KR102355568B1 - A Biomarker for diagnosing Intestinal Bechet's disease - Google Patents

Abstract

A biomarker of the present invention can predict the onset and possibility of disease based on comparison with a normal control group, and can very effectively differentiate and diagnose Crohn's disease and Behcet's enteritis, which are difficult to diagnose based on symptoms alone.

Description

A biomarker for diagnosis of Bechet's disease {A Biomarker for diagnosing Intestinal Bechet's disease}

The present invention relates to a biomarker for diagnosis of Behçet's enteritis.

Behcet's disease is a systemic vasculitis with a relatively high incidence in regions from the Mediterranean coast to Far East Asia, such as Korea, China, Japan, and Turkey, compared to the West. And it is a disease of unknown cause characterized by symptoms such as invasion of important organs such as the central nervous system. That is, it corresponds to systemic vasculitis that can occur anywhere blood flows, and shows various clinical symptoms and symptoms of severity. Early diagnosis of Behcet's disease is very important because Behcet's disease has various clinical manifestations and can cause serious complications and disability in some cases. Based on Turkey, which is known for having a high incidence of Behçet's disease, it costs about $3,300 per patient per year to treat general Behcet's disease, and about $5,000 for patients with Behcet's disease showing neurological symptoms. has been reported In particular, since it shows the highest disease activity in the 20s and 40s, it is a disease that causes a very large economic and social loss due to the occurrence of serious complications at a young age. In addition, about 42% of patients with Behçet's disease are unable to work most days of the year, so it is a disease that causes a greater social loss. Since Behcet's disease is a disease with a higher prevalence in Korea than in the West, direct and indirect medical expenses due to Behcet's disease are expected to be very large.

On the other hand, since there is currently no objective diagnostic biomarker that can distinguish between a patient with Behcet's disease and a healthy person, the diagnosis of Behcet's disease mainly relies on clinical symptoms. However, since Behcet's disease shows various clinical symptoms, diagnosis based on clinical symptoms shows low sensitivity and specificity. In addition, there is a problem that it takes a long time from the onset to the confirmation of Behcet's disease. In particular, Bechet's disease (Intestinal Bechet's disease) and Crohn's disease (Crohn's disease) have similar symptoms, and endoscopic and histological findings are also similar, making it difficult to distinguish them. However, clinical pathological findings, experimental methods, or histological diagnostic tests for differentiating such diseases have not yet been studied.

In order to overcome this limitation, it is very important to invent an objective diagnostic biomarker. If an objective diagnostic biomarker that can diagnose Behçet's disease, particularly Behcet's enteritis, can be discovered, by diagnosing such a disease at an early stage, the time taken for confirmation can be reduced, and treatment tailored to the level of the disease can be made possible, resulting in symptoms Exacerbations and expensive unnecessary treatments can be avoided. In addition, better treatment outcomes can be achieved by providing accurate information on disease-related prognosis.

One object of the present invention is to provide a biomarker for diagnosing Behcet's disease.

Another object of the present invention is a composition for diagnosis of Behcet's disease; And to provide a kit comprising the same.

Another object of the present invention is to provide an information providing method for diagnosing Behcet's disease.

However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problems, and other problems not mentioned will be clearly understood by those of ordinary skill in the art from the following description.

One embodiment of the present invention provides a biomarker for diagnosing Behcet's disease.

In one embodiment of the present invention, there is provided a biomarker for diagnosis of Behçet's enteritis.

The biomarker of the present invention is a maltase-glucoamylase (maltase-glucoamylase; MGAM) protein; or a gene encoding it.

The "maltase-glucoamylase (MGAM)" of the present invention is a digestive enzyme present in the intestine, and corresponds to an alpha-glucosidase digestive enzyme. These proteins are composed of two subunits with different substrate specificities. The N-terminal catalytic domain has high activity towards maltose, and the C-terminal domain has broader substrate specificity and activity towards glucose oligomers. In the gut, these proteins act as enzymes to work synergistically with sucrase-isomaltase and alpha-amylase to digest dietary starch.

The maltase-glucoamylase protein of the present invention may consist of the amino acid sequence shown in SEQ ID NO: 1.

The "biomarker" of the present invention generally refers to a protein or a gene encoding the protein present in various biological samples, and refers to an indicator capable of representing various changes or the onset of diseases that may occur in an individual. For the purpose of the present invention, when the expression level of the biomarker is increased compared to a normal control or Crohn's disease patient, the onset or likelihood of Behçet's disease, for example, Behçet's enteritis, can be represented with very high accuracy.

In one embodiment of the present invention, the maltase-glucoamylase (maltase-glucoamylase; MGAM) protein; Or, when the expression level of the gene encoding it is increased compared to a normal control group or a patient with Crohn's disease, Behcet's disease, for example, Behçet's enteritis, has already occurred or it can be expected that the incidence is high.

The "diagnosis" of the present invention means identifying the presence or characteristics of a pathological condition. For the purpose of the present invention, the diagnosis is to predict whether or not the onset of Behçet's disease, for example, Behcet's enteritis, or the possibility of its onset, and whether or not the disease occurs through the protein of the present invention or a gene encoding the same. can be predicted very effectively.

