diagnosis or prognosis diagnosis using the biomarker, and a method for diagnosing ischemic disease or prognosis using the biomarker.

The present invention is a research carried out with the support of the National Research Foundation of Korea Biomedical Technology Development Project with funding from the Ministry of Science and ICT (NRF-2016M3A9B6900776).

Stroke is a neurological symptom that occurs when a blood vessel supplying blood to the brain is blocked or ruptured and the brain is damaged in that part. Ischemic stroke occurs when the brain tissue becomes ischemic due to a decrease or blockage of blood supply to the brain tissue. is known to occupy. When an ischemic stroke occurs, blood flow is temporarily cut off and cells cannot obtain oxygen and nutrients. ) process can minimize nerve damage.

An ischemic stroke is a disease in which brain nerve cells are damaged due to insufficient blood flow to the brain tissue due to blockage of cerebral blood vessels. can However, recently, thrombectomy in cerebral arteries has developed rapidly, and if a successful thrombectomy is performed by promptly visiting a hospital after the onset of symptoms of cerebral infarction, the sequelae of cerebral infarction can be reduced to a minimum.

Early detection of stroke through rapid diagnosis is the best defense to reduce the damage. Stroke diagnosis methods are divided into preventive diagnosis and post-onset diagnosis. The preventive diagnosis method is a blood test method, and the post-onset diagnosis method is first of all due to the extremely conflicting treatment mechanisms of ischemic stroke and hemorrhagic stroke. For the etiological diagnosis of cognitive infarct cognition, computer aided tomography (CT) or magnetic resonance imaging (MRI) is performed. In addition, cerebral blood flow measurement using radioactive isotopes or ultrasound, brain Angiography, etc. CT is the most helpful for the differential diagnosis of ischemic stroke and hemorrhagic stroke, and MRI is evaluated to be excellent for determining the degree of ischemic stroke and vascular status in the acute phase. However, there are disadvantages such as high cost due to the use of expensive equipment and limitation of diagnostic sites.

In the conventional stroke diagnosis method, there is no preventive diagnosis method other than blood test, and the diagnosis method after the onset is diagnosis using radioactive isotopes harmful to the human body. Not only that, but it also creates a problem that it takes a considerable period of time for the diagnosis results to come out.

However, since brain tissue does not regenerate once damaged and the sequelae are serious once it occurs, the importance of diagnosis for treatment decisions after the onset is not only in terms of reducing socioeconomic loss costs, but also in reducing mortality and national disease management. The net functional value is very high. Therefore, there is an urgent need to develop a new stroke marker that effectively enables early diagnosis of stroke.

In contrast, the present inventors made efforts to discover new biomarkers of ischemic diseases including cerebral infarction. As a result, gene and protein expression changes in human and monkey cerebral infarction disease models were more than doubled compared to normal controls, and in particular, genes and By discovering eight new biomarkers that significantly change protein expression, the present invention was completed.

Republic of Korea Patent 10-2170826 Republic of Korea Patent 10-2140581 Korean Patent Registration 10-1478426 Republic of Korea Patent Publication 10-2011-0133133

An object of the present invention is to provide a biomarker for diagnosing ischemic disease or prognosis, which includes an adhesion G protein-coupled receptor G3 (ADGRG3) protein or a gene encoding the protein.

Another object of the present invention is the ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), Any one protein selected from the group consisting of ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6), or any one of the genes encoding the protein It is to provide a biomarker for diagnosis of ischemic disease or prognosis further comprising one or more.

Another object of the present invention is to provide a composition for diagnosing ischemic disease or prognosis comprising a substance specifically binding to the biomarker according to the present invention.

Another object of the present invention is to provide a kit or biochip for diagnosing ischemic disease or prognosis, which includes a substance specifically binding to the biomarker according to the present invention.

Another object of the present invention is to provide a method for providing information on diagnosis of ischemic disease or prognosis of an individual by confirming the expression level of the biomarker according to the present invention in a biological sample isolated from an individual.

Another object of the present invention is to provide a method for screening a drug for preventing or treating ischemic disease by confirming the expression level of the biomarker according to the present invention in a biological sample treated with a candidate drug for preventing or treating ischemic disease.

In order to achieve the above object, the present invention is an antibody or aptamer that specifically binds to the ADGRG3 (adhesion G protein-coupled receptor G3) protein represented by the amino acid sequence of SEQ ID NO: 1, or a gene encoding the protein Provided is a kit for diagnosing ischemic disease or prognosis comprising a primer, a probe, or an antisense nucleotide that binds positively.

In addition, the present invention is ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 ( Intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) antibody or aptamer that specifically binds to any one protein selected from the group consisting of, or Provided is a kit for diagnosing ischemic disease or prognosis comprising a primer, a probe, or an antisense nucleotide that specifically binds to a gene encoding the protein.

In addition, the present invention provides an antibody or aptamer that specifically binds to the ADGRG3 (adhesion G protein-coupled receptor G3) protein represented by the amino acid sequence of SEQ ID NO: 1, or a primer that specifically binds to a gene encoding the protein , to provide a biochip for diagnosing ischemic disease or prognosis comprising a probe or antisense nucleotide.

In addition, the present invention is ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 ( Intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) antibody or aptamer that specifically binds to any one protein selected from the group consisting of, or Provided is a biochip for diagnosing ischemic disease or prognosis comprising a primer, a probe, or an antisense nucleotide that specifically binds to a gene encoding the protein.

In addition, the present invention provides an antibody or aptamer that specifically binds to the ADGRG3 (adhesion G protein-coupled receptor G3) protein represented by the amino acid sequence of SEQ ID NO: 1, or a primer that specifically binds to a gene encoding the protein , to provide a composition for diagnosing ischemic disease or prognosis comprising a probe or antisense nucleotide.

In addition, the present invention is ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 ( Intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) antibody or aptamer that specifically binds to any one protein selected from the group consisting of, or Provided is a composition for diagnosing ischemic disease or prognosis comprising a primer, a probe, or an antisense nucleotide that specifically binds to a gene encoding the protein.

In addition, the present invention comprises the steps of measuring the expression level of the ADGRG3 (adhesion G protein-coupled receptor G3) protein, or a gene encoding the protein, represented by the amino acid sequence of SEQ ID NO: 1 in a biological sample isolated from a subject of interest It provides an information providing method for diagnosing ischemic disease, including diagnosis or prognosis.

In addition, the present invention provides ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP) in a biological sample isolated from a subject of interest. responsive element binding protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) any one protein selected from the group consisting of, or the above It provides an information providing method for diagnosing or prognosing an ischemic disease, comprising measuring the expression level of a gene encoding a protein.

In addition, the present invention comprises the steps of treating a candidate substance expected to prevent or treat an ischemic disease in the isolated biological sample; and measuring the expression level of an adhesion G protein-coupled receptor G3 (ADGRG3) protein represented by the amino acid sequence of SEQ ID NO: 1, or a gene encoding the protein, in the biological sample treated with the candidate substance; A method for screening a drug for preventing or treating ischemic disease is provided.

In addition, the present invention comprises the steps of treating a candidate substance expected to prevent or treat ischemic disease in the isolated biological sample; And, in the biological sample treated with the candidate substance, ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding) protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) any one protein selected from the group consisting of, or encoding the protein It provides a screening method of a drug for the prevention or treatment of ischemic disease, comprising the step of measuring the expression level of the gene.

In the biomarkers ADGRG3, AATK, ALOX5, CD177, CREB5, ICAM3, LSM3 and S100A6 according to the present invention, gene and protein expression changes were more than doubled in the patient cerebral infarction model and the primate cerebral infarction induction model compared to the normal control group. By showing significant changes in gene and protein expression changes, the kit using the biomarker can be usefully used for diagnosing or prognosing cerebral infarction in humans and animals, or screening drugs for treating cerebral infarction.

1 is a diagram showing a change in the expression of a biomarker according to the present invention for each cerebral infarction period.
2 is a diagram showing that the expression of the biomarker according to the present invention is increased in the cerebral infarct group as a result of Dot blot analysis.
3 is a diagram showing that the expression of the biomarker according to the present invention is increased in the cerebral infarction group as a result of dot blot analysis (long-term exposure).

Hereinafter, the present invention will be described in detail.

The present invention provides an adhesion G protein-coupled receptor G3 (ADGRG3) protein or a biomarker for ischemic disease diagnosis or prognosis diagnosis including a gene encoding the protein.

In addition, the present invention is ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 ( intercellular adhesion molecule 3), any one protein selected from the group consisting of LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6), or one or two genes encoding the protein Provided is a biomarker for diagnosis of ischemic disease including abnormality or prognosis.

The adhesion G protein-coupled receptor G3 (ADGRG3) is represented by the amino acid sequence of SEQ ID NO: 1, apoptosis associated tyrosine kinase (AATK) is represented by the amino acid sequence of SEQ ID NO: 2, and arachidonate 5-lipoxygenase (ALOX5) is represented by SEQ ID NO: 3, CD177 (CD177 molecule) is represented by the amino acid sequence of SEQ ID NO: 4, CREB5 (cAMP responsive element binding protein 5) is represented by the amino acid sequence of SEQ ID NO: 5, ICAM3 (intercellular adhesion molecule 3) ) is represented by the amino acid sequence of SEQ ID NO: 6, LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) is represented by the amino acid sequence of SEQ ID NO: 7, S100A6 (S100 calcium binding protein A6) of SEQ ID NO: 8 It may be one represented by an amino acid sequence.

The 'diagnosis' means identifying the presence or characteristics of a pathological condition. For the purposes of the present invention, the diagnosis may include all of the onset of the ischemic disease, the likelihood of the onset, the onset time, and/or the degree of progression.

The 'ischemic disease' includes cerebral infarction, cerebral ischemia, cerebral hypoxia, ischemic heart disease, heart failure, myocardial ischemia, ischemic renal failure, skeletal muscle ischemia, intestinal ischemia, sleep apnea, hypoxia during sleep, focal perfusion defect, retinal ischemia, neonatal hypoxia, fetal hypoxia. , may be any one selected from the group consisting of thrombosis, altitude sickness, carbon monoxide poisoning, sudden infant death syndrome, and congenital circulatory disorders, but may include all other ischemic diseases known in the art.

