WO2014148429A1 - Biomarqueur pour le diagnostic d'une pneumonie interstitielle non spécifique - Google Patents

Biomarqueur pour le diagnostic d'une pneumonie interstitielle non spécifique Download PDF

Info

Publication number
WO2014148429A1
WO2014148429A1 PCT/JP2014/057128 JP2014057128W WO2014148429A1 WO 2014148429 A1 WO2014148429 A1 WO 2014148429A1 JP 2014057128 W JP2014057128 W JP 2014057128W WO 2014148429 A1 WO2014148429 A1 WO 2014148429A1
Authority
WO
WIPO (PCT)
Prior art keywords
antibody
hla
nsip
antigen
biomarker
Prior art date
Application number
PCT/JP2014/057128
Other languages
English (en)
Japanese (ja)
Inventor
博 木田
淳 熊ノ郷
尚 荒瀬
井上　義一
Original Assignee
国立大学法人大阪大学
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 国立大学法人大阪大学 filed Critical 国立大学法人大阪大学
Priority to JP2015506765A priority Critical patent/JP6315832B2/ja
Publication of WO2014148429A1 publication Critical patent/WO2014148429A1/fr

Links

Images

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6854—Immunoglobulins
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/12—Pulmonary diseases

Definitions

  • the present invention relates to a biomarker that can easily and highly accurately diagnose nonspecific interstitial pneumonia.
  • Interstitial pneumonia is a general term for diseases that cause inflammation mainly in the interstitium of the lung. There are various causes, including cases where the cause of drugs, smoking, etc. is clear, cases where the cause is associated with collagen disease, and cases where the cause cannot be identified. Interstitial pneumonia whose cause cannot be specified is collectively referred to as idiopathic interstitial pneumonia (IIPs). IIPs includes two diseases, idiopathic pulmonary fibrosis (IPF) / normal interstitial pneumonia (UIP) and nonspecific interstitial pneumonia (NSIP), which are classified according to prognosis and responsiveness to steroids. included.
  • IIPs idiopathic interstitial pneumonia
  • IPF idiopathic pulmonary fibrosis
  • UIP normal interstitial pneumonia
  • NSIP nonspecific interstitial pneumonia
  • IPF idiopathic pulmonary fibrosis
  • UIP normal interstitial pneumonia
  • NSIP nonspecific interstitial pneumonia
  • IPF is an abbreviation for “Idiopathic pulmonary fibrosis”
  • UIP is an abbreviation for “Usual interstitial pneumonia”.
  • IPF is used as a clinical diagnosis name
  • UIP is a name used as a histopathological diagnosis name
  • NSIP is an abbreviation of “Non-specific interstitial pneumonia” and is a name used in both clinical diagnosis and histopathological diagnosis.
  • UIP pattern and “NSIP pattern”.
  • IPF / UIP and NSIP in idiopathic interstitial pneumonia is an important index for predicting the prognosis of a patient.
  • IPF / UIP is regarded as a progressive poor prognosis disease, and a treatment plan is made after ascertaining the balance between long-term symptomatic treatment and drug therapy and side effects caused thereby.
  • NSIP has a good therapeutic response and generally has a good prognosis, so an aggressive treatment policy is adopted. Therefore, distinguishing between IPF / UIP and NSIP can be an important factor in determining the dosing schedule and treatment policy.
  • Non-Patent Document 2 A non-invasive examination method includes chest CT image examination and the like, but it is impossible to distinguish IPF / UIP and NSIP except for typical examples (see Non-Patent Document 3, for example).
  • KL-6 sialylated sugar chain antigen KL-6 (Sialylated carbohydrate antigen KL-6) is known as a biomarker.
  • KL-6 has high specificity in patients who have developed interstitial pneumonia, and correlates with the activity of interstitial pneumonia. Therefore, KL-6 is used as a highly clinically significant marker.
  • IPF / UIP and NSIP cannot be differentiated by this marker.
  • An object of the present invention is to provide a biomarker that enables simple and highly accurate diagnosis of nonspecific interstitial pneumonia. Furthermore, this invention aims at providing the test
  • the present inventors have intensively studied to solve the above-mentioned problems. Based on the strong nature of autoimmune diseases such as high antinuclear antibody values in NISP patients, IPF / UIP is included among autoantibodies. Whether or not there is an antibody that can be a biomarker that can be used to differentiate between two groups of NSIP and NSIP. As a result, it was found that the antibody titer of a specific autoantibody was increased in NSIP patients.
  • the present invention has been completed as a result of further studies based on such findings. That is, the present invention provides a biomarker, a test method, a test reagent, a diagnostic kit, and the like of the following modes.
  • Non-specific interstitial pneumonia comprising at least one antibody selected from the group consisting of anti-Mx1 antibody, anti-NINJ2 antibody, anti-DCX antibody, anti-C10orf49 antibody, anti-ZMAT4 antibody, anti-CDC42SE1 antibody and anti-GPT2 antibody ( NSIP) biomarker.