The "Behcet's disease" of the present invention is a systemic vasculitis, and is a disease of unknown cause characterized by symptoms such as ulcers and arthritis in mucous membranes such as the oral cavity, genitals or intestines, and invasion of important organs such as blood vessels and central nervous system.

The Behcet's disease of the present invention may be Behçet's enteritis, but is not limited thereto. Ulcers caused by Behçet's enteritis do not respond well to treatment, unlike oral ulcers, and intestinal bleeding or perforation may be combined. Therefore, it is very important to diagnose Behçet's enteritis early and distinguish it from other inflammatory bowel diseases. For the purpose of the present invention, the biomarker can be differentiated and diagnosed by clarifying the distinction from various other diseases corresponding to inflammatory bowel disease having similar clinical symptoms, in particular, Crohn's disease.

Another embodiment of the present invention provides a composition for diagnosing Behcet's disease.

In one embodiment of the present invention, there is provided a composition for diagnosis of Behçet's enteritis.

In the diagnostic composition of the present invention, the content related to the Behcet's disease, Behçet's enteritis, maltase-glucoamylase protein, etc. are the same as those described in the biomarker, and thus are omitted to avoid excessive complexity of the specification.

The agent capable of measuring the expression level of the protein of the present invention is not limited as long as it is an agent capable of measuring the expression level of the protein by specifically binding to the protein. For example, the agent capable of measuring the expression level of the protein is at least selected from the group consisting of an antibody, oligopeptide, ligand, PNA (peptide nucleic acid) and aptamer that specifically binds to the protein. may be one.

The "antibody" of the present invention 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 said 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. For example, the polyclonal antibody can be produced by a method well known in the art, including the process of injecting an antigen of the protein into an animal and collecting blood from the animal to obtain a serum containing the antibody. Such polyclonal antibodies can be prepared from any animal such as goat, rabbit, sheep, monkey, horse, pig, cow, dog, and the like. In addition, monoclonal antibodies can be prepared using the hybridoma method well known in the art (see Kohler and Milstein (1976) European Journal of Immunology 6:511-519), or the phage antibody library technology (Clackson et al, Nature, 352). :624-628, 1991; Marks et al, J. Mol. Biol., 222:58, 1-597, 1991). The antibody prepared by the above method may be separated and purified using methods such as gel electrophoresis, dialysis, salt precipitation, ion exchange chromatography, and affinity chromatography. In addition, the antibody of the present invention includes a complete form having two full-length light chains and two full-length heavy chains, as well as functional fragments of the antibody.

The functional fragment of the antibody of the present invention means a fragment having at least an antigen-binding function, and includes Fab, F(ab'), F(ab')2 and Fv.

In the present invention, the "PNA (Peptide Nucleic Acid)" refers to an artificially synthesized polymer similar to DNA or RNA, and was first introduced by Professors Nielsen, Egholm, Berg and Buchardt of the University of Copenhagen, Denmark in 1991. Whereas DNA has a phosphate-ribose sugar backbone, PNA has a repeated N-(2-aminoethyl)-glycine backbone linked by peptide bonds, which greatly increases binding strength and stability to DNA or RNA, resulting in molecular biology , diagnostic assays 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" of the present invention is an oligonucleic acid or 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 cell-cycle inhibitor isolated from a combinatorial library". Proc Natl Acad Sci USA. 95(24): 142727 (1998).

The antibody, PNA and aptamer of the present invention can be easily prepared by those skilled in the art based on the amino acid sequence of the protein of the present invention.

The agent capable of measuring the expression level of the gene of the present invention is not limited as long as it can measure the expression level by complementary binding to the gene of the present invention. For example, the agent capable of measuring the expression level of the gene may be at least one selected from the group consisting of a primer, a probe, and an antisense nucleotide complementary to the gene.

The "primer" of the present invention is a fragment recognizing a target gene sequence, and includes a pair of forward and reverse primers, but preferably, a primer pair that provides analysis results having specificity and sensitivity. High specificity can be conferred when the primer's nucleic acid sequence is a sequence that is inconsistent with a non-target sequence present in the sample, thus amplifying only the target gene sequence containing the complementary primer binding site and not causing non-specific amplification. .

The "probe" of the present invention means a substance capable of complementary binding to a target substance to be detected in a sample, and means a substance capable of specifically confirming the presence of a 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, and most preferably It is PNA. More specifically, the probe is a biomaterial derived from or similar thereto, or manufactured in vitro, and includes, for example, enzymes, proteins, antibodies, microorganisms, animal and plant cells and organs, neurons, DNA, and It may be RNA, and DNA includes cDNA, genomic DNA, and oligonucleotides, and RNA includes genomic RNA, mRNA, and oligonucleotides.

The "LNA (Locked nucleic acids)" of the present invention means a nucleic acid analog comprising 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 an ideal shape at the Watson-Crick bond. When LNA is incorporated into DNA or RNA oligonucleotides, LNA can pair with complementary nucleotide chains more rapidly, increasing the stability of the double helix.

The "antisense" of the present invention means that the antisense oligomer is hybridized with a target sequence in RNA by Watson-Crick base pairing, and typically mRNA and RNA: a sequence of nucleotide bases allowing the formation of an oligomeric heteroduplex in the target sequence. and oligomers having an inter-subunit backbone. An oligomer may have exact sequence complementarity or approximate complementarity to a target sequence.