The agent for measuring the protein expression level of the biomarker is not particularly limited, but for example, an antibody, oligopeptide, ligand, PNA (peptide nucleic acid) and aptamer that specifically binds to the protein. It may include one or more selected from.

The antibody refers to a substance that specifically binds to an antigen and causes an antigen-antibody reaction. For the purposes of the present invention, an antibody refers to an antibody that specifically binds to the biomarker protein. Antibodies of the present invention include polyclonal antibodies, monoclonal antibodies and recombinant antibodies. The antibody can be readily prepared using techniques well known in the art.

The PNA (Peptide Nucleic Acid) refers to an artificially synthesized, DNA or RNA-like polymer, and DNA has a phosphate-ribose sugar backbone, whereas PNA is repeated N- (2-) linked by a peptide bond. It has an aminoethyl)-glycine backbone, which greatly increases binding strength and stability to DNA or RNA, and is used in molecular biology, diagnostic analysis, and antisense therapy. PNA is described in Nielsen PE, Egholm M, Berg RH, Buchardt O (December 1991). "Sequence-selective recognition of DNA by strand displacement with a thymine-substituted polyamide". Science 254 (5037): 1497-1500.

The aptamer is an oligonucleic acid or a peptide molecule, and the general description of the aptamer is described in Bock LC et al., Nature 355(6360):5646(1992); Hoppe-Seyler F, Butz K "Peptide aptamers: powerful new tools for molecular medicine". J Mol Med. 78(8):42630(2000); Cohen BA, Colas P, Brent R. "An artificial cellcycle inhibitor isolated from a combinatorial library". Proc Natl Acad Sci USA. 95(24): 142727 (1998).

The agent for measuring the expression level of the gene encoding the biomarker protein may include at least one selected from the group consisting of primers, probes and antisense nucleotides that specifically bind to the gene encoding the biomarker protein.

The primer is a fragment recognizing a target gene sequence, and includes a pair of forward and reverse primers, but is preferably a primer pair that provides analysis results with specificity and sensitivity. When the nucleic acid sequence of the primer is a sequence that is inconsistent with a non-target sequence present in the sample, and thus amplifies only the target gene sequence containing the complementary primer binding site and does not cause non-specific amplification, high specificity can be conferred. .

The probe means a substance capable of specifically binding to a target substance to be detected in the sample, and means a substance capable of specifically confirming the presence of the target substance in the sample through the binding. The type of probe is not limited as a material commonly used in the art, but preferably PNA (peptide nucleic acid), LNA (locked nucleic acid), peptide, polypeptide, protein, RNA or DNA. The probe may be derived from, or similar to, a biological material as a biomaterial or manufactured in vitro, and may be, for example, enzymes, proteins, antibodies, microorganisms, animal and plant cells and organs, neurons, DNA, and RNA. , DNA includes cDNA, genomic DNA, and oligonucleotides, RNA includes genomic RNA, mRNA, and oligonucleotides, and examples of proteins include antibodies, antigens, enzymes, peptides, and the like.

The LNA (Locked nucleic acids) means a nucleic acid analog including a 2'-O, 4'-C methylene bridge [J Weiler, J Hunziker and J Hall Gene Therapy (2006) 13, 496.502]. LNA nucleosides include common nucleic acid bases in DNA and RNA, and can form base pairs according to Watson-Crick base pairing rules. However, due to the 'locking' of the molecule due to the methylene bridge, the LNA does not form the ideal shape in the Watson-Crick bond. When LNAs are incorporated into DNA or RNA oligonucleotides, LNAs can pair with complementary nucleotide chains more rapidly, increasing the stability of the double helix.

The antisense oligomer hybridizes to a target sequence in RNA by Watson-Crick base pairing, typically allowing the formation of mRNA and RNA:oligomeric heteroduplexes within the target sequence, a sequence of nucleotide bases and an intersubunit backbone means an oligomer having An oligomer may have exact sequence complementarity or approximate complementarity to a target sequence.

Since the biomarker protein according to the present invention or information on the gene encoding it is known, those skilled in the art can easily design a primer, probe or antisense nucleotide that specifically binds to the gene encoding the protein based on this information. .

In addition, the present invention is ischemic comprising an antibody or aptamer that specifically binds to ADGRG3 (adhesion G protein-coupled receptor G3) protein, or a primer, probe or antisense nucleotide that specifically binds to a gene encoding the protein. A kit for diagnosing a disease or prognosis is provided.

In addition, the present invention is ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 ( Intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) antibody or aptamer that specifically binds to any one protein selected from the group consisting of, or Provided is a kit for diagnosing ischemic disease or prognosis comprising a primer, a probe, or an antisense nucleotide that specifically binds to a gene encoding the protein.

The adhesion G protein-coupled receptor G3 (ADGRG3) is represented by the amino acid sequence of SEQ ID NO: 1, apoptosis associated tyrosine kinase (AATK) is represented by the amino acid sequence of SEQ ID NO: 2, and arachidonate 5-lipoxygenase (ALOX5) is represented by SEQ ID NO: 3, CD177 (CD177 molecule) is represented by the amino acid sequence of SEQ ID NO: 4, CREB5 (cAMP responsive element binding protein 5) is represented by the amino acid sequence of SEQ ID NO: 5, ICAM3 (intercellular adhesion molecule 3) ) is represented by the amino acid sequence of SEQ ID NO: 6, LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) is represented by the amino acid sequence of SEQ ID NO: 7, S100A6 (S100 calcium binding protein A6) of SEQ ID NO: 8 It may be one represented by an amino acid sequence.

The ischemic disease is cerebral infarction, cerebral ischemia, cerebral hypoxia, ischemic heart disease, heart failure, myocardial ischemia, ischemic renal failure, skeletal muscle ischemia, intestinal ischemia, sleep apnea, hypoxia during sleep, focal perfusion defect, retinal ischemia, neonatal hypoxia, fetal hypoxia, thrombosis , may be any one selected from the group consisting of altitude sickness, carbon monoxide poisoning, sudden infant death syndrome, and congenital circulatory disorders, but may include all other ischemic diseases known in the art.

The kit consists of a RT-PCR kit, a PCR kit, a nucleic acid amplification kit, a DNA chip kit, an ELISA kit, a protein chip kit, a rapid kit, a multiple reaction monitoring (MRM) kit, and a lateral flow analysis (LFA) kit. It may be any one selected from.

The kit may further include one or more other component compositions, solutions or devices suitable for the assay method.

For example, the diagnostic or prognostic diagnostic kit of the present invention may further include essential elements necessary for performing the reverse transcription polymerase reaction. The reverse transcription polymerase reaction kit includes a pair of primers specific for a gene encoding a marker protein. The primer is a nucleotide having a sequence specific to the nucleic acid sequence of the gene, and may have a length of about 7 bp to 50 bp, more preferably about 10 bp to 30 bp. It may also include a primer specific for the nucleic acid sequence of the control gene. Other Reverse Transcription Polymerase Reaction Kits include test tubes or other suitable containers, reaction buffers (varying pH and magnesium concentrations), deoxynucleotides (dNTPs), enzymes such as Taq-polymerase and reverse transcriptase, DNase, RNase inhibitor DEPC -Water (DEPC-water), sterile water, etc. may be included.

In addition, the kit for diagnosis or prognostic diagnosis of the present invention may include essential elements necessary for performing a DNA chip. The DNA chip kit may include a substrate to which cDNA or oligonucleotide corresponding to a gene or fragment thereof is attached, and reagents, agents, enzymes, etc. for preparing a fluorescently-labeled probe. The substrate may also contain cDNA or oligonucleotides corresponding to control genes or fragments thereof.

In addition, the kit for diagnosis or prognostic diagnosis of the present invention may include essential elements necessary for performing ELISA. The ELISA kit contains an antibody specific for this protein. Antibodies are antibodies with high specificity and affinity for a marker protein and little cross-reactivity with other proteins, and are monoclonal antibodies, polyclonal antibodies, or recombinant antibodies. The ELISA kit may also include an antibody specific for a control protein. Other ELISA kits include reagents capable of detecting bound antibody, for example, labeled secondary antibodies, chromophores, enzymes (eg, conjugated with an antibody) and substrates thereof or capable of binding the antibody. other materials and the like.

In addition, the present invention is ischemic comprising an antibody or aptamer that specifically binds to ADGRG3 (adhesion G protein-coupled receptor G3) protein, or a primer, probe or antisense nucleotide that specifically binds to a gene encoding the protein. Provided is a biochip for disease diagnosis or prognosis diagnosis.

In addition, the present invention is ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 ( Intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) antibody or aptamer that specifically binds to any one protein selected from the group consisting of, or Provided is a biochip for diagnosing ischemic disease or prognosis comprising a primer, a probe, or an antisense nucleotide that specifically binds to a gene encoding the protein.

The adhesion G protein-coupled receptor G3 (ADGRG3) is represented by the amino acid sequence of SEQ ID NO: 1, apoptosis associated tyrosine kinase (AATK) is represented by the amino acid sequence of SEQ ID NO: 2, and arachidonate 5-lipoxygenase (ALOX5) is represented by SEQ ID NO: 3, CD177 (CD177 molecule) is represented by the amino acid sequence of SEQ ID NO: 4, CREB5 (cAMP responsive element binding protein 5) is represented by the amino acid sequence of SEQ ID NO: 5, ICAM3 (intercellular adhesion molecule 3) ) is represented by the amino acid sequence of SEQ ID NO: 6, LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) is represented by the amino acid sequence of SEQ ID NO: 7, S100A6 (S100 calcium binding protein A6) of SEQ ID NO: 8 It may be one represented by an amino acid sequence.

The ischemic disease is cerebral infarction, cerebral ischemia, cerebral hypoxia, ischemic heart disease, heart failure, myocardial ischemia, ischemic renal failure, skeletal muscle ischemia, intestinal ischemia, sleep apnea, hypoxia during sleep, focal perfusion defect, retinal ischemia, neonatal hypoxia, fetal hypoxia, thrombosis , may be any one selected from the group consisting of altitude sickness, carbon monoxide poisoning, sudden infant death syndrome, and congenital circulatory disorders, but may include all other ischemic diseases known in the art.