  • Item 2. The biomarker according to Item 1, wherein the antibody is at least one selected from the group consisting of an anti-Mx1 antibody and an anti-NINJ2 antibody.
  • Item 3. Item 3. The biomarker according to Item 1 or 2, wherein the antibody is a combination of an anti-Mx1 antibody and an anti-NINJ2 antibody.
  • a method for examining nonspecific interstitial pneumonia comprising: Item 5.
  • Item 5. The examination method according to Item 4, wherein the antibody is at least one selected from the group consisting of an anti-Mx1 antibody and an anti-NINJ2 antibody.
  • Item 6. The test method according to Item 4 or 5, wherein the antibody is a combination of an anti-Mx1 antibody and an anti-NINJ2 antibody.
  • Item 7. Item 7.
  • the detection of the antibody is carried out using at least one protein selected from the group consisting of Mx1, NINJ2, DCX, C10orf49, ZMAT4, CDC42SE1, and GPT2, or an antigen consisting of a partial peptide thereof. Inspection method according to crab.
  • Item 9. Item 9. The inspection method according to any one of Items 4 to 8, wherein the detection step is performed by an ELISA method.
  • At least one selected from the group consisting of anti-Mx1 antibody, anti-NINJ2 antibody, anti-DCX antibody, anti-C10orf49 antibody, anti-ZMAT4 antibody, anti-CDC42SE1 antibody and anti-GPT2 antibody in a blood sample collected from a test animal A step of detecting an antibody, and a step of diagnosing NSIP based on the detection result obtained in the step,
  • a reagent for testing nonspecific interstitial pneumonia comprising at least one protein selected from the group consisting of Mx1, NINJ2, DCX, C10orf49, ZMAT4, CDC42SE1 and GPT2 or an antigen consisting of these partial peptides .
  • Item 13 A diagnostic kit for nonspecific interstitial pneumonia (NSIP), comprising the test reagent according to Item 12.
  • NSIP that could only be diagnosed by a highly invasive method such as surgical lung biopsy so far can be diagnosed with high accuracy by a simpler and less invasive method. Can do.
  • a test reagent capable of easily detecting the biomarker and a diagnostic kit including the test reagent are provided.
  • the biomarker of the present invention is detected at a high concentration in the blood sample of NSIP patients, while it is not significantly different from the normal control group in UIP / IPF patients. Therefore, by detecting the biomarker of the present invention, it is possible to distinguish UIP / IPF and NSIP with high accuracy. There is a big difference in prognosis between UIP / IPF and NSIP, and NSIP patients generally respond well to therapeutic drugs and have a good prognosis. Therefore, when it is determined that the patient suffers from NSIP, an appropriate therapeutic agent can be selected and a dosing schedule can be planned.
  • A is a diagram schematically showing the principle of autoantibody (anti-Mx1 antibody) measurement using cells presenting an Mx1-HLA-DR complex.
  • B is a graph showing the results of measuring autoantibodies (anti-Mx1 antibodies) using cells presenting Mx1-HLA-DR complexes.
  • Biomarkers of nonspecific interstitial pneumonia (NSIP) biomarkers present invention anti Mx1 antibody, anti NINJ2 antibody, anti-DCX antibody, the group consisting of anti-C10orf49 antibody, anti ZMAT4 antibody, anti CDC42SE1 and anti GPT2 antibody It consists of at least one antibody selected from the above.
  • the biomarker of the present invention will be described in detail.
  • nonspecific interstitial pneumonia is used as a clinical diagnosis name in a narrow sense, and when used as a pathological diagnosis name, it is expressed as “NSIP pattern”. On the other hand, in some cases, it is simply expressed as “NSIP” and used without distinction in the meaning of both the clinical diagnosis name and the histopathological diagnosis name.
  • NSIP nonspecific interstitial pneumonia
  • the biomarker of the present invention comprises an antibody that specifically recognizes at least one of the antibodies shown in the following (a) to (g).
  • (A) Anti-Mx1 antibody The anti-Mx1 antibody is an antibody that specifically recognizes Mx1.
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza virus) resistance 1
  • Mx1 myxovirus (influenza
  • the anti-NINJ2 antibody is an antibody that specifically recognizes NINJ2.
  • NINJ2 (ninjurin 2) is one type of adhesion molecule, and its expression is enhanced in Schwann cells in the distal nerve after peripheral nerve injury, and it is considered that NINJ2 (ninjurin 2) may promote neurite outgrowth.
  • the specific amino acid sequence and base sequence of NINJ2 are registered in the NCBI database with the accession number NM_0166533.4.
  • DCX Neuronal migration protein doublecortin
  • NM_178152.1 The specific amino acid sequence and base sequence of DCX are registered in the NCBI database with the accession number NM_178152.1.
  • the anti-C10orf49 antibody is an antibody that specifically recognizes C10orf49.