Since the agent capable of measuring the expression level of the gene of the present invention can derive the nucleotide sequence of the gene based on the amino acid sequence of the protein of the present invention, those skilled in the art can complement the gene based on this. A primer, a probe, etc. that bind can be easily prepared.

In the diagnostic composition of the present invention, the maltase-glucoamylase protein; Alternatively, when the expression level of the gene encoding it is increased compared to a normal control group or a Crohn's disease patient, Behcet's disease, for example, Behçet's enteritis, has already occurred or it can be expected that the incidence is high.

Another embodiment of the present invention provides a kit for diagnosing Behcet's disease.

One embodiment of the present invention provides a kit for diagnosis of Behçet's enteritis.

The diagnostic kit of the present invention includes the composition for diagnosis of Behcet's disease of the present invention.

In the diagnostic kit of the present invention, the content related to the agent capable of measuring the expression level of Behçet's disease, Behçet's enteritis, maltase-glucoamylase protein, protein or gene is the same as described in the biomarker and diagnostic composition, is omitted to avoid excessive complexity.

The kit of the present invention may be an RT-PCR kit, a DNA chip kit, an ELISA kit, a protein chip kit, a rapid kit, or a multiple reaction monitoring (MRM) kit, but is not limited thereto.

The kit of the present invention may further include one or more other component compositions, solutions or devices suitable for the assay method. For example, the 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 the gene encoding the 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 complementary to the nucleic acid sequence of the control gene. Other reverse transcription polymerase reaction kits include test tubes or other suitable containers, reaction buffers (with 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.

The diagnostic kit 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.

The diagnostic kit 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 their substrates or capable of binding the antibody. other materials and the like.

Another embodiment of the present invention provides a method for providing information for diagnosing Behcet's disease.

In one embodiment of the present invention, there is provided a method for providing information for diagnosing Behçet's enteritis.

In the method for providing information of the present invention, in a biological sample isolated from a subject, maltase-glucoamylase protein; or measuring the expression level of a gene encoding it.

In the information providing method of the present invention, the contents related to the Behcet's disease, Behcet's enteritis, maltase-glucoamylase protein, etc. are the same as those described in the biomarker and the diagnostic composition, and thus are omitted to avoid excessive complexity of the specification.

In the information providing method of the present invention, the maltase-glucoamylase protein; Or, when the expression level of the gene encoding it is increased compared to the normal control, it can be predicted that the likelihood of Behcet's disease, for example, Behçet's enteritis, is high.

For the purposes of the present invention said protein; Or, when the expression level of the gene encoding it is high compared to the expression level of a Crohn's disease patient, Behcet's disease, for example, Behçet's enteritis, has already occurred, or it can be predicted that the onset of the gene is high.

In the information providing method of the present invention, when the onset or high probability of Behcet's disease, for example, Behcet's enteritis, is predicted, appropriate treatment such as administering a drug for the disease to the target subject It may further include the step of performing.

In the present invention, the "target individual" refers to an individual whose onset of Behcet's disease, for example, Behcet's enteritis, is uncertain, and an individual who has the disease or has a high probability of developing the disease.

The "biological sample" of the present invention refers to any material, biological fluid, tissue or cell obtained from or derived from an individual, for example, whole blood, leukocytes, peripheral blood mononuclear blood, sputum, tears, mucus, nasal washes, including peripheral blood mononuclear cells, buffy coat, plasma and serum ), nasal aspirate, breath, urine, semen, saliva, peritoneal washings, pelvic fluids, cystic fluid, meningeal fluid, amniotic fluid, glandular fluid, pancreatic fluid, lymph fluid, pleural fluid, nipple aspirate, bronchial aspiration It may include bronchial aspirate, synovial fluid, joint aspirate, organ secretions, cell, cell extract or cerebrospinal fluid, However, the present invention is not limited thereto.

The agent capable of measuring the expression level of the protein of the present invention is at least one selected from the group consisting of an antibody, oligopeptide, ligand, PNA (peptide nucleic acid) and aptamer that specifically binds to the protein. may be

The agent capable of measuring the expression level of the gene of the present invention may be at least one selected from the group consisting of primers, probes and antisense nucleotides that complementarily bind to the gene.

The method of measuring the expression level of the protein of the present invention is a protein chip analysis, immunoassay, ligand binding assay, MALDI-TOF (Matrix Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry) analysis, SELDI-TOF (Sulface Enhanced Laser) Desorption/Ionization Time of Flight Mass Spectrometry) analysis, radioimmunoassay, radioimmunodiffusion method, Oukteroni immunodiffusion method, rocket immunoelectrophoresis, tissue immunostaining, complement fixation assay, 2D electrophoresis analysis, liquid chromatography-mass Analysis (liquid chromatography-Mass Spectrometry, LC-MS), LC-MS/MS (liquid chromatography-Mass Spectrometry/ Mass Spectrometry), Western blotting and ELISA (enzyme linked immunosorbent assay) may be at least one selected from the group consisting of have.

The method of measuring the expression level of the gene of the present invention is 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 It may be at least one selected from the group consisting of a protection assay (RPA; RNase protection assay), Northern blotting, and a DNA chip.