The biochip may be a protein chip or a DNA chip.

In addition, the present invention is ischemic comprising an antibody or aptamer that specifically binds to ADGRG3 (adhesion G protein-coupled receptor G3) protein, or a primer, probe or antisense nucleotide that specifically binds to a gene encoding the protein. Provided is a composition for disease diagnosis or prognosis diagnosis.

In addition, the present invention is ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 ( Intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) antibody or aptamer that specifically binds to any one protein selected from the group consisting of, or Provided is a composition for diagnosing ischemic disease or prognosis comprising a primer, a probe, or an antisense nucleotide that specifically binds to a gene encoding the protein.

The adhesion G protein-coupled receptor G3 (ADGRG3) is represented by the amino acid sequence of SEQ ID NO: 1, apoptosis associated tyrosine kinase (AATK) is represented by the amino acid sequence of SEQ ID NO: 2, and arachidonate 5-lipoxygenase (ALOX5) is represented by SEQ ID NO: 3, CD177 (CD177 molecule) is represented by the amino acid sequence of SEQ ID NO: 4, CREB5 (cAMP responsive element binding protein 5) is represented by the amino acid sequence of SEQ ID NO: 5, ICAM3 (intercellular adhesion molecule 3) ) is represented by the amino acid sequence of SEQ ID NO: 6, LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) is represented by the amino acid sequence of SEQ ID NO: 7, S100A6 (S100 calcium binding protein A6) of SEQ ID NO: 8 It may be one represented by an amino acid sequence.

The ischemic disease is cerebral infarction, cerebral ischemia, cerebral hypoxia, ischemic heart disease, heart failure, myocardial ischemia, ischemic renal failure, skeletal muscle ischemia, intestinal ischemia, sleep apnea, hypoxia during sleep, focal perfusion defect, retinal ischemia, neonatal hypoxia, fetal hypoxia, thrombosis , may be any one selected from the group consisting of altitude sickness, carbon monoxide poisoning, sudden infant death syndrome, and congenital circulatory disorders, but may include all other ischemic diseases known in the art.

In addition, the present invention provides a diagnosis or prognosis of an ischemic disease comprising measuring the expression level of an adhesion G protein-coupled receptor G3 (ADGRG3) protein, or a gene encoding the protein, in a biological sample isolated from a subject of interest A method for providing information for diagnosis is provided.

In addition, the present invention provides ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP) in a biological sample isolated from a subject of interest. responsive element binding protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) any one protein selected from the group consisting of, or the above It provides a method for providing information for diagnosing ischemic disease or diagnosing a prognosis, comprising measuring the expression level of a gene encoding a protein.

The adhesion G protein-coupled receptor G3 (ADGRG3) is represented by the amino acid sequence of SEQ ID NO: 1, apoptosis associated tyrosine kinase (AATK) is represented by the amino acid sequence of SEQ ID NO: 2, and arachidonate 5-lipoxygenase (ALOX5) is represented by SEQ ID NO: 3, CD177 (CD177 molecule) is represented by the amino acid sequence of SEQ ID NO: 4, CREB5 (cAMP responsive element binding protein 5) is represented by the amino acid sequence of SEQ ID NO: 5, ICAM3 (intercellular adhesion molecule 3) ) is represented by the amino acid sequence of SEQ ID NO: 6, LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) is represented by the amino acid sequence of SEQ ID NO: 7, S100A6 (S100 calcium binding protein A6) of SEQ ID NO: 8 It may be one represented by an amino acid sequence.

The ischemic disease is cerebral infarction, cerebral ischemia, cerebral hypoxia, ischemic heart disease, heart failure, myocardial ischemia, ischemic renal failure, skeletal muscle ischemia, intestinal ischemia, sleep apnea, hypoxia during sleep, focal perfusion defect, retinal ischemia, neonatal hypoxia, fetal hypoxia, thrombosis , may be any one selected from the group consisting of altitude sickness, carbon monoxide poisoning, sudden infant death syndrome, and congenital circulatory disorders, but may include all other ischemic diseases known in the art.

The target subject refers to an individual whose onset is uncertain or highly likely to develop an ischemic disease, and refers to all mammals including humans, monkeys, apes, rats and mice.

The biological sample refers to any material, biological fluid, tissue or cell obtained from or derived from an individual, and is preferably selected from the group consisting of tissues, cells, blood, serum, plasma, saliva and urine. do.

The agent for measuring the expression level of the protein is not particularly limited, but preferably an antibody, oligopeptide, ligand, PNA (peptide nucleic acid) and aptamer that specifically binds to the protein encoded by the biomarker gene ( aptamer) may include one or more selected from the group consisting of.

Methods for measuring or comparing the expression level of the protein include protein chip analysis, immunoassay, ligand binding assay, MALDI-TOF analysis, SELDI-TOF analysis, radioimmunoassay, radioimmunodiffusion method, Oukteroni immunodiffusion method, There are rocket immunoelectrophoresis, tissue immunostaining, complement fixation assay, two-dimensional electrophoresis analysis, liquid chromatography-mass spectrometry (LC-MS), LC-MS/MS, Western blotting or ELISA (enzyme linked immunosorbent assay), etc. , but is not limited thereto.

The agent for measuring the expression level of the gene may include at least one selected from the group consisting of primers, probes, and antisense nucleotides that specifically bind to the biomarker gene.

As an analysis method for measuring the expression level of the gene, reverse transcription polymerase reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RT-PCR), real-time reverse transcription polymerase reaction (Real-time RT-PCR), RNase protection RNase protection assay (RPA), Northern blotting, or DNA chip, but is not limited thereto.

In addition, the present invention comprises the steps of treating a candidate substance expected to prevent or treat an ischemic disease in the isolated biological sample; And, measuring the expression level of ADGRG3 (adhesion G protein-coupled receptor G3) protein, or a gene encoding the protein in the biological sample treated with the candidate substance; A screening method is provided.

In addition, the present invention comprises the steps of treating a candidate substance expected to prevent or treat ischemic disease in the isolated biological sample; And, in the biological sample treated with the candidate substance, ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding) protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) and S100A6 (S100 calcium binding protein A6) any one protein selected from the group consisting of, or encoding the protein It provides a screening method of a drug for the prevention or treatment of ischemic disease, comprising the step of measuring the expression level of the gene.

The adhesion G protein-coupled receptor G3 (ADGRG3) is represented by the amino acid sequence of SEQ ID NO: 1, apoptosis associated tyrosine kinase (AATK) is represented by the amino acid sequence of SEQ ID NO: 2, and arachidonate 5-lipoxygenase (ALOX5) is represented by SEQ ID NO: 3, CD177 (CD177 molecule) is represented by the amino acid sequence of SEQ ID NO: 4, CREB5 (cAMP responsive element binding protein 5) is represented by the amino acid sequence of SEQ ID NO: 5, ICAM3 (intercellular adhesion molecule 3) ) is represented by the amino acid sequence of SEQ ID NO: 6, LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated) is represented by the amino acid sequence of SEQ ID NO: 7, S100A6 (S100 calcium binding protein A6) of SEQ ID NO: 8 It may be one represented by an amino acid sequence.

The ischemic disease is cerebral infarction, cerebral ischemia, cerebral hypoxia, ischemic heart disease, heart failure, myocardial ischemia, ischemic renal failure, skeletal muscle ischemia, intestinal ischemia, sleep apnea, hypoxia during sleep, focal perfusion defect, retinal ischemia, neonatal hypoxia, fetal hypoxia, thrombosis , may be any one selected from the group consisting of altitude sickness, carbon monoxide poisoning, sudden infant death syndrome, and congenital circulatory disorders, but may include all other ischemic diseases known in the art.

The biological sample refers to any material, biological fluid, tissue or cell obtained from or derived from an individual, and is preferably selected from the group consisting of tissues, cells, blood, serum, plasma, saliva and urine. do.

The candidate material includes any substance, molecule, element, compound, entity, or a combination thereof. Examples include, but are not limited to, proteins, polypeptides, small organic molecules, polysaccharides, polynucleotides, and the like. It may also be a natural product, a synthetic compound, or a combination of two or more substances.

When the expression level of the biomarker gene or a protein encoded by the biomarker gene in the biological sample after the treatment of the candidate material is increased or decreased compared to before the treatment of the candidate material, the candidate material is a drug for preventing or treating ischemic disease It may include the step of determining

The description of the agent for measuring the expression level and the method of measuring the expression level in the screening method of the present invention overlaps with that described in the information providing method for diagnosis of the present invention, and the description thereof is omitted below to avoid excessive complexity of the specification. .

Hereinafter, the present invention will be described in more detail through examples. These examples are only for illustrating the present invention in more detail, and it will be apparent to those of ordinary skill in the art that the scope of the present invention is not limited by these examples according to the gist of the present invention. .

< Example 1> Patient cerebral infarction and monkey cerebral infarction model

The gene expression of each group compared to the control group was compared in order to identify the human cerebral infarction patient blood sample for each genome/proteome at each time period: control (Pre), post-perfusion (Post), 1 day (1d), 1 week, and 4 weeks. First of all, since it is impossible to collect normal blood samples from humans, which should be originally used as a control group, the Pre sample that arrived at the hospital was set as a control group, and the change in gene expression in other groups compared to the Pre sample was observed (genome). For proteomic analysis, commercially available human serum was purchased and protein change analysis was performed between the control group and the experimental group (proteomic protein).

After induction of a monkey cerebral infarction induction model (cynomolgus), after blood collection/RNA separation before/after occlusion, 1 day, 1 week, and 4 weeks for gene expression change/analysis, genome, cytokine/chemokines were used using 44K Agilent GX array. The proteomic was analyzed using the array.

If the data analysis analyzed the functions of the genes showing a 2-fold or more expression change in each group, statistically significant results were combined and analyzed by classifying them as p-values.

< Example 2> Analysis of cytokine/chemokines array in human cerebral infarction and monkey cerebral infarction induction model blood samples

Blood from human cerebral infarction patients was isolated at Normal, Pre, Post, 1 day, 1 week, and 4 weeks. The separated blood was centrifuged at 2000 g for 20 minutes to separate serum, and the separated serum was analyzed for protein expression using a cytokine/chemokines array kit.