  • C10orf49 chromosome 10 open reading frame 49
  • the specific amino acid sequence and base sequence of C10orf49 are registered in the NCBI database with the accession number NM_14514.1.
  • the anti-ZMAT4 antibody is an antibody that specifically recognizes ZMAT4.
  • the function of ZMAT4 (Zinc fingerer matrin-type protein 4) as a protein is unknown.
  • the specific amino acid sequence and base sequence of ZMAT4 are registered in the NCBI database with accession number BC019598.1.
  • the anti-CDC42SE1 antibody is an antibody that specifically recognizes CDC42SE1.
  • CDC42SE1 (CDC42 small effector 1) is a protein involved in accumulation of polymerized filamentous actin in T lymphocytes. The specific amino acid sequence and base sequence of CDC42SE1 are registered in the NCBI database with the accession number NM_020239.2.
  • G Anti-GPT2 antibody
  • GPT2 Glacial pyruvate transaminase (alanine aminotransferase) 2
  • GPT2 is an enzyme that catalyzes a transamination reaction that produces pyruvate and glutamate from alanine and 2-oxoglutarate.
  • the specific amino acid sequence and base sequence of GPT2 are registered in the NCBI database with the accession number NM_1333443.1.
  • any of the antibodies (a) to (g) may be used as a biomarker, but the antibody titer in the blood sample of NSIP patient is high, and the amount of antibody in the blood sample of IPF / UIP patient Since the difference is more remarkable, anti-Mx1 antibody and anti-NINJ2 antibody are preferable examples of the biomarker of the present invention.
  • any one antibody selected from the above (a) to (g) may be used as a biomarker, or a combination of two or more antibodies may be used.
  • NSIP nonspecific interstitial pneumonia
  • the antibody class is not particularly limited as long as it can be detected, and may be any of IgG (including IgG1, IgG2, IgG3, IgG4, etc.), IgD, IgE, IgA, sIgA, IgM and the like.
  • IgG is exemplified as a preferred antibody class in the present invention.
  • the biomarker of the present invention can also be used for antibody binding fragments (for example, F (ab ′) 2 , Fab ′, Fab, Fv, sFv, dsFv, sdAb). Can be used as
  • Nonspecific Interstitial Pneumonia (NSIP) Testing Method provides a NSIP testing method.
  • the test method of the present invention is selected from the group consisting of anti-Mx1 antibody, anti-NINJ2 antibody, anti-DCX antibody, anti-C10orf49 antibody, anti-ZMAT4 antibody, anti-CDC42SE1 antibody and anti-GPT2 antibody in a blood sample collected from a test animal Detecting at least one antibody produced.
  • the biomarker that is, the antibody
  • the biomarker is a detection target.
  • the origin of the blood sample is not particularly limited, and any animal can be used. More specifically, laboratory animals such as rodents and rabbits such as mice, rats, hamsters and guinea pigs; domestic animals such as pigs, cows, goats, horses and sheep; pets such as dogs and cats; humans, monkeys and orangutans Primates such as chimpanzees.
  • the blood sample is preferably derived from a primate, more preferably a human.
  • the type of blood sample is not particularly limited as long as the biomarker can be detected, and examples thereof include whole blood, serum, and plasma.
  • Serum is a preferred example from the viewpoint that it can be easily prepared and more accurately detects the biomarker.
  • serum can be prepared by removing blood cells and blood coagulation factors such as fibrinogen (factor I), prothrombin (factor II), factor V, and factor VIII from the collected blood (whole blood). It can be obtained as a supernatant obtained after allowing the blood to stand or a supernatant obtained by subjecting the blood to centrifugation.
  • blood sample is used after being diluted to an appropriate concentration as necessary.
  • the detection of each antibody uses at least one selected from the group consisting of Mx1, NINJ2, DCX, C10orf49, ZMAT4, CDC42SE1, and GPT2 or a partial peptide thereof as an antigen, This is performed by detecting the biomarker (antibody).
  • the antigen used to detect the biomarker in the blood sample in the test method of the present invention can be specifically recognized by the biomarker (antibody) present in the blood sample collected from the test animal.
  • an antigen derived from an animal belonging to the same species as the test animal is preferable.
  • the antigen used for detection of the biomarker is preferably derived from a human.
  • the antigen used for detecting the biomarker in the blood sample may be a full-length protein, or a partial peptide thereof as long as it can be specifically recognized by the antibody.
  • the partial peptide used as an antigen is not particularly limited as long as it contains an antigenic determinant that can be recognized by an antibody derived from a test animal, and the antibody detected as the biomarker Depending on the type, it can be set as appropriate. In general, since the number of amino acid residues constituting an antigenic determinant is about 5 to 20, partial peptides include, for example, those composed of 5 or more amino acid residues.
  • the antigen can be obtained by a known method.
  • the antigen can be collected from an animal such as a human exemplified as the test animal.
  • the collection can be performed according to a conventionally known method for isolating and purifying a protein or peptide from tissue or cultured cells.