[Sequence List]

SEQ ID NO: 1: MGAM protein amino acid sequence

MARKKLKKFTTLEIVLSVLLLVLFIISIVLIVLLAKESLKSTAPDPGTTGTPDPGTTGTP

DPGTTGTTHARTTGPPDPGTTGTTPVSAECPVVNELERINCIPDQPPTKATCDQRGCCWN

PQGAVSVPWCYYSKNHSYHVEGNLVNTNAGFTARLKNLPSSPVFGSNVDNVLLTAEYQTS

NRFHFKLTDQTNNRFEVPHEHVQSFSGNAAASLTYQVEISRQPFSIKVTRRSNNRVLFDS

SIGPLLFADQFLQLSTRLPSTNVYGLGEHVHQQYRHDMNWKTWPIFNRDTTPNGNGTNLY

GAQTFFLCLEDASGLSFGVFLMNSNAMEVVLQPAPAITYRTIGGILDFYVFLGNTPEQVV

QEYLELIGRPALPSYWALGFHLSRYEYGTLDNMREVVERNRAAQLPYDVQHADIDYMDER

RDFTYDSVDFKGFPEFVNELHNNGQKLVIIVDPAISNNSSSSKPYGPYDRGSDMKIWVNS

SDGVTPLIGEVWPGQTVFPDYTNPNCAVWWTKEFELFHNQVEFDGIWIDMNEVSNFVDGS

VSGCSTNNLNNPPFTPRILDGYLFCKTLCMDAVQHWGKQYDIHNLYGYSMAVATAEAAKT

VFPNKRSFILTRSTFAGSGKFAAHWLGDNTATWDDLRWSIPGVLEFNKLFGIPMVGPDICG

FALDTPEELCRRWMQLGAFYPFSRNHNGQGYKDQDPASFGADSLLLNSSRHYLNIRYTLL

PYLYTLFFRAHSRGDTVARPLLHEFYEDNSTWDVHQQFLWGPGLLITPVLDEGAEKVMAY

VPDAVWYDYETGSQVRWRKQKVEMELPGDKIGLHLRGGYIFPTQQPNTTTLASRKNPLGL

IIALDENKEAKGELFWDNGETKDTVANKVYLLCEFSVTQNRLEVNISQSTYKDPNNLAFN

EIKILGTEEPSNVTVKHNGVPSQTSPTVTYDSNLKVAIITDIDLLLGEAYTVEWSIKIRD

EEKIDCYPDENGASAENCTARGCIWEASNSSGVPFCYFVNDLYSVSDVQYNSHGATADIS

LKSSVYANAFPSTPVNPLRLDVTYHKNEMLQFKIYDPNKNRYEVPVPLNIPSMPSSTPEG

QLYDVLIKKNPFGIEIRRKSTGTIIWDSQLLGFTFSDMFIRISTRLPSKYLYGFGETEHR

SYRRDLEWHTWGMFSRDQPPGYKKNSYGVHPYYMGLEEDGSAHGVLLLNSNAMDVTFQPL

PALTYRTTGGVLDFYVFLGPTPELVTQQYTELIGRPVMVPYWSLGFQLCRYGYQNDSEIA

SLYDEMVAAQIPYDVQYSDIDYMERQLDFTLSPKFAGFPALINRMKADGMRVILILDPAI

SGNETQPYPAFTRGVEDDVFIKYPNDGDIVWGKVWPDFPDVVVNGSLDWDSQVELYRAYV

AFPDFFRNSTAKWWKREIEELYNNPQNPERSLKFDGMWIDMNEPSSFVNGAVSPGCRDAS

LNHPPYMPHLESRDRGLSSKTLCMESQQILPDGSLVQHYNVHNLYGWSQTRPTYEAVQEV

TGQRGVVITRSTFPSSGRWAGHWLGDNTAAWDQLKKSIIGMMEFSLFGISYTGADICGFF

QDAEYEMCVRWMQLGAFYPFSRNHNTIGTRRQDPVSWDVAFVNISRTVLQTRYTLLPYLY

TLMHKAHTEGVTVVRPLLHEFVSDQVTWDIDSQFLLGPAFLVSPVLERNARNVTAYFPRA

RWYDYYTGVDINARGEWKTLPAPLDHINLHVRGGYILPWQEPALNTHLSRQKFMGFKIAL

DDEGTAGGWLFWDDGQSIDTYGKGLYYLASFSASQNTMQSHIIFNNYITGTNPLKLGYIE

IWGVGSVPVTSVSISVSGMVITPSFNNDPTTQVLSIDVTDRNISLHNFTSLTWISTL

The biomarker of the present invention can predict the onset and possibility of the disease compared to the normal control group, and can very effectively distinguish and diagnose Crohn's disease and Behçet's disease, for example, Behcet's enteritis, which are difficult to diagnose by distinguishing only the symptoms. can

1 shows the results of confirming the expression level of the biomarker according to the present invention in the intestinal mucosal tissue according to an embodiment of the present invention through immunostaining.
2 and 3 show the results of confirming the expression level of the biomarker according to the present invention in serum according to an embodiment of the present invention through ELISA.