Patient cerebral infarction cytokine/chemokines expression differed slightly between patients. When looking at the patient's cerebral infarction expression pattern, if there was a significant change in the cytokine/chemokines expression pattern from normal to pre and post, there was a change in the increase or decrease of the expressed protein at 24 hours, 1 week, and 4 weeks. In the monkey cerebral infarction induction model, cytokine/chemokines expression varied significantly between subjects. Compared to the baseline (normal), there was no significant protein change.

In the patient's total signification, there are protein changes in the order of extracellular matrix, secretion, neurogenesis, cell migration, immune response, inflammatory response, and cell differentiation in the pre and post phases, and secretion, inflammatory response, cell migration, and immune at 24 hours and 1 week. In response, cell differentiation, and 4 weeks, total signification expression trends were shown in the order of extracellular matrix, cell migration, neurogenesis, secretion, immune response, inflammatory response, and cell differentiation. In the monkey cerebral infarction induction model, apoptotic process & cell death, cell differentiation, neurogenesis, cell cycle, cell migration, inflammatory response, secretion, immune response, aging, angiogenesis, and extracellular matrix were sequential at D0, 24 hrs, 1 week, and 4 weeks. Total signification showed a tendency to express, in particular, angiogenesis-related proteins showed expression only at D0 after induction of cerebral infarction, and extracellular matrix-related proteins only at 24 hours after induction of cerebral infarction.

In order to find similar proteins for cytokine/chemokine expression pattern and network analysis of patient cerebral infarction/monkey cerebral infarction, heatmap analysis was performed with the quantified values of these cytokine/chemokine expression levels using the imageJ program.

< Example 3> Next generation sequencing of human and monkey cerebral infarction model blood samples ( NGS ) analysis

<3-1> Gene expression analysis by human cerebral infarction ontology

After the occurrence of a human cerebral infarction, blood is separated immediately before reperfusion (Pre-control), immediately after reperfusion (Post), 1 day, and 1 week, and some isolates serum, some isolates total RNA, and mRNA expression next generation sequencing (next generation sequencing) , NGS) was performed. Total RNA quality was measured using the Agilent 2100 Bioanalyzer system, and it was confirmed that there was no problem in performing NGS.

Hypoxia, cytokine/chemokines, inflammatory response, apoptotic process & cell death, autophagy & phagocytosis, excitotoxicity, peri-infarct depolarization, and mitochondrial clustering related to the mechanism of cerebral infarction were analyzed. After the occurrence of human cerebral infarction, the expression patterns of 24 hrs and 1 wks after post were similar in most treatment groups, although there were differences according to the gene ontology as a whole for each time period before reperfusion (Pre-control), immediately after reperfusion (Post), 24 hrs, and 1 wks. , the gene expression pattern in the post group showed many changes compared to other treatment groups.

In the human cerebral infarction model, excitotoxicity, acidotoxicity, inflammation, and peri-infarct by gene ontology to confirm changes in the physiological/pathological expression of major cellular genes immediately before reperfusion (Pre), immediately after reperfusion (Post), 24 hrs, and 1 week after occurrence of human cerebral infarction in a human cerebral infarction model. Depolarization, cell death, hypoxia, oxidative stress, ER stress, mTOR, autophagy & phagocytosis, astrocyte, mitochondria, neuron, astrocyte, microglia, endothelial, and cell pericyte were classified and analyzed. As a result of the overall analysis, the expression of all gene groups showed a tendency to decrease at 24 hrs, and then to increase again at 1 week. .

<3-2> monkey cerebral infarction Gene expression analysis by induced model ontology

After induction of cerebral infarction in monkeys, blood was separated from the control group, immediately after reperfusion (Post), 1 day, 1 week (1wks), and 4 weeks (4wks). next generation sequencing (NGS) was performed. Total RNA quality was measured using the Agilent 2100 Bioanalyzer system, and it was confirmed that there was no problem in performing NGS.

Hypoxia, cytokine/chemokines, inflammatory response, apoptotic process & cell death, autophagy & phagocytosis, excitotoxicity, peri-infarct depolarization, and mitochondrial clustering were analyzed as in the mechanism of cerebral infarction. After induction of cerebral infarction in monkeys, D0 1st, D0 2nd, 24hrs, 1wks, and 4wks were similar in expression patterns of D0 1st, D0 2nd, 24hrs, and 1wks in most control groups, although there were differences according to gene ontology as a whole. Gene expression patterns showed many changes compared to other treatment groups.

Excitotoxicity, acidotoxicity, inflammation, and peritoneal changes in major cell physiology/pathological expression of genes immediately before reperfusion (Pre), immediately after reperfusion (Post), 24 hrs, and 1 week after occurrence of human cerebral infarction in a monkey cerebral infarction induction model. -infarct depolarization, cell death, hypoxia, oxidative stress, ER stress, mTOR, autophagy & phagocytosis, astrocyte, mitochondria, neuron, astrocyte, microglia, endothelial, and cell pericyte were classified and analyzed. As a result of the overall analysis, when looking at all cellular, physiological and pathological mechanisms, when the highest expression pattern was shown at 24 hours, the gene expression pattern showed a tendency to continuously decrease at the 1st and 4th weeks, but ER stress, mTOR signal In the mechanism, it was confirmed that the gene expression showed the highest point from the 2nd of D0, but the gene expression pattern decreased from the 24th hour.

As a result, ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 (intercellular adhesion) It was confirmed that molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated), and S100A6 (S100 calcium binding protein A6) are major biomarkers showing significant expression changes at each cerebral infarction period.

< Example 4> Analysis of the efficacy of transplantation of cell therapy in a monkey cerebral infarction induction model

<4-1> Used in cerebral infarction induction model human neural stem cells characterization analysis

Human neural stem cells extracted from 15-week-old fetal brain tissue were prepared using the v-myc gene. As a result of immunofluorescence staining and RT-PCR analysis of the LK1.NSCc neural stem cell line, it was confirmed that it had the characteristics of a typical neural stem cell line.

On the second day after transplantation, fluorescent nanoparticles were loaded into the human neural stem cell line to be transplanted into the monkey brain infarction induction model to obtain an MRI image. Nanoparticles If fluorescent nanoparticles can be observed in the cells from 11 hours, the presence of nanoparticles was observed up to 3 times of subculture.

<4-2> In a monkey cerebral infarction induction model human neural stem cell line Efficacy evaluation of therapeutic transplantation

At 90 minutes after induction of cerebral infarction in monkeys, IA-2X106, SWI, and DWI 30-minute images were taken with IA-2X106, SWI, and DWI cell therapy loaded with fluorescent nanoparticles, and IV-6X106 cells were transplanted on the 1st and 2nd days after induction of cerebral infarction. Blood cells were collected at D8 (baseline), 90 minutes after cerebral infarction (D0), 1wks, and 4wks, and analyzed for NGS and cytokine/chemokines array.

Overall, it was confirmed that apoptosis, inflammatory response, and cytokine/chemokines-related gene expression were efficacious in the human neural stem cell line treatment group transplanted.

As a result of analyzing the genomic expression patterns of neurons, astrocytes, microglia, perivascular cells, and cerebrovascular endothelial cells that make up the brain, the expression increased in the cell treatment group transplanted compared to the control group that induced cerebral infarction. However, in the case of microglia, it was confirmed that the gene expression pattern was increased in the control group than in the treatment group at the D0, 1wks sample period.

As a result of analyzing the cytokine/chemokines array after transplanting the cell therapy into the monkey brain infarction model, it was confirmed that the overall cytokine/chemokines protein expression pattern decreased in the treatment group transplanted with the cell therapy.

As a result, ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 (intercellular adhesion) It was confirmed that molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated), and S100A6 (S100 calcium binding protein A6) show changes in expression after transplantation of cell therapy into a cerebral infarction induction model.

< Example 5> dot Dot blot used biomarker verification

Biomarkers discovered as control and cerebral infarction samples were verified using dot blot.

Specifically, the nitrocellulose membrane was pre-wetted in TBS before using the dot equipment. After inserting the nitrocellulose membrane into the dot device, the screw was securely locked to prevent cross-well contamination. To ensure uniform binding of antigen, the membrane was re-wetted with 100 ul TBS per well. The flow valve was set to air and about 250ul of the antigen was added. A gentle vacuum was created to allow the entire sample to be filtered through the membrane by gravity flow. Washing was performed by adding 200 ul of TBS to each well. When washing of each well was completely finished, blocking was performed by removing the NC membrane from the equipment. The remaining part was performed in the same manner as western blot.

As a result, ADGRG3 (adhesion G protein-coupled receptor G3), AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 (intercellular adhesion) It was confirmed that molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and mRNA degradation associated), and S100A6 (S100 calcium binding protein A6) are major biomarkers showing significant expression changes according to the cerebral infarction period ( FIGS. 1 to 3 ). .