  • tissue or cell expressing the antigen is disrupted with a homogenizer and then lysed.
  • the antigen can also be isolated and purified from a culture by culturing a transformant into which an expression vector containing a nucleic acid encoding the antigen has been introduced.
  • it can be prepared as a polypeptide by a conventionally known peptide synthesis method based on the amino acid sequence of the antigen. Examples of peptide synthesis methods include solid phase synthesis and liquid phase synthesis, and any method may be used in the present invention.
  • the partial peptide used as an antigen may be prepared by cleaving each polypeptide produced by the above method with a peptidase corresponding to the type of each antigen.
  • the antigen may be modified with phosphate, sugar or sugar chain, phospholipid, lipid, nucleotide or the like.
  • the antigen may be one to which a known tag is linked in order to easily perform a purification process or the like. Examples of such tags include glutathione-S-transferase (GST), FLAG tag, His tag, and the like.
  • the antigen may be used in the form of a complex formed with a major histocompatibility complex (MHC) class II molecule.
  • MHC major histocompatibility complex
  • the MHC class II molecule presenting the antigen may be referred to as “antigen / MHC class II”.
  • antigen / MHC class II will be described.
  • the origin of the MHC class II molecule is not particularly limited, and examples thereof include humans, mice, rats, dogs, monkeys, rabbits, sheep and horses.
  • the origin of the MHC class II molecule may be the same as or different from the origin of the test animal to be examined.
  • the MHC class II molecule is a complex of an ⁇ chain and a ⁇ chain, the type of each is not particularly limited, and the halotype of the gene encoding them is not particularly limited.
  • the MHC class II molecule When the MHC class II molecule is derived from a human, the MHC class II molecule encoded by, for example, the HLA-DPA locus, the HLA-DQA locus, or the HLA-DRA locus as the ⁇ chain of the MHC class II molecule As the ⁇ chain of the MHC class II molecule, for example, the ⁇ chain of the MHC class II molecule encoded by the HLA-DPB locus, the HLA-DQB locus, or the HLA-DRB locus is Can be mentioned.
  • the haplotypes of MHC class ⁇ chain and ⁇ chain at each locus are not particularly limited.
  • the MHC class II molecule is preferably a molecule containing both an ⁇ chain and a ⁇ chain.
  • the MHC class II molecule is not particularly limited as long as it can form a complex with the antigen, and specific examples include HLA-DR, HLA-DP, HLA-DQ, and the like.
  • HLA-DR examples include HLA-DR1, HLA-DR2, HLA-DR3, HLA-DR4, HLA-DR5, HLA-DR6, HLA-DR7, HLA-DR8, HLA-DR9, HLA-DR10, HLA -DR11, HLA-DR12, HLA-DR13, HLA-DR14, HLA-DR15, HLA-DR52, HLA-DR53 and the like.
  • HLA-DR examples include molecules containing HLA-DRA1 as the ⁇ chain and HLA-DRB such as HLA-DRB1, HLA-DRB3, HLA-DRB4, or HLA-DRB5 as the ⁇ chain.
  • HLA-DR As the HLA-DR, more specifically, as an ⁇ chain, an allyl such as HLA-DRA1 * 01, as a ⁇ chain, for example, HLA-DRB1 * 01, HLA-DRB1 * 03, HLA-DRB1 * 04, HLA -DRB1 * 07, HLA-DRB1 * 08, HLA-DRB1 * 09, HLA-DRB1 * 10, HLA-DRB1 * 11, HLA-DRB1 * 12, HLA-DRB1 * 13, HLA-DRB1 * 14, HLA-DRB1 * 15, alleles such as HLA-DRB1 * 16, HLA-DRB3 * 01, HLA-DRB4 * 01, HLA-DRB5 * 01 and the like.
  • HLA-DQ examples include HLA-DQ1, HLA-DQ2, HLA-DQ3, HLA-DQ4, HLA-DQ5, HLA-DQ6, HLA-DQ7, and HLA-DQ8.
  • HLA-DQ examples include molecules containing HLA-DQA such as HLA-DQA1 as the ⁇ chain and HLA-DQB such as HLA-DQB1 as the ⁇ chain.
  • ⁇ chain for example, allele such as HLA-DQA1 * 01, HLA-DQA1 * 02, HLA-DQA1 * 03, HLA-DQA1 * 04, HLA-DQA1 * 05, HLA-DQA1 * 06
  • examples of the ⁇ chain include alleles such as HLA-DQB1 * 02, HLA-DQB1 * 03, HLA-DQB1 * 04, HLA-DQB1 * 05, HLA-DQB1 * 06, and the like.
  • HLA-DP examples include HLA-DP1, HLA-DP2, HLA-DP3, HLA-DP4, and HLA-DP5.
  • HLA-DP examples include molecules containing HLA-DPA such as HLA-DPA1 as an ⁇ chain and HLA-DPB such as HLA-DPB1 as a ⁇ chain.