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 skilled 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

[Example 1] Confirmation of biomarker expression level in intestinal mucosal tissue

A colonoscopy-guided biopsy was performed on 7 patients with Bechet's disease or 7 patients with Crohn's disease (CD) being treated at the inflammatory bowel disease clinic at Yonsei University Severance Hospital (Seoul, Sinchon). Colonic mucosal tissue was collected through the

The tissue obtained from the Behcet's disease or Crohn's disease patient was fixed with 10% formalin, embedded in paraffin, and a 7 μm thick section was attached to a slide. Then, the sections were deparaffinized using xylene and then treated with ethanol in order from high concentration to low concentration. Thereafter, immunostaining was performed using an antibody that specifically binds to maltase-glucoamylase, and the expression level of the proteins was measured using a microscope, and the results are shown in FIG. 1 .

1, when comparing Crohn's disease and Behcet's enteritis, which correspond to inflammatory bowel disease, maltase-glucoamylase ( It was confirmed that the expression level of MGAM) was significantly increased.

From the above results, it can be seen that when the biomarker according to the present invention is used, Crohn's disease and Behcet's disease, which are difficult to diagnose by distinguishing only by symptoms, can be distinguished and diagnosed very effectively.

[Example 2] Identification of biomarker expression levels in serum

As in Example 1, sera were obtained from 7 patients with Behçet's enteritis, 7 patients with Crohn's disease, or normal controls being treated at the inflammatory bowel disease clinic of Yonsei University Severance Hospital. The serum thus obtained was subjected to ELISA analysis using an antibody that specifically binds to maltase-glucoamylase by a conventional method, and the results are shown in FIGS. 2 and 3 .

As shown in FIGS. 2 and 3 , it was confirmed that the expression level of maltase-glucoamylase protein was significantly higher in patients with Behçet's enteritis (intestinal BD) compared to normal controls (HC) and patients with Crohn's disease (CD).

Through the above results, the maltase-glucoamylase according to the present invention can predict the onset and possibility of the disease compared to the normal control group, as well as other inflammatory bowel diseases such as Crohn's disease and Behcet's disease, which are difficult to diagnose based on symptoms alone. , in particular, it can be seen that Behcet's enteritis can be distinguished and diagnosed very effectively.

As described above in detail a specific part of the present invention, for those of ordinary skill in the art, this specific description is only a preferred embodiment, and it is clear that the scope of the present invention is not limited thereto. Accordingly, the substantial scope of the present invention will be defined by the appended claims and their equivalents.