<110> EBIOGEN CO., LTD. <120> Biomarker for diagnosis or prognosis of ischemic disease <130> NP21-1107 <160> 8 <170> KoPatentIn 3.0 <210> 1 <211> 549 <212> PRT <213> Homo sapiens <400> 1 Met Ala Thr Pro Arg Gly Leu Gly Ala Leu Leu Leu Leu Leu Leu Leu Leu 1 5 10 15 Pro Thr Ser Gly Gln Glu Lys Pro Thr Glu Gly Pro Arg Asn Thr Cys 20 25 30 Leu Gly Ser Asn Asn Met Tyr Asp Ile Phe Asn Leu Asn Asp Lys Ala 35 40 45 Leu Cys Phe Thr Lys Cys Arg Gln Ser Gly Ser Asp Ser Cys Asn Val 50 55 60 Glu Asn Leu Gln Arg Tyr Trp Leu Asn Tyr Glu Ala His Leu Met Lys 65 70 75 80 Glu Gly Leu Thr Gln Lys Val Asn Thr Pro Phe Leu Lys Ala Leu Val 85 90 95 Gln Asn Leu Ser Thr Asn Thr Ala Glu Asp Phe Tyr Phe Ser Leu Glu 100 105 110 Pro Ser Gln Val Pro Arg Gln Val Met Lys Asp Glu Asp Lys Pro Pro 115 120 125 Asp Arg Val Arg Leu Pro Lys Ser Leu Phe Arg Ser Leu Pro Gly Asn 130 135 140 Arg Ser Val Val Arg Leu Ala Val Thr Ile Leu Asp Ile Gly P ro Gly 145 150 155 160 Thr Leu Phe Lys Gly Pro Arg Leu Gly Leu Gly Asp Gly Ser Gly Val 165 170 175 Leu Asn Asn Arg Leu Val Gly Leu Ser Val Gly Gln Met His Val Thr 180 185 190 Lys Leu Ala Glu Pro Leu Glu Ile Val Phe Ser His Gln Arg Pro Pro 195 200 205 Pro Asn Met Thr Leu Thr Cys Val Phe Trp Asp Val Thr Lys Gly Thr 210 215 220 Thr Gly Asp Trp Ser Ser Glu Gly Cys Ser Thr Glu Val Arg Pro Glu 225 230 235 240 Gly Thr Val Cys Cys Cys Asp His Leu Thr Phe Phe Ala Leu Leu Leu 245 250 255 Arg Pro Thr Leu Asp Gln Ser Thr Val His Ile Leu Thr Arg Ile Ser 260 265 270 Gln Ala Gly Cys Gly Val Ser Met Ile Phe Leu Ala Phe Thr Ile Ile 275 280 285 Leu Tyr Ala Phe Leu Arg Leu Ser Arg Glu Arg P he Lys Ser Glu Asp 290 295 300 Ala Pro Lys Ile His Val Ala Leu Gly Gly Ser Leu Phe Leu Leu Asn 305 310 315 320 Leu Ala Phe Leu Val Asn Val Gly Ser Gly Ser Lys Gly Ser Asp Ala 325 330 335 Ala Cys Trp Ala Arg Gly Ala Val Phe His Tyr Phe Leu Leu Cys Ala 340 345 350 Phe Thr Trp Met Gly Leu Glu Ala Phe His Leu Tyr Leu Leu Ala Val 355 360 365 Arg Val Phe Asn Thr Tyr Phe Gly His Tyr Phe Leu Lys Leu Ser Leu 370 375 380 Val Gly Trp Gly Leu Pro Ala Leu Met Val Ile Gly Thr Gly Ser Ala 385 390 395 400 Asn Ser Tyr Gly Leu Tyr Thr Ile Arg Asp Arg Glu Asn Arg Thr Ser 405 410 415 Leu Glu Leu Cys Trp Phe Arg Glu Gly Thr Thr Met Tyr Ala Leu Tyr 420 425 430 Ile Thr Val His Gly Tyr Phe Leu I le Thr Phe Leu Phe Gly Met Val 435 440 445 Val Leu Ala Leu Val Val Trp Lys Ile Phe Thr Leu Ser Arg Ala Thr 450 455 460 Ala Val Lys Glu Arg Gly Lys Asn Arg Lys Lys Val Leu Thr Leu Leu 465 470 475 480 Gly Leu Ser Ser Leu Val Gly Val Thr Trp Gly Leu Ala Ile Phe Thr 485 490 495 Pro Leu Gly Leu Ser Thr Val Tyr Ile Phe Ala Leu Phe Asn Ser Leu 500 505 510 Gln Gly Val Phe Ile Cys Cys Trp Phe Thr Ile Leu Tyr Leu Pro Ser 515 520 525 Gln Ser Thr Thr Val Ser Ser Ser Thr Ala Arg Leu Asp Gln Ala His 530 535 540 Ser Ala Ser Gln Glu 545 <210> 2 <211> 1374 <212> PRT <213> Homo sapiens < 400> 2 Met Ser Ser Ser Phe Phe Asn Pro Ser Phe Ala Phe Ser Ser His Phe 1 5 10 15 Asp Pro Asp Gly Ala Pro Leu Ser Glu Leu Ser Trp Pro Ser Ser Leu 20 25 30 Ala Val Val Ala Val Ser Phe Ser Gly Leu Phe Ala Val Ile Val Leu 35 40 45 Met Leu Ala Cys Leu Cys Cys Lys Lys Gly Gly Ile Gly Phe Lys Glu 50 55 60 Phe Glu Asn Ala Glu Gly Asp Glu Tyr Ala Ala Asp Leu Ala Gln Gly 65 70 75 80 Ser Pro Ala Thr Ala Ala Gln Asn Gly Pro Asp Val Tyr Val Leu Pro 85 90 95 Leu Thr Glu Val Ser Leu Pro Met Ala Lys Gln Pro Gly Arg Ser Val 100 105 110 Gln Leu Leu Lys Ser Thr Asp Val Gly Arg His Ser Leu Leu Tyr Leu 115 120 125 Lys Glu Ile Gly Arg Gly Trp Phe Gly Lys Val Phe Leu Gly Glu Val 130 135 140 Asn Ser Gly Ile Ser Ser Ser Ala Gln Val Val Val Lys Glu Leu Gln Ala 145 150 155 160 Ser Ala Ser Val Gln Glu Gln Met Gln Phe Leu Glu Glu Val Gln Pro 165 170 175 Tyr Arg Ala Leu Lys His Ser Asn Leu Leu Gln Cys Leu Ala Gln Cys 180 185 190 Ala Glu Val Thr Pro Tyr Leu Leu Val Met Glu Phe Cys P ro Leu Gly 195 200 205 Asp Leu Lys Gly Tyr Leu Arg Ser Cys Arg Val Ala Glu Ser Met Ala 210 215 220 Pro Asp Pro Arg Thr Leu Gln Arg Met Ala Cys Glu Val Ala Cys Gly 225 230 235 240 Val Leu His Leu His Arg Asn Asn Phe Val His Ser Asp Leu Ala Leu 245 250 255 Arg Asn Cys Leu Leu Thr Ala Asp Leu Thr Val Lys Ile Gly Asp Tyr 260 265 270 Gly Leu Ala His Cys Lys Tyr Arg Glu Asp Tyr Phe Val Thr Ala Asp 275 280 285 Gln Leu Trp Val Pro Leu Arg Trp Ile Ala Pro Glu Leu Val Asp Glu 290 295 300 Val His Ser Asn Leu Leu Val Val Asp Gln Thr Lys Ser Gly Asn Val 305 310 315 320 Trp Ser Leu Gly Val Thr Ile Trp Glu Leu Phe Glu Leu Gly Thr Gln 325 330 335 Pro Tyr Pro Gln His Ser Asp Gln Gln Val L eu Ala Tyr Thr Val Arg 340 345 350 Glu Gln Gln Leu Lys Leu Pro Lys Pro Gln Leu Gln Leu Thr Leu Ser 355 360 365 Asp Arg Trp Tyr Glu Val Met Gln Phe Cys Trp Leu Gln Pro Glu Gln 370 375 380 Arg Pro Thr Ala Glu Glu Val His Leu Leu Leu Ser Tyr Leu Cys Ala 385 390 395 400 Lys Gly Ala Thr Glu Ala Glu Glu Glu Phe Glu Arg Arg Trp Arg Ser 405 410 415 Leu Arg Pro Gly Gly Gly Gly Val Gly Pro Gly Pro Gly Ala Ala Gly 420 425 430 Pro Met Leu Gly Gly Val Val Glu Leu Ala Ala Ala Ser Ser Phe Pro 435 440 445 Leu Leu Glu Gln Phe Ala Gly Asp Gly Phe His Ala Asp Gly Asp Asp 450 455 460 Val Leu Thr Val Thr Glu Thr Ser Arg Gly Leu Asn Phe Glu Tyr Lys 465 470 475 480 Trp Glu Ala Gly Arg Gly Ala G lu Ala Phe Pro Ala Thr Leu Ser Pro 485 490 495 Gly Arg Thr Ala Arg Leu Gln Glu Leu Cys Ala Pro Asp Gly Ala Pro 500 505 510 Pro Gly Val Val Pro Val Leu Ser Ala His Ser Pro Ser Leu Gly Ser 515 520 525 Glu Tyr Phe Ile Arg Leu Glu Glu Ala Ala Pro Ala Ala Gly His Asp 530 535 540 Pro Asp Cys Ala Gly Cys Ala Pro Ser Pro Pro Ala Thr Ala Asp Gln 545 550 555 560 Asp Asp Asp Ser Asp Gly Ser Thr Ala Ala Ser Leu Ala Met Glu Pro 565 570 575 Leu Leu Gly His Gly Pro Pro Val Asp Val Pro Trp Gly Arg Gly Asp 580 585 590 His Tyr Pro Arg Arg Ser Leu Ala Arg Asp Pro Leu Cys Pro Ser Arg 595 600 605 Ser Pro Ser Pro Ser Ala Gly Pro Leu Ser Leu Ala Glu Gly Gly Ala 610 615 620 Glu Asp Ala Asp Trp Gly Val Ala Ala P he Cys Pro Ala Phe Phe Glu 625 630 635 640 Asp Pro Leu Gly Thr Ser Pro Leu Gly Ser Ser Gly Ala Pro Pro Leu 645 650 655 Pro Leu Thr Gly Glu Asp Glu Leu Glu Glu Val Gly Ala Arg Arg Ala 660 665 670 Ala Gln Arg Gly His Trp Arg Ser Asn Val Ser Ala Asn Asn Asn Ser 675 680 685 Gly Ser Arg Cys Pro Glu Ser Trp Asp Pro Val Ser Ala Gly Gly His 690 695 700 Ala Glu Gly Cys Pro Ser Pro Lys Gln Thr Pro Arg Ala Ser Pro Glu 705 710 715 720 Pro Gly Tyr Pro Gly Glu Pro Leu Leu Leu Gly Leu Gln Ala Ala Ser Ala 725 730 735 Gln Glu Pro Gly Cys Cys Pro Gly Leu Pro His Leu Cys Ser Ala Gln 740 745 750 Gly Leu Ala Pro Ala Pro Cys Leu Val Thr Pro Ser Trp Thr Glu Thr 755 760 765 Ala Ser Ser Gly Gly Asp H is Pro Gln Ala Glu Pro Lys Leu Ala Thr 770 775 780 Glu Ala Glu Gly Thr Thr Gly Pro Arg Leu Pro Leu Pro Ser Val Pro 785 790 795 800 Ser Pro Ser Gln Glu Gly Ala Pro Leu Pro Ser Glu Glu Ala Ser Ala 805 810 815 Pro Asp Ala Pro Asp Ala Leu Pro Asp Ser Pro Thr Pro Ala Thr Gly 820 825 830 Gly Glu Val Ser Ala Ile Lys Leu Ala Ser Ala Leu Asn Gly Ser Ser 835 840 845 Ser Ser Ser Pro Glu Val Glu Ala Pro Ser Ser Glu Asp Glu Asp Thr Ala 850 855 860 Glu Ala Thr Ser Gly Ile Phe Thr Asp Thr Ser Ser Asp Gly Leu Gln 865 870 875 880 Ala Arg Arg Pro Asp Val Val Pro