  • HLA-DPA1 * 01, HLA-DPA1 * 02, HLA-DPA1 * 03, HLA-DPA1 * 04 and the like examples include allyl such as HLA-DPB1 * 04, HLA-DPB1 * 05, HLA-DPB1 * 09 and the like.
  • the MHC class II molecule may be an autoimmune disease sensitive MHC class II molecule.
  • the autoimmune disease-susceptible MHC class II molecule is an MHC class ⁇ and ⁇ chain having a relatively high probability of developing an autoimmune disease with respect to other MHC class ⁇ and ⁇ chain haplotypes (allyl).
  • MHC class II molecules containing at least one.
  • the MHC class II molecules may be used alone or in combination of two or more.
  • the type of the MHC class II molecule may be appropriately set according to the type of antigen to be presented.
  • the MHC class II molecule suitable for presenting Mx1 is preferably HLA-DR; more preferably ⁇ chain is HLA-DRA1 * 01 and ⁇ chain is HLA-DRB1 * 01, HLA-DRB1 * 03, HLA-DRB1 * 04, HLA-DRB1 * 07, HLA-DRB1 * 08, HLA-DRB1 * 09 , HLA-DRB1 * 10, HLA-DRB1 * 11, HLA-DRB1 * 12, HLA-DRB1 * 13, HLA-DRB1 * 14, HLA-DRB1 * 15, HLA-DRB1 * 16, HLA-DRB3 * 01, HLA -HLA-DR that is at least one of DRB4 * 01 and HLA-DRB5 * 01; more preferred
  • the ⁇ chain is HLA-DRA1 * 01 and the ⁇ chain is HLA-DRB1 * 04, HLA-DR
  • the MHC class II molecule on which the antigen is presented can be obtained by introducing a gene encoding the antigen or a partial peptide thereof into a cell that expresses the MHC class II molecule.
  • the cell expressing the MHC class II molecule may be a cell having the gene encoding the MHC class II molecule as an endogenous gene (hereinafter referred to as an MHC class II molecule endogenous cell) or a cell having an exogenous gene (hereinafter referred to as an exogenous gene). MHC class II molecule non-endogenous cells).
  • the antigen-encoding gene may be introduced into a host cell having the MHC class II molecule-encoding gene as an endogenous gene.
  • the antigen-encoding gene and the MHC class II molecule-encoding gene may be co-introduced into the host cell.
  • a method for introducing a gene into a cell can be performed by a known transformation method.
  • MHC class II molecular endogenous cells examples include the immune system cells.
  • the MHC class II molecule non-endogenous cells are not particularly limited, and examples thereof include animal cells, plant cells, and insect cells.
  • animal cells specifically, HeLa cells, 293 cells, 293T cells, NIH3T3 cells, COS cells, CHO cells and other cultured cells, ES cells, stem cells such as hematopoietic stem cells, T cells, B cells, dendritic cells And immune system cells such as macrophages and glial cells, and cells isolated from living organisms such as primary cultured cells.
  • a cell that presents the antigen / MHC class II on the cell surface can be obtained.
  • cells presenting antigen / MHC class II may be used as they are for the detection of the biomarker, or antigen / MHC class II is separated from cells presenting antigen / MHC class II and used as biomarkers. It may be used for measuring a marker.
  • the test method of the present invention is performed by detecting the presence or absence of the biomarker in a blood sample collected from a test animal.
  • the biomarker may be detected by measuring the amount of the biomarker in a blood sample. By measuring the amount of the biomarker, it can be quantitatively compared with the biomarker in a normal control group or a subject suffering from IPF / UIP, and a test with higher accuracy is possible.
  • the method is preferably performed by measuring the amount of the biomarker in a blood sample.
  • the antigen and the blood sample (antibody) are specifically contacted by bringing the antigen into contact with the blood sample.
  • a method for directly or indirectly detecting the binding can be mentioned.
  • Specific examples of such detection methods include immunoassays such as ELISA, Western blot, immunoprecipitation, radioimmunoassay (RIA), and fluorescent immunoassay.
  • a label such as an enzyme label, a chromogenic label, a radiolabel, or a luminescent label is bound to an antibody that specifically binds to the biomarker (antibody), and the label is detected. Or it can carry out by measuring.
  • NSIP clinical diagnosis or histopathological diagnosis can be carried out simply and with high accuracy, regardless of which detection method is employed.
  • ELISA is a preferable detection method.
  • the conditions for performing the immunoassay are not particularly limited as long as specific binding between the biomarker and the antigen in a blood sample can be detected, and is set based on conventionally known conditions.
  • the biomarker of the present invention is detected by the ELISA method, a blood sample collected from a test animal is added to each well of the multiwell plate on which the antigen or the antigen is fixed, and the antigen in the well and the blood sample A biomarker (antibody) is reacted.
  • a labeled antibody that specifically binds to an antibody derived from a test animal is added to each well and reacted, and then a reaction product obtained by adding an enzyme substrate is detected and / or quantified to obtain a blood sample
  • the biomarker in it can be detected and / or quantified.