<110> Industry-Academic Cooperation Foundation, Yonsei University <120> A Biomarker for diagnosing Intestinal Bechet's disease <130> PDPB204102 <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1857 <212> PRT <213> Homo sapiens <400> 1 Met Ala Arg Lys Lys Leu Lys Lys Phe Thr Thr Leu Glu Ile Val Leu 1 5 10 15 Ser Val Leu Leu Leu Val Leu Phe Ile Ile Ser Ile Val Leu Ile Val 20 25 30 Leu Leu Ala Lys Glu Ser Leu Lys Ser Thr Ala Pro Asp Pro Gly Thr 35 40 45 Thr Gly Thr Pro Asp Pro Gly Thr Thr Gly Thr Pro Asp Pro Gly Thr 50 55 60 Thr Gly Thr Thr His Ala Arg Thr Thr Gly Pro Pro Asp Pro Gly Thr 65 70 75 80 Thr Gly Thr Thr Pro Val Ser Ala Glu Cys Pro Val Val Asn Glu Leu 85 90 95 Glu Arg Ile Asn Cys Ile Pro Asp Gln Pro Pro Thr Lys Ala Thr Cys 100 105 110 Asp Gln Arg Gly Cys Cys Trp Asn Pro Gln Gly Ala Val Ser Val Pro 115 120 125 Trp Cys Tyr Tyr Ser Lys Asn His Ser Tyr His Val Glu Gly Asn Leu 130 135 140 Val Asn Thr Asn Ala Gly Phe Thr Ala Arg Leu Lys Asn Leu Pro Ser 145 150 155 160 Ser Pro Val Phe Gly Ser Asn Val Asp Asn Val Leu Leu Thr Ala Glu 165 170 175 Tyr Gln Thr Ser Asn Arg Phe His Phe Lys Leu Thr Asp Gln Thr Asn 180 185 190 Asn Arg Phe Glu Val Pro His Glu His Val Gln Ser Phe Ser Gly Asn 195 200 205 Ala Ala Ala Ser Leu Thr Tyr Gln Val Glu Ile Ser Arg Gln Pro Phe 210 215 220 Ser Ile Lys Val Thr Arg Arg Ser Asn Asn Arg Val Leu Phe Asp Ser 225 230 235 240 Ser Ile Gly Pro Leu Leu Phe Ala Asp Gln Phe Leu Gln Leu Ser Thr 245 250 255 Arg Leu Pro Ser Thr Asn Val Tyr Gly Leu Gly Glu His Val His Gln 260 265 270 Gln Tyr Arg His Asp Met Asn Trp Lys Thr Trp Pro Ile Phe Asn Arg 275 280 285 Asp Thr Thr Pro Asn Gly Asn Gly Thr Asn Leu Tyr Gly Ala Gln Thr 290 295 300 Phe Phe Leu Cys Leu Glu Asp Ala Ser Gly Leu Ser Phe Gly Val Phe 305 310 315 320 Leu Met Asn Ser Asn Ala Met Glu Val Val Leu Gln Pro Ala Pro Ala 325 330 335 Ile Thr Tyr Arg Thr Ile Gly Gly Ile Leu Asp Phe Tyr Val Phe Leu 340 345 350 Gly Asn Thr Pro Glu Gln Val Val Gln Glu Tyr Leu Glu Leu Ile Gly 355 360 365 Arg Pro Ala Leu Pro Ser Tyr Trp Ala Leu Gly Phe His Leu Ser Arg 370 375 380 Tyr Glu Tyr Gly Thr Leu Asp Asn Met Arg Glu Val Val Glu Arg Asn 385 390 395 400 Arg Ala Ala Gln Leu Pro Tyr Asp Val Gln His Ala Asp Ile Asp Tyr 405 410 415 Met Asp Glu Arg Arg Asp Phe Thr Tyr Asp Ser Val Asp Phe Lys Gly 420 425 430 Phe Pro Glu Phe Val Asn Glu Leu His Asn Asn Gly Gln Lys Leu Val 435 440 445 Ile Ile Val Asp Pro Ala Ile Ser Asn Asn Ser Ser Ser Ser Lys Pro 450 455 460 Tyr Gly Pro Tyr Asp Arg Gly Ser Asp Met Lys Ile Trp Val Asn Ser 465 470 475 480 Ser Asp Gly Val Thr Pro Leu Ile Gly Glu Val Trp Pro Gly Gln Thr 485 490 495 Val Phe Pro Asp Tyr Thr Asn Pro Asn Cys Ala Val Trp Trp Thr Lys 500 505 510 Glu Phe Glu Leu Phe His Asn Gln Val Glu Phe Asp Gly Ile Trp Ile 515 520 525 Asp Met Asn Glu Val Ser Asn Phe Val Asp Gly Ser Val Ser Gly Cys 530 535 540 Ser Thr Asn Asn Leu Asn Asn Pro Pro Phe Thr Pro Arg Ile Leu Asp 545 550 555 560 Gly Tyr Leu Phe Cys Lys Thr Leu Cys Met Asp Ala Val Gln His Trp 565 570 575 Gly Lys Gln Tyr Asp Ile His Asn Leu Tyr Gly Tyr Ser Met Ala Val 580 585 590 Ala Thr Ala Glu Ala Ala Lys Thr Val Phe Pro Asn Lys Arg Ser Phe 595 600 605 Ile Leu Thr Arg Ser Thr Phe Ala Gly Ser Gly Lys Phe Ala Ala His 610 615 620 Trp Leu Gly Asp Asn Thr Ala Thr Trp Asp Asp Leu Arg Trp Ser Ile 625 630 635 640 Pro Gly Val Leu Glu Phe Asn Leu Phe Gly Ile Pro Met Val Gly Pro 645 650 655 Asp Ile Cys Gly Phe Ala Leu Asp Thr Pro Glu Glu Leu Cys Arg Arg 660 665 670 Trp Met Gln Leu Gly Ala Phe Tyr Pro Phe Ser Arg Asn His Asn Gly 675 680 685 Gln Gly Tyr Lys Asp Gln Asp Pro Ala Ser Phe Gly Ala Asp Ser Leu 690 695 700 Leu Leu Asn Ser Ser Arg His Tyr Leu Asn Ile Arg Tyr Thr Leu Leu 705 710 715 720 Pro Tyr Leu Tyr Thr Leu Phe Phe Arg Ala His Ser Arg Gly Asp Thr 725 730 735 Val Ala Arg Pro Leu Leu His Glu Phe Tyr Glu Asp Asn Ser Thr Trp 740 745 750 Asp Val His Gln Gln Phe Leu Trp Gly Pro Gly Leu Leu Ile Thr Pro 755 760 765 Val Leu Asp Glu Gly Ala Glu Lys Val Met Ala Tyr Val Pro Asp Ala 770 775 780 Val Trp Tyr Asp Tyr Glu Thr Gly Ser Gln Val Arg Trp Arg Lys Gln 785 790 795 800 Lys Val Glu Met Glu Leu Pro Gly Asp Lys Ile Gly Leu His Leu Arg 805 810 815 Gly Gly Tyr Ile Phe Pro Thr Gln Gln Pro Asn Thr Thr Thr Leu Ala 820 825 830 Ser Arg Lys Asn Pro Leu Gly Leu Ile Ile Ala Leu Asp Glu Asn Lys 835 840 845 Glu Ala Lys Gly Glu Leu Phe Trp Asp Asn Gly Glu Thr Lys Asp Thr 850 855 860 Val Ala Asn Lys Val Tyr Leu Leu Cys Glu Phe Ser Val Thr Gln Asn 865 870 875 880 Arg Leu Glu Val Asn Ile Ser Gln Ser Thr Tyr Lys Asp Pro Asn Asn 885 890 895 Leu Ala Phe Asn Glu Ile Lys Ile Leu Gly Thr Glu Glu Pro Ser Asn 900 905 910 Val Thr Val Lys His Asn Gly Val Pro Ser Gin Thr Ser