Ala Phe Arg Ser Leu Gln Lys Gln 885 890 895 Val Gly Thr Pro Asp Ser Leu Asp Ser Leu Asp Ile Pro Ser Ser Ala 900 905 910 Ser Asp Gly G ly Tyr Glu Val Phe Ser Pro Ser Ala Thr Gly Pro Ser 915 920 925 Gly Gly Gln Pro Arg Ala Leu Asp Ser Gly Tyr Asp Thr Glu Asn Tyr 930 935 940 Glu Ser Pro Glu Phe Val Leu Lys Glu Ala Gln Glu Gly Cys Glu Pro 945 950 955 960 Gln Ala Phe Ala Glu Leu Ala Ser Glu Gly Glu Gly Pro Gly Pro Glu 965 970 975 Thr Arg Leu Ser Thr Ser Leu Ser Gly Leu Asn Glu Lys Asn Pro Tyr 980 985 990 Arg Asp Ser Ala Tyr Phe Ser Asp Leu Glu Ala Glu Ala Glu Ala Thr 995 1000 1005 Ser Gly Pro Glu Lys Lys Cys Gly Gly Asp Arg Ala Pro Gly Pro Glu 1010 1015 1020 Leu Gly Leu Pro Ser Thr Gly Gln Pro Ser Glu Gln Val Cys Leu Arg 1025 1030 1035 1040 Pro Gly Val Ser Gly Glu Ala Gln Gly Ser Gly Pro Gly Glu Val Leu 1045 1050 1055 Pro Pro Leu Leu Gln Leu Glu Gly Ser Ser Pro Glu Pro Ser Thr Cys 1060 1065 1070 Pro Ser Gly Leu Val Pro Glu Pro Pro Glu Pro Gln Gly Pr o Ala Lys 1075 1080 1085 Val Arg Pro Gly Pro Ser Pro Ser Cys Ser Gln Phe Phe Leu Leu Thr 1090 1095 1100 Pro Val Pro Leu Arg Ser Glu Gly Asn Ser Ser Glu Phe Gln Gly Pro 1105 1110 1115 1120 Pro Gly Leu Leu Ser Gly Pro Ala Pro Gln Lys Arg Met Gly Gly Pro 1125 1130 1135 Gly Thr Pro Arg Ala Pro Leu Arg Leu Ala Leu Pro Gly Leu Pro Ala 1140 1145 1150 Ala Leu Glu Gly Arg Pro Glu Glu Glu Glu Glu Asp Ser Glu Asp Ser 1155 1160 1165 Asp Glu Ser Asp Glu Glu Leu Arg Cys Tyr Ser Val Gln Glu Pro Ser 1170 1175 1180 Glu Asp Ser Glu Glu Glu Ala Pro Ala Val Pro Val Val Val Ala Glu 1185 1190 1195 1200 Ser Gln Ser Ala Arg Asn Leu Arg Ser Leu Leu Lys Met Pro Ser Leu 1205 1210 1215 Leu Ser Glu Thr Phe Cys Glu Asp Leu Glu Arg Lys Lys Lys Lys Ala Val 1220 1225 1230 Ser Phe Phe Asp Asp Val Thr Val Tyr Leu Phe Asp Gln Glu Ser Pro 1235 1240 1245 Thr Arg Glu Leu Gly Glu Pro Phe Pro Gly Ala Lys Glu Ser Pro Pro 1250 1255 1260 Thr Phe Leu Arg Gly Ser Pro Gly Ser Pro Ser Ala Pro Asn Arg Pro 1265 1270 1275 1280 Gln Gln Ala Asp Gly Ser Pro Asn Gly Ser Thr Ala Glu Glu Gly Gly 1285 1290 1295 Gly Phe Ala Trp Asp Asp Asp Phe Pro Leu Met Thr Ala Lys Ala Ala 1300 1305 1310 Phe Ala Met Ala Leu Asp Pro Ala Ala Pro Ala Pro Ala Ala Pro Thr 1315 1320 1325 Pro Thr Pro Ala Pro Phe Ser Arg Phe Thr Val Ser Pro Ala Pro Thr 1330 1335 1340 Ser Arg Phe Ser Ile Thr His Val Ser Asp Ser Asp Ala Glu Ser Lys 1345 1350 1355 1360 Arg Gly Pro Glu Ala Gly Ala Gly Gly Glu Ser Lys Glu Ala 1365 1370 <210> 3 <211> 674 <212> PRT <213> Homo sapiens <400> 3 Met Pro Ser Tyr Thr Val Thr Val Ala Thr Gly Ser Gln Trp Phe Ala 1 5 10 15 Gly Thr Asp Asp Tyr Ile Tyr Leu Ser Leu Val Gly Ser Ala Gly Cys 20 25 30 Ser Glu Lys His Leu Leu Asp Lys Pro Phe Tyr Asn Asp Phe Glu Arg 35 40 45 Gly Ala Val Asp Ser Tyr Asp Val Thr Val Asp Glu Glu Leu Gly Glu 50 55 60 Ile Gln Leu Val Arg Ile Glu Lys Arg Lys Tyr Trp Leu Asn Asp Asp 65 70 75 80 Trp Tyr Leu Lys Tyr Ile Thr Leu Lys Thr Pro His Gly Asp Tyr Ile 85 90 95 Glu Phe Pro Cys Tyr Arg Trp Ile Thr Gly Asp Val Glu Val Val Leu 100 105 110 Arg Asp Gly Arg Ala Lys Leu Ala Arg Asp Asp Gln Ile His Ile Leu 115 120 125 Lys Gln His Arg Arg Lys Glu Leu Glu Thr Arg Gln Lys Gln Tyr Arg 130 135 140 Trp Met Glu Trp Asn Pro Gly Phe Pro Leu Ser Ile Asp Ala Lys Cys 145 150 155 160 His Lys Asp Leu Pro Arg Asp Ile Gln Phe Asp Ser Glu Lys Gly Val 165 170 175 Asp Phe Val Leu Asn Tyr Ser Lys Ala Met Glu Asn Leu Phe Ile Asn 180 185 190 Arg Phe Met His Met Phe Gln Ser Ser Trp Asn Asp Phe Ala Asp Phe 195 200 205 Glu Lys Ile Phe Val Lys Ile Ser Asn Thr Ile Ser Glu Arg Val Met 210 215 220 Asn His Trp Gln Glu Asp Leu Met Phe Gly Tyr Gln Phe Leu Asn Gly 225 230 235 240 Cys Asn Pro Val Leu Ile Arg Arg Cys Thr Glu Leu Pro Glu Lys Leu 245 250 255 Pro Val Thr Thr Glu Met Val Glu Cys Ser Leu Glu Arg Gln Leu Ser 260 265 270 Leu Glu Gln Glu Val Gln Gln Gly Asn Ile Phe Ile Val Asp Phe Glu 275 280 285 Leu Leu Asp Gly Ile Asp Ala Asn Lys Thr Asp Pro Cys Thr Leu Gln 290 295 300 Phe Leu Ala Ala Pro Ile Cys Leu Leu Tyr Lys Asn Leu Ala Asn Lys 305 310 315 320 Ile Val Pro Ile Ala Ile Gln Leu Asn Gln Ile Pro Gly Asp Glu Asn 325 330 335 Pro Ile Phe Leu Pro Ser Asp Ala Lys Tyr Asp Trp Leu Leu Ala Lys 340 345 350 Ile Trp Val Arg Ser Ser Asp Phe His Val His Gln Thr Ile Thr His 355 360 365 Leu Leu Arg Thr His Leu Val Ser Glu Val Phe Gly Ile Ala Met Tyr 370 375 380 Arg Gln Leu Pro Ala Val His Pro Ile Phe Lys Leu Leu Val Ala His 385 390 395 400 Val Arg Phe Thr Ile Ala Ile Asn Thr Lys Ala Arg Glu Gln Leu Ile 405 410 415 Cys Glu Cys Gly Leu Phe Asp Lys Ala Asn Ala Thr Gly Gly Gly Gly 420 425 430 His Val Gln Met Val Gln Arg Ala Met Lys Asp Leu Thr Tyr Ala Ser 435 440 445 Leu Cys Phe Pro Glu Ala Ile Lys Ala Arg Gly Met Glu Ser Lys Glu 450 455 460 Asp Ile Pro Tyr Tyr Phe Tyr Arg Asp Asp Gly Leu Leu Val Trp Glu 465 470 475 480 Ala Ile Arg Thr Phe Thr Ala Glu Val Val Asp Ile Tyr Tyr Glu Gly 485 490 495 Asp Gln Val Val Glu Glu Asp Pro Glu Leu Gln Asp Phe Val Asn Asp 500 505 510 Val Tyr Val Tyr Gly Met Arg Gly Arg Lys Ser Ser Gly Phe Pro Lys 515 520 525 Ser Val Lys Ser Arg Glu Gln Leu Ser Glu Tyr Leu Thr Val Val Ile 530 535 540 Phe Thr Ala Ser Ala Gln His Ala Ala Val Asn Phe Gly Gln Tyr Asp 545 550 555 560 Trp Cys Ser Trp Ile Pro Asn Ala Pro Pro Thr Met Arg Ala Pro Pro 565 570 575 Pro Thr Ala Lys Gly Val Val Thr Ile Glu Gln Ile Val Asp Thr Leu 580 585 590 Pro Asp Arg Gly Arg Ser Cys Trp His Leu Gly Ala Val Trp Ala Leu 595 600 605 Ser Gln Phe Gln Glu Asn Glu Leu Phe Leu Gly Met Tyr Pro Glu Glu 610 615 620 His Phe Ile Glu Lys Pro Val Lys Glu Ala Met Ala Arg Phe Arg Lys 625 630 635 640 Asn Leu Glu Ala Ile Val Ser Val Ile Ala Glu Arg Asn Lys Lys Lys 645 650 655 Gln Leu Pro Tyr Tyr Tyr Leu Ser Pro Asp Arg Ile Pro Asn Ser Val 660 665 670 Ala Ile <210> 4 <211> 437 <212> PRT <213 > Homo sapiens <400> 4 Met Ser Ala Val Leu Leu Leu Ala Leu Leu Gly Phe Ile Leu Pro Leu 1 5 10 15 Pro Gly Val Gln Ala Leu Leu Cys Gln Phe Gly Thr Val Gln His Val 20 25 30 Trp Lys Val Ser Asp Leu Pro Arg Gln Trp Thr Pro Lys Asn Thr Ser 35 40 45 Cys Asp Ser Gly Leu Gly Cys Gln Asp Thr Leu Met Leu Ile Glu Ser 50 55 60 Gly Pro Gln Val Ser Leu Val Leu Ser Lys Gly Cys Thr Glu Ala Lys 65 70 75 80 Asp Gln Glu Pro Arg Val Thr Glu His Arg Met Gly Pro Gly Leu Ser 85 90 95 Leu Ile Ser Tyr Thr Phe Val Cys Arg Gln Glu Asp Phe Cys Asn Asn 100 105 110 Leu Val Asn Ser Leu Pro Leu Trp Ala Pro Gln Pro Pro Ala Asp Pro 115 120 125 Gly Ser Leu Arg Cys Pro Val Cys Leu Ser Met Glu Gly Cys Leu Glu 130 135 140 Gly Thr Thr Glu Glu Ile Cys Pro Lys Gly Thr Thr His Cys Tyr Asp 145 150 155 160 Gly Leu Leu Arg Leu Arg Gly Gly Gly Ile Phe Ser Asn Leu Arg Val 165 170 175 Gln Gly Cys Met Pro Gln Pro Val Cys Asn Leu Leu Asn Gly Thr Gln 180 185 190 Glu Ile Gly Pro Val Gly Met Thr Glu A sn Cys Asp Met Lys Asp Phe 195 200 205 Leu Thr Cys His Arg Gly Thr Thr Ile Met Thr His Gly Asn Leu Ala 210 215 220 Gln Glu Pro Thr Asp Trp Thr Thr Ser Asn Thr Glu Met Cys Glu Val 225 230 235 240 Gly Gln Val Cys Gln Glu Thr Leu Leu Leu Leu Asp Val Gly Leu Thr 245 250 255 Ser Thr Leu Val Gly Thr Lys Gly Cys Ser Thr Val Gly Ala Gln Asn 260 265 270 Ser Gln Lys Thr Thr Ile His Ser Ala Pro Pro Gly Val Leu Val Ala 275 280 285 Ser Tyr Thr