  • the labeled antibody that specifically binds to the antibody derived from the test animal can be appropriately selected based on the test animal species from which the blood sample is collected. For example, when the test animal is a human, And non-human labeled antibodies that specifically bind human antibodies (for example, rabbit-derived anti-human IgG antibodies).
  • an enzyme used for labeling a labeled antibody that specifically binds to an antibody derived from a test animal can be appropriately selected from those usually used.
  • peroxidase alkaline phosphatase, luciferase
  • Examples include esterase, glucose oxidase, ⁇ -D-galactosidase, ⁇ -D-glucuronidase and the like.
  • the enzyme substrate can be appropriately selected from known substrates depending on the type of enzyme. For example, when the enzyme is peroxidase, 3,3 ′, 5,5′-tetramethylbenzidine (TMB) Can be used as a substrate.
  • TMB 5,5′-tetramethylbenzidine
  • Detection and / or quantification of the reaction product generated by the reaction between the enzyme and the substrate can be performed by measuring the absorbance of the reaction product, for example, 3,3 ′, 5,5′-tetramethylbenzidine (TMB). ) Can be used by measuring the absorbance at 450 nm.
  • TMB 3,3 ′, 5,5′-tetramethylbenzidine
  • the antigen is labeled with a radioisotope, reacted with the biomarker in a blood sample to form an immune complex, and detected based on the radioactivity released from the radioisotope. be able to.
  • the antigen is solid-phased on a plate or the like, a blood sample is added thereto and reacted, and then an antibody that specifically binds to an antibody present in the blood sample of the test animal is further reacted. It can be performed by detecting fluorescence development.
  • an antibody that specifically binds to a test animal-derived antibody one labeled with a fluorescent dye is used as described in the ELISA method. Examples of fluorescent dyes include FITC, PE, APC, Cy-3, and Cy-5.
  • the immunoprecipitation method can be detected by reacting the antigen with a blood sample to form an immune complex and precipitating it as an insolubilized product using an active adsorbent such as protein A or protein G. Furthermore, it can also be detected by a combination of immunoprecipitation and Western blotting. More specifically, the antigen to which a tag such as FLAG is linked is reacted with a blood sample, and if the biomarker is present in the sample, an immune complex is formed, which is precipitated by the aforementioned active adsorbent. . The obtained precipitate is subjected to Western blotting.
  • the precipitate is separated and developed by SDS-PAGE, transferred to a nitrocellulose membrane, PVDF membrane, etc., and then the antibody against the tag is subjected to an antigen-antibody reaction on the transfer membrane, whereby the biomarker is present in the blood sample. If it is, it can be detected as a band.
  • a blood sample may be brought into contact with the cell.
  • the addition ratio of the cells and the blood sample is not particularly limited. For example, 0.01 to 1 ml of the blood sample can be mentioned for 10 to 20 million cells (for example, about 5 million cells).
  • the incubation conditions are not particularly limited, but the temperature is, for example, 0 to 37 ° C., preferably 0 to 10 ° C., more preferably 0 to 5 ° C., and the pH is, for example, pH 6 to 9, preferably pH 7 to 8. More preferably, the pH is 7.2 to 7.6, and the time is, for example, 3 to 120 minutes, preferably 10 to 90 minutes, more preferably 30 to 60 minutes. If a biomarker is present in the blood sample, the biomarker specifically binds to the antigen / MHC class II antigen presented by the cell. Alternatively, it may be measured indirectly.
  • the biomarker of the present invention was detected in a test animal (preferably human) known to be affected with interstitial pneumonia by the conventionally used interstitial pneumonia marker KL-6 In this case, it can be determined that the subject animal suffers from NSIP of idiopathic interstitial pneumonia.
  • the amount of the biomarker of the present invention in the blood sample is significantly increased compared to the normal control group. Therefore, when the biomarker of the present invention is quantitatively detected, NSIP diagnosis is performed by comparing the amount of biomarker in the blood sample of the normal control group and the amount of biomarker in the blood sample of the test animal. It can be performed. Furthermore, although the amount of the biomarker of the present invention is increased in the blood sample of NSIP patients, it is not significantly different from the normal control group in IPF / UIP patients. It can also be used for NSIP identification.
  • the amount of the biomarker contained in the blood sample is larger than that in the normal control group (or the IPF / UIP group that is a disease to be differentiated).
  • the amount of biomarker in the test animal is normal control.
  • the 95th percentile value of a group (or IPF / UIP group that is a disease to be differentiated) is set as a cut-off value, and a case where the value is equal to or higher than the cut-off value can be mentioned. If it is above the cut-off value, it can be determined that the test animal is highly likely to be affected by NSIP, or that it is highly likely that NSIP has a tissue lesion.
  • the test animal is less likely to be affected by NSIP or is less likely to have NSIP tissue lesions. Also, the greater the amount of the biomarker of the invention in the blood sample, the more severe the NSIP symptoms can be predicted.