Pro Thr Val 915 920 925 Thr Tyr Asp Ser Asn Leu Lys Val Ala Ile Ile Thr Asp Ile Asp Leu 930 935 940 Leu Leu Gly Glu Ala Tyr Thr Val Glu Trp Ser Ile Lys Ile Arg Asp 945 950 955 960 Glu Glu Lys Ile Asp Cys Tyr Pro Asp Glu Asn Gly Ala Ser Ala Glu 965 970 975 Asn Cys Thr Ala Arg Gly Cys Ile Trp Glu Ala Ser Asn Ser Ser Gly 980 985 990 Val Pro Phe Cys Tyr Phe Val Asn Asp Leu Tyr Ser Val Ser Asp Val 995 1000 1005 Gln Tyr Asn Ser His Gly Ala Thr Ala Asp Ile Ser Leu Lys Ser Ser 1010 1015 1020 Val Tyr Ala Asn Ala Phe Pro Ser Thr Pro Val Asn Pro Leu Arg Leu 1025 1030 1035 1040 Asp Val Thr Tyr His Lys Asn Glu Met Leu Gln Phe Lys Ile Tyr Asp 1045 1050 1055 Pro Asn Lys Asn Arg Tyr Glu Val Pro Val Pro Leu Asn Ile Pro Ser 1060 1065 1070 Met Pro Ser Ser Thr Pro Glu Gly Gin Leu Tyr Asp Val Leu Ile Lys 1075 1080 1085 Lys Asn Pro Phe Gly Ile Glu Ile Arg Arg Lys Ser Thr Gly Thr Ile 1090 1095 1100 Ile Trp Asp Ser Gln Leu Leu Gly Phe Thr Phe Ser Asp Met Phe Ile 1105 1110 1115 1120 Arg Ile Ser Thr Arg Leu Pro Ser Lys Tyr Leu Tyr Gly Phe Gly Glu 1125 1130 1135 Thr Glu His Arg Ser Tyr Arg Arg Asp Leu Glu Trp His Thr Trp Gly 1140 1145 1150 Met Phe Ser Arg Asp Gln Pro Gly Tyr Lys Lys Asn Ser Tyr Gly 1155 1160 1165 Val His Pro Tyr Tyr Met Gly Leu Glu Glu Asp Gly Ser Ala His Gly 1170 1175 1180 Val Leu Leu Leu Asn Ser Asn Ala Met Asp Val Thr Phe Gln Pro Leu 1185 1190 1195 1200 Pro Ala Leu Thr Tyr Arg Thr Thr Gly Gly Val Leu Asp Phe Tyr Val 1205 1210 1215 Phe Leu Gly Pro Thr Pro Glu Leu Val Thr Gln Gln Tyr Thr Glu Leu 1220 1225 1230 Ile Gly Arg Pro Val Met Val Pro Tyr Trp Ser Leu Gly Phe Gln Leu 1235 1240 1245 Cys Arg Tyr Gly Tyr Gln Asn Asp Ser Glu Ile Ala Ser Leu Tyr Asp 1250 1255 1260 Glu Met Val Ala Ala Gln Ile Pro Tyr Asp Val Gln Tyr Ser Asp Ile 1265 1270 1275 1280 Asp Tyr Met Glu Arg Gln Leu Asp Phe Thr Leu Ser Pro Lys Phe Ala 1285 1290 1295 Gly Phe Pro Ala Leu Ile Asn Arg Met Lys Ala Asp Gly Met Arg Val 1300 1305 1310 Ile Leu Ile Leu Asp Pro Ala Ile Ser Gly Asn Glu Thr Gln Pro Tyr 1315 1320 1325 Pro Ala Phe Thr Arg Gly Val Glu Asp Asp Val Phe Ile Lys Tyr Pro 1330 1335 1340 Asn Asp Gly Asp Ile Val Trp Gly Lys Val Trp Pro Asp Phe Pro Asp 1345 1350 1355 1360 Val Val Val Asn Gly Ser Leu Asp Trp Asp Ser Gln Val Glu Leu Tyr 1365 1370 1375 Arg Ala Tyr Val Ala Phe Pro Asp Phe Phe Arg Asn Ser Thr Ala Lys 1380 1385 1390 Trp Trp Lys Arg Glu Ile Glu Glu Leu Tyr Asn Asn Pro Gln Asn Pro 1395 1400 1405 Glu Arg Ser Leu Lys Phe Asp Gly Met Trp Ile Asp Met Asn Glu Pro 1410 1415 1420 Ser Ser Phe Val Asn Gly Ala Val Ser Pro Gly Cys Arg Asp Ala Ser 1425 1430 1435 1440 Leu Asn His Pro Pro Tyr Met Pro His Leu Glu Ser Arg Asp Arg Gly 1445 1450 1455 Leu Ser Ser Lys Thr Leu Cys Met Glu Ser Gln Gln Ile Leu Pro Asp 1460 1465 1470 Gly Ser Leu Val Gln His Tyr Asn Val His Asn Leu Tyr Gly Trp Ser 1475 1480 1485 Gln Thr Arg Pro Thr Tyr Glu Ala Val Gln Glu Val Thr Gly Gln Arg 1490 1495 1500 Gly Val Val Ile Thr Arg Ser Thr Phe Pro Ser Ser Gly Arg Trp Ala 1505 1510 1515 1520 Gly His Trp Leu Gly Asp Asn Thr Ala Ala Trp Asp Gln Leu Lys Lys 1525 1530 1535 Ser Ile Ile Gly Met Met Glu Phe Ser Leu Phe Gly Ile Ser Tyr Thr 1540 1545 1550 Gly Ala Asp Ile Cys Gly Phe Phe Gln Asp Ala Glu Tyr Glu Met Cys 1555 1560 1565 Val Arg Trp Met Gln Leu Gly Ala Phe Tyr Pro Phe Ser Arg Asn His 1570 1575 1580 Asn Thr Ile Gly Thr Arg Arg Gln Asp Pro Val Ser Trp Asp Val Ala 1585 1590 1595 1600 Phe Val Asn Ile Ser Arg Thr Val Leu Gln Thr Arg Tyr Thr Leu Leu 1605 1610 1615 Pro Tyr Leu Tyr Thr Leu Met His Lys Ala His Thr Glu Gly Val Thr 1620 1625 1630 Val Val Arg Pro Leu Leu His Glu Phe Val Ser Asp Gln Val Thr Trp 1635 1640 1645 Asp Ile Asp Ser Gln Phe Leu Leu Gly Pro Ala Phe Leu Val Ser Pro 1650 1655 1660 Val Leu Glu Arg Asn Ala Arg Asn Val Thr Ala Tyr Phe Pro Arg Ala 1665 1670 1675 1680 Arg Trp Tyr Asp Tyr Tyr Thr Gly Val Asp Ile Asn Ala Arg Gly Glu 1685 1690 1695 Trp Lys Thr Leu Pro Ala Pro Leu Asp His Ile Asn Leu His Val Arg 1700 1705 1710 Gly Gly Tyr Ile Leu Pro Trp Gln Glu Pro Ala Leu Asn Thr His Leu 1715 1720 1725 Ser Arg Gln Lys Phe Met Gly Phe Lys Ile Ala Leu Asp Asp Glu Gly 1730 1735 1740 Thr Ala Gly Gly Trp Leu Phe Trp Asp Asp Gly Gln Ser Ile Asp Thr 1745 1750 1755 1760 Tyr Gly Lys Gly Leu Tyr Tyr Leu Ala Ser Phe Ser Ala Ser Gln Asn 1765 1770 1775 Thr Met Gln Ser His Ile Ile Phe Asn Asn Tyr Ile Thr Gly Thr Asn 1780 1785 1790 Pro Leu Lys Leu Gly Tyr Ile Glu Ile Trp Gly Val Gly Ser Val Pro 1795 1800 1805 Val Thr Ser Val Ser Ile Ser Val Ser Gly Met Val Ile Thr Pro Ser 1810 1815 1820 Phe Asn Asn Asp Pro Thr Thr Gln Val Leu Ser Ile Asp Val Thr Asp 1825 1830 1835 1840 Arg Asn Ile Ser Leu His Asn Phe Thr Ser Leu Thr Trp Ile Ser Thr 1845 1850 1855 Leu