His Phe Cys Ser Ser Asp Leu Cys Asn Ser Ala Ser Ser 290 295 300 Ser Ser Val Leu Leu Asn Ser Leu Pro Pro Gln Ala Ala Pro Val Pro 305 310 315 320 Gly Asp Arg Gln Cys Pro Thr Cys Val Gln Pro Leu Gly Thr Cys Ser 325 330 335 Ser Gly Ser Pro Arg Met T hr Cys Pro Arg Gly Ala Thr His Cys Tyr 340 345 350 Asp Gly Tyr Ile His Leu Ser Gly Gly Gly Leu Ser Thr Lys Met Ser 355 360 365 Ile Gln Gly Cys Val Ala Gln Pro Ser Ser Phe Leu Leu Asn His Thr 370 375 380 Arg Gln Ile Gly Ile Phe Ser Ala Arg Glu Lys Arg Asp Val Gln Pro 385 390 395 400 Pro Ala Ser Gln His Glu Gly Gly Gly Ala Glu Gly Leu Glu Ser Leu 405 410 415 Thr Trp Gly Val Gly Leu Ala Leu Ala Pro Ala Leu Trp Trp Gly Val 420 425 430 Val Cys Pro Ser Cys 435 <210> 5 <211> 568 <212> PRT <213> Homo sapiens <400> 5 Met Ile Tyr Glu Glu Ser Lys Met Asn Leu Glu Gln Glu Arg Pro Phe 1 5 10 15 Val Cys Ser Ala Pro Gly Cys Ser Gln Arg Phe Pro Thr Glu Asp His 20 25 30 Leu Met Ile His Arg His Lys His Glu Met Thr Leu Lys Phe Pro Ser 35 40 45 Ile Lys Thr Asp Asn Met Leu Ser Asp Gln Thr Pro Met Ile Tyr Glu 50 55 60 Glu Ser Lys Met Asn Leu Glu Gln Glu Arg Pro Phe Val Cys Ser Ala 65 70 75 80 Pro Gly Cys Ser Gln Arg Phe Pro Thr Glu Asp His Leu Met Ile His 85 90 95 Arg His Lys His Glu Met Thr Leu Lys Phe Pro Ser Ile Lys Thr Asp 100 105 110 Asn Met Leu Ser Asp Gln Thr Pro Thr Pro Thr Arg Phe Leu Lys Asn 115 120 125 Cys Glu Glu Val Gly Leu Phe Ser Glu Leu Asp Cys Ser Leu Glu His 130 135 140 Glu Phe Arg Lys Ala Gln Glu Glu Glu Ser Ser Lys Arg Asn Ile Ser 145 150 155 160 Met His Asn Ala Val Gly Gly Ala Met Thr Gly Pro Gly Thr His Gln 165 170 175 Leu Ser Ser Ala Arg Leu Pro Asn His Asp Thr Asn Val Val Ile Gln 180 185 190 Gln Ala Met Pro Ser Pro Gln Ser Ser Ser Val Ile Thr Gln Ala Pro 195 200 205 Ser Thr Asn Arg Gln Ile Gly Pro Val Pro Gly Se r Leu Ser Ser Leu 210 215 220 Leu His Leu His Asn Arg Gln Arg Gln Pro Met Pro Ala Ser Met Pro 225 230 235 240 Gly Thr Leu Pro Asn Pro Thr Met Pro Gly Ser Ser Ala Val Leu Met 245 250 255 Pro Met Glu Arg Gln Met Ser Val Asn Ser Ser Ile Met Gly Met Gln 260 265 270 Gly Pro Asn Leu Ser Asn Pro Cys Ala Ser Pro Gln Val Gln Pro Met 275 280 285 His Ser Glu Ala Lys Met Arg Leu Lys Ala Ala Leu Thr His His Pro 290 295 300 Ala Ala Met Ser Asn Gly Asn Met Asn Thr Met Gly His Met Met Glu 305 310 315 320 Met Met Gly Ser Arg Gln Asp Gln Thr Pro His His His Met His Ser 325 330 335 His Pro His Gln His Gln Thr Leu Pro Pro His His Pro Tyr Pro His 340 345 350 Gln His Gln His Pro Ala His His Pr o His Pro Gln Pro His His Gln 355 360 365 Gln Asn His Pro His His His Ser His Ser His Leu His Ala His Pro 370 375 380 Ala His His Gln Thr Ser Pro His Pro Leu His Thr Gly Asn Gln 385 390 395 400 Ala Gln Val Ser Pro Ala Thr Gln Gln Met Gln Pro Thr Gln Thr Ile 405 410 415 Gln Pro Pro Gln Pro Thr Gly Gly Arg Arg Arg Arg Val Val Asp Glu 420 425 430 Asp Pro Asp Glu Arg Arg Arg Lys Phe Leu Glu Arg Asn Arg Ala Ala 435 440 445 Ala Thr Arg Cys Arg Gln Lys Arg Lys Val Trp Val Met Ser Leu Glu 450 455 460 Lys Lys Ala Glu Glu Leu Thr Gln Thr Asn Met Gln Leu Gln Asn Glu 465 470 475 480 Val Ser Met Leu Lys Asn Glu Val Ala Gln Leu Lys Gln Leu Leu Leu 485 490 495 Thr His Lys Asp Cys Pr o Ile Thr Ala Met Gln Lys Glu Ser Gln Gly 500 505 510 Tyr Leu Ser Pro Glu Ser Ser Pro Pro Ala Ser Pro Val Pro Ala Cys 515 520 525 Ser Gln Gln Gln Val Ile Gln His Asn Thr Ile Thr Thr Ser Ser Ser 530 535 540 Val Ser Glu Val Val Gly Ser Ser Thr Leu Ser Gln Leu Thr Thr His 545 550 555 560 Arg Thr Asp Leu Asn Pro Ile Leu 565 <210> 6 <211> 547 <212> PRT <213> Homo sapiens <400 > 6 Met Ala Thr Met Val Pro Ser Val Leu Trp Pro Arg Ala Cys Trp Thr 1 5 10 15 Leu Leu Val Cys Cys Leu Leu Thr Pro Gly Val Gln Gly Gln Glu Phe 20 25 30 Leu Leu Arg Val Glu Pro Gln Asn Pro Val Leu Ser Ala Gly Gly Ser 35 40 45 Leu Phe Val Asn Cys Ser Thr Asp Cys Pro Ser Ser Glu Lys Ile Ala 50 55 60 Leu Glu Thr Ser Leu Ser Lys Glu Leu Val Ala Ser Gly Met Gly Trp 65 70 75 80 Ala Ala Phe Asn Leu Ser Asn Val Thr Gly Asn Ser Arg Ile Leu Cys 85 90 95 Ser Val Tyr Cys Asn Gly Ser Gln Ile Thr Gly Ser Ser Asn Ile Thr 100 105 110 Val Tyr Arg Leu Pro Glu Arg Val Glu Leu Ala Pro Leu Pro Pro Trp 115 120 125 Gln Pro Val Gly Gln Asn Phe Thr Leu Arg Cys Gln Val Glu Asp Gly 130 135 140 Ser Pro Arg Thr Ser Leu Thr Val Val Leu Leu Arg Trp Glu Glu Glu 145 150 155 160 Leu Ser Arg Gln Pro Ala Val Glu Glu Pro Ala Glu Val Thr Ala Thr 165 170 175 Val Leu Ala Ser Arg Asp Asp His Gly Ala Pro Phe Ser Cys Arg Thr 180 185 190 Glu Leu Asp Met Gln Pro Gln Gly Leu Gly Leu Phe Val Asn Thr Ser 195 200 205 Ala Pro Arg Gln Leu Arg Thr Phe Val Leu Pro Val Thr Pro Pro Arg 210 215 220 Leu Val Ala Pro Arg Phe Leu Glu Val Glu Thr Ser Trp Pro Val Asp 225 230 235 240 Cys Thr Leu Asp Gly Leu Phe Pro Ala Ser Glu Ala Gln Val Tyr Leu 245 250 255 Ala Leu Gly Asp Gln Met Leu Asn Ala Thr Val Met Asn His Gly Asp 260 265 270 Thr Leu Thr Ala Thr Ala Thr Ala Thr Ala Arg Ala Asp Gln Glu Gly 275 280 285 Ala Arg Glu Ile Val Cys Asn Val Thr Leu Gly Gly Glu Arg Arg Glu 290 295 300 Ala Arg Glu Asn Leu Thr Val Phe Ser Phe Leu Gly Pro Ile Val Asn 305 310 315 320 Leu Ser Glu Pro Thr Ala His Glu Gly Ser Thr Val Thr Val Ser Cys 325 330 335 Met Ala Gly Ala Arg Val Gln Val Thr Leu Asp Gly Val Pro Ala Ala 340 345 350 Ala Pro Gly Gln Pro Ala Gln Leu Gln Leu Asn Ala Thr Glu Ser Asp 355 360 365 Asp Gly Arg Ser Phe Phe Cys Ser Ala Thr Leu Glu Val Asp Gly Glu 370 375 380 Phe Leu His Arg Asn Ser Ser Val Gln Leu Arg Val Leu Tyr Gly Pro 385 390 395 400 Lys Ile Asp Arg Ala Thr Cys Pro Gln His Leu Lys Trp Lys Asp Lys 405 410 415 Thr Arg His Val Leu Gln Cys Gln Ala Arg Gly Asn Pro Tyr Pro Glu 420 425 430 Leu Arg Cys Leu Lys Glu Gly Ser Ser Arg Glu Val Pro Val Gly Ile 435 440 445 Pro Phe Phe Val Asn Val Thr His Asn Gly Thr Tyr Gln Cys Gln Ala 450 455 460 Ser Ser Ser Arg Gly Lys Tyr Thr Leu Val Val Val Met Asp Ile Glu 465 470 475 480 Ala Gly Ser Ser His Phe Val Pro Val Phe Val Ala Val Leu Leu Thr 485 490 495 Leu Gly Val Val Thr Ile Val Leu Ala Leu Met Tyr Val Phe Arg Glu 500 505 510 His Gln Arg Ser Gly Ser Tyr His Val Arg Glu Glu Ser Thr Tyr Leu 515 520 525 Pro Leu Thr Ser Met Gln Pro Thr Glu Ala Met Gly Glu Glu Pro Ser 530 535 540 Arg Ala Glu 545 <210> 7 <211> 102 <212> PRT <213> Homo sapiens <400> 7 Met Ala Asp Asp Val Asp Gln Gln Gln Thr Thr Asn Thr Val Glu Glu 1 5 10 15 Pro Leu Asp Leu Ile Arg Leu Ser Leu Asp Glu Arg Ile Tyr Val Lys 20 25 30 Met Arg Asn Asp Arg Glu Leu Arg Gly Arg Leu His Ala Tyr Asp Gln 35 40 45 His Leu Asn Met Ile Leu Gly Asp Val Glu Glu Thr Val Thr Thr Ile 50 55 60 Glu Ile Asp Glu Glu Thr Tyr Glu Glu Ile Tyr Lys Ser Thr Lys Arg 65 70 75 80 Asn Ile Pro Met Leu Phe Val Arg Gly Asp Gly Val Val Leu Val Ala 85 90 95 Pro Pro Leu Arg Val Gly 100 <210> 8 <211> 90 <212> PRT <213> Homo sapiens <400> 8 Met Ala Cys Pro Leu Asp Gln Ala Ile Gly Leu Leu Val Ala Ile Phe 1 5 10 15 His Lys Tyr Ser Gly Arg Glu Gly Asp Lys His Thr Leu Ser Lys Lys 20 25 30 Glu Leu Lys Glu Leu Ile Gln Lys Glu Leu Thr Ile Gly Ser Lys Leu 35 40 45 Gln Asp Ala Glu Ile Ala Arg Leu Met Glu Asp Leu Asp Arg Asn Lys 50 55 60 Asp Gln Glu Val Asn Phe Gln Glu Tyr Val Thr Phe Leu Gly Ala Leu 65 70 75 80Ala Leu Ile Tyr Asn Glu Ala Leu Lys Gly 85 90