  • NSIP When the biomarker of the present invention is detected, it is diagnosed that NSIP generally has a good prognosis among idiopathic interstitial pneumonia.
  • the prognosis can be predicted.
  • NSIP can make an appropriate dosing schedule and treatment plan for suppressing NSIP treatment and / or progression of symptoms based on its good therapeutic response.
  • NSIP is, Mx1, NINJ2, DCX, C10orf49 , ZMAT4, CDC42SE1 and at least one or antigen consisting of a partial peptide selected from the group consisting of GPT2, non-specific interstitial pneumonia (NSIP) test reagents are provided.
  • the antigen used in the test reagent of the present invention is as described above.
  • the antigen may be provided immobilized on an insolubilized carrier.
  • the material of the insolubilized carrier is not particularly limited as long as it does not prevent the detection of the biomarker.
  • polystyrene, polyethylene, polypropylene, polyester, polyacrylonitrile, polyvinyl chloride, fluororesin, crosslinked dextran, paper, silicon, glass, metal, Agarose etc. can be illustrated. Two or more of these materials may be used in combination.
  • the shape of the insolubilized carrier may be any shape such as a microplate, a tray shape, a spherical shape, a fiber shape, a rod shape, a disk shape, a container shape, a cell, and a test tube.
  • the antigen may be provided as antigen / MHC class II, as described above, or may be provided in the state of a cell expressing antigen / MHC class II.
  • Immobilization of the antigen on the insolubilized carrier can be performed according to a conventionally known method.
  • the amount of the antigen immobilized on the insolubilized carrier is not particularly limited as long as it is sufficient to specifically bind to the antibody against each antigen.
  • the solution used for immobilization on the insolubilized carrier is 2 to 10 ⁇ g / mL, preferably 2 to 8 ⁇ g / mL, more preferably 2 to 6 ⁇ g / mL.
  • the biomarker detected by the test reagent of the present invention is an antibody capable of recognizing a plurality of antigenic determinants
  • an antibody specifically recognizing each of the plurality of antigenic determinants present in each antigen is a full-length protein from the viewpoint that the detection sensitivity can be improved by comprehensively detecting a).
  • test reagent of the present invention may contain a buffer, a stabilizer, a preservative, and the like in addition to the antigen, and may be formulated according to a conventionally known method.
  • NSIP Diagnostic Kit The present invention provides a diagnostic kit for nonspecific interstitial pneumonia (NSIP) comprising the reagent for testing nonspecific interstitial pneumonia (NSIP).
  • a labeled antibody that specifically binds to an antibody derived from a test animal that may be required to carry out antibody detection.
  • a non-human labeled antibody that specifically binds a human antibody a detection agent for a labeled substance, a solubilizer, a cleaning agent, a reaction stop solution, a control sample, a test protocol, and the like may be included.
  • the present invention provides a NSIP diagnosis method.
  • the diagnostic method is selected from the group consisting of anti-Mx1 antibody, anti-NINJ2 antibody, anti-DCX antibody, anti-C10orf49 antibody, anti-ZMAT4 antibody, anti-CDC42SE1 antibody and anti-GPT2 antibody in a blood sample collected from a test animal.
  • the antibody measurement method, criteria for determining that the test animal suffers from NSIP, and the like are as described above.
  • Patient background serum was collected with written consent from a patient visiting the National Hospital Organization Kinki Central Chest Disease Center. After collecting blood from 10 cases of IPF, 8 cases of NSIP, 10 cases of autoimmune alveolar proteinosis (PAP), 10 cases of sarcoidosis (SAR), and 10 cases of healthy subjects (may be referred to as control) , Centrifuged at 2500 rpm for 10 minutes, and the supernatant was collected and stored at ⁇ 80 ° C. until used for experiments (Table 1). This study was conducted in accordance with the Ethics Guidelines for Clinical Research of the Ministry of Health, Labor and Welfare, after approval by the Clinical Research Ethics Review Committee of Osaka University Hospital and National Hospital Organization Kinki Chuo Chest Disease Center.
  • Protein array method ProtoArray Human Protein Microarray v5.0 (Invitrogen) was purchased and used according to the instruction manual. In the same array, about 8,000 kinds of glutathione S-transferase (GST) fusion full-length proteins prepared with insect cells (Sf9) are spotted at two points each. Place the array in a quadriPERM culture dish (Greiner Bio One) and react with blocking buffer at 4 ° C for 60 minutes, then react with 1: 500 diluted serum for 90 minutes at 4 ° C on a horizontal shaker (50 rpm). It was. After washing, Alexa Fluor 647 goat anti-human IgG (Invitrogen) was diluted 1: 2,000 and reacted at 4 ° C. for 90 minutes.
  • GST glutathione S-transferase
  • the fluorescence intensity median value (relative fluorescence units, rfu) of each spot on the array dried by centrifugation was read with a microarray scanner (Axon 4200AL) equipped with GenePix Pro Software (Molecular Devices).