Claims (13)

delete delete maltase-glucoamylase protein; Or, a composition for diagnosing Behcet's disease, comprising an agent capable of measuring the expression level of the gene encoding it. 4. The method of claim 3,
The Behcet's disease will be Behcet's enteritis, a diagnostic composition. 4. The method of claim 3,
The agent capable of measuring the expression level of the protein is at least one selected from the group consisting of an antibody, oligopeptide, ligand, PNA (peptide nucleic acid) and aptamer that specifically binds to the protein. , a diagnostic composition. 4. The method of claim 3,
The agent capable of measuring the expression level of the gene is at least one selected from the group consisting of primers, probes and antisense nucleotides that complementarily bind to the gene, diagnostic composition. A diagnostic kit for Behçet's disease comprising the diagnostic composition of any one of claims 3 to 6. 8. The method of claim 7,
The Behcet's disease is Behcet's enteritis, a diagnostic kit. In a biological sample isolated from a subject of interest, maltase-glucoamylase protein; Or comprising the step of measuring the expression level of the gene encoding the same, information providing method for diagnosing Behcet's disease. 10. The method of claim 9,
Methods for measuring the expression level of the protein include protein chip analysis, immunoassay, ligand binding assay, MALDI-TOF (Matrix Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry) analysis, SELDI-TOF (Sulface Enhanced Laser Desorption/Ionization) assay. Time of Flight Mass Spectrometry analysis, radioimmunoassay, radioimmunodiffusion method, octeroni immunodiffusion method, rocket immunoelectrophoresis, tissue immunostaining, complement fixation assay, two-dimensional electrophoresis analysis, liquid chromatography-mass spectrometry (liquid) At least one selected from the group consisting of chromatography-Mass Spectrometry, LC-MS, LC-MS/MS (liquid chromatography-Mass Spectrometry/ Mass Spectrometry), Western blotting, and ELISA (enzyme linked immunosorbent assay), an information providing method. 10. The method of claim 9,
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 ( RPA; RNase protection assay), Northern blotting (Northern blotting) and at least one selected from the group consisting of a DNA chip, information providing method. 10. The method of claim 9,
The Behcet's disease will be Behcet's enteritis, information providing method. 10. The method of claim 9,
The method for providing the information includes a maltase-glucoamylase protein; Or, when the expression level of the gene encoding it is increased compared to the normal control, it is predicted that the likelihood of developing Behcet's disease is high, the information providing method.

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