Claims (14)

An antibody or aptamer that specifically binds to the ADGRG3 (adhesion G protein-coupled receptor G3) protein represented by the amino acid sequence of SEQ ID NO: 1, or a primer, probe or antisense nucleotide that specifically binds to a gene encoding the protein A kit for diagnosing ischemic disease or prognosis comprising a.
The method of claim 1,
AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and An antibody or aptamer that specifically binds to any one protein selected from the group consisting of mRNA degradation associated) and S100A6 (S100 calcium binding protein A6), or a primer, probe or A kit for diagnosing ischemic disease or prognosis, characterized in that it further comprises an antisense nucleotide.
The method of claim 1,
The kit consists of a RT-PCR kit, a PCR kit, a nucleic acid amplification kit, a DNA chip kit, an ELISA kit, a protein chip kit, a rapid kit, a multiple reaction monitoring (MRM) kit, and a lateral flow analysis (LFA) kit. A kit for diagnosing ischemic disease or prognosis, characterized in that it is any one selected from
The method of claim 1,
The ischemic disease is cerebral infarction, cerebral ischemia, cerebral hypoxia, ischemic heart disease, heart failure, myocardial ischemia, ischemic renal failure, skeletal muscle ischemia, intestinal ischemia, sleep apnea, hypoxia during sleep, focal perfusion defect, retinal ischemia, neonatal hypoxia, fetal hypoxia, thrombosis , Altitude sickness, carbon monoxide poisoning, ischemic disease diagnosis or prognosis diagnosis kit, characterized in that any one selected from the group consisting of sudden infant death syndrome and congenital circulatory disorders.
An antibody or aptamer that specifically binds to the ADGRG3 (adhesion G protein-coupled receptor G3) protein represented by the amino acid sequence of SEQ ID NO: 1, or a primer, probe or antisense nucleotide that specifically binds to a gene encoding the protein A composition for diagnosing ischemic disease or prognosis comprising a.
6. The method of claim 5,
AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and An antibody or aptamer that specifically binds to any one protein selected from the group consisting of mRNA degradation associated) and S100A6 (S100 calcium binding protein A6), or a primer, probe or A composition for diagnosis or prognosis of ischemic disease, characterized in that it further comprises an antisense nucleotide.
ADGRG3 (adhesion G protein-coupled receptor G3) protein represented by the amino acid sequence of SEQ ID NO: 1 in a biological sample isolated from a subject of interest, or an ischemic disease comprising the step of measuring the expression level of a gene encoding the protein A method of providing information for making a diagnosis or prognosis.
8. The method of claim 7,
AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and Any one protein selected from the group consisting of mRNA degradation associated) and S100A6 (S100 calcium binding protein A6), or ischemic disease, characterized in that it further comprises the step of measuring the expression level of a gene encoding the protein Methods of providing information for prognosis diagnosis.
8. The method of claim 7,
The biological sample is an information providing method for diagnosing or prognosing an ischemic disease, characterized in that it is selected from the group consisting of tissue, cells, blood, serum, plasma, saliva and urine.
8. The method of claim 7,
Methods for measuring the expression level of the protein include protein chip analysis, immunoassay, ligand binding assay, MALDITOF (Matrix Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry) analysis, SELDITOF (Sulface Enhanced Laser Desorption/Ionization Time of Flight Mass) analysis. Spectrometry analysis, radioimmunoassay, radioimmunodiffusion method, Oukteroni immunodiffusion method, rocket immunoelectrophoresis, tissue immunostaining, complement fixation assay, two-dimensional electrophoresis analysis, liquid chromatography-Mass Spectrometry , LC-MS), LCMS/MS (liquid chromatography-Mass Spectrometry/ Mass Spectrometry), Western blotting, and ELISA (enzyme linked immunosorbent assay) information for diagnosing or prognosing an ischemic disease, characterized in that it is selected from the group consisting of How to provide.
8. The method of claim 7,
Methods for measuring the expression level of the gene include reverse transcription polymerase reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RT-PCR), real-time reverse transcription polymerase reaction (Real-time RT-PCR), RNase protection assay ( An information providing method for diagnosing or prognosing an ischemic disease, characterized in that it is selected from the group consisting of RPA; RNase protection assay), Northern blotting, and a DNA chip.
treating the isolated biological sample with a candidate substance expected to prevent or treat ischemic disease; and
Measuring the expression level of an adhesion G protein-coupled receptor G3 (ADGRG3) protein, or a gene encoding the protein, represented by the amino acid sequence of SEQ ID NO: 1 in the biological sample treated with the candidate substance; A method of screening a drug for the prevention or treatment of
13. The method of claim 12,
AATK (apoptosis associated tyrosine kinase), ALOX5 (arachidonate 5-lipoxygenase), CD177 (CD177 molecule), CREB5 (cAMP responsive element binding protein 5), ICAM3 (intercellular adhesion molecule 3), LSM3 (LSM3 homolog, U6 small nuclear RNA and Ischemic characterized in that it further comprises the step of measuring the expression level of any one protein selected from the group consisting of mRNA degradation associated) and S100A6 (S100 calcium binding protein A6), or a gene encoding the protein. A method for screening a drug for preventing or treating a disease.
13. The method of claim 12,
The biological sample is a screening method for a drug for preventing or treating ischemic disease, characterized in that it is selected from the group consisting of tissue, cells, blood, serum, plasma, saliva and urine.

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