  • the fluorescence intensity of the antigen protein was converted into a ratio of interquartile differences (RoIQD) value according to the following formula (1).
  • X is the fluorescence intensity of each antigen protein
  • Q1 array is the 25th percentile value from the lowest of the fluorescence intensity of all antigen proteins in the array
  • Q3 array represents the 75th percentile from the lowest of the fluorescence intensities of all antigenic proteins in the array
  • quadrature normalization was performed to determine the serum autoantibody titer.
  • Quartile normalization antigen proteins are arranged in the order of the RoIQD value for each array, the average value of RoIQD values of the same rank (48 RoIQD values in this test example) is calculated, and this average value is the same for each array.
  • the operation of assigning to the antigen protein of the rank was performed. Since a series of operations can suppress variation in data between arrays, it is possible to compare serum autoantibody titers between arrays.
  • the cut-off value of each serum autoantibody was determined according to the following formula (2).
  • Q1 Ag is the 25th percentile of the total antibody titer of each autoantibody (48 in this study)
  • Q3 Ag represents the 75th percentile of the total antibody titer of each autoantibody (48 in this study) Only when the serum autoantibody titer was larger than the cutoff value, the autoantibody titer ratio (s / c) to the cutoff value was calculated.
  • the score value as an autoantibody specific to a specific disease was determined according to the following formula (3).
  • the following formula (3) describes the case of a PAP patient as an example. Here, 10 cases of aPAP were used as the disease group, and 38 cases of other disease groups and healthy persons were used as the control group.
  • Freq is the percentage of cases with an autoantibody titer higher than the cutoff value
  • Internsity represents the average value of autoantibody titer ratio (s / c) to cutoff value)
  • the threshold value of the score value for determining a disease-specific autoantibody was examined.
  • Gnjetic S. Proc Natl Acad Sci USA 2010; 107: 5088-5093 and J Immunol Methods 2009; 341: 50-58
  • adopted a high score> 5 but targeted inflammatory diseases It was considered unsuitable for examination in this test example (FIG. 1). Therefore, when the threshold value was gradually increased, it was found that there was no overlap between each disease when the score ⁇ 57.1 was set as the threshold value.
  • 51 NSIPs, 40 IPFs, 57 aPAPs, 44 SARs were found.
  • the disease-specific autoantibodies could be determined. Among them, seven types of antibodies showing particularly high scores were selected from autoantibodies specific to NSIP disease. Table 2 below shows the scores of the seven autoantibodies.
  • FIG. 2 shows antibody titers in graphs for each disease.
  • the serum anti-Mx1 antibody titer by the protein array was also increased in alveolar proteinosis (PAP), similar to KL-6, which is a conventional interstitial pneumonia marker.
  • PAP alveolar proteinosis
  • the antibody titer was clearly increased in NSIP patients, indicating that it could be a differential marker between IPF and NSIP (FIG. 2).
  • FIG. 3 it was shown that, in addition to the anti-Mx1 antibody, six types of autoantibodies can serve as differential markers between IPF and NSIP.
  • FIG. 3 shows that, in addition to the anti-Mx1 antibody, six types of autoantibodies can serve as differential markers between IPF and NSIP.
  • FIG. 4 shows a graph in which the anti-Mx1 antibody titer and the anti-NINJ2 antibody titer are identified for each NSIP patient.
  • NSIP is roughly divided into patients with high anti-Mx1 antibody titers and patients with high anti-NINJ2 antibodies, and there was little overlap between anti-Mx1 antibody positive and anti-NINJ2 antibody positive. It was revealed that NSIP can be diagnosed with higher accuracy by using both antibodies and anti-NINJ2 antibodies as differential markers.
  • the antigenic protein Mx1 is called Interferon-stimulated genes (ISGs) and belongs to a group of antiviral proteins induced by type 1-interferon. The relationship between interferon signaling and interstitial pneumonia has been reported so far. Specific test methods are shown below.
  • Formalin-fixed paraffin-embedded specimens of normal lung tissue were purchased from SUPER BIO CHIPS (Seoul, Korea).
  • NSIP patient's surgical lung biopsy specimen (formalin fixed paraffin embedded) was provided by National Hospital Organization Kinki Central Chest Disease Center.
  • Rabbit anti-human Mx1 polyclonal antibody was used as the primary antibody.
  • a goat anti-rabbit IgG antibody was used as the secondary antibody.
  • a glass slide with a paraffin section of each specimen was allowed to stand at 60 ° C. overnight and then immersed in xylene (10 minutes (4 times). Thereafter, 100% ethanol, 95% ethanol, 80% ethanol, 50% ethanol) Then, deparaffinization was carried out step by step in the order of water, and after antigen activation using a citrate buffer, endogenous peroxidase inhibition was carried out.
  • the specimen reacted with the secondary antibody was washed with PBST, and then developed using Streptavidin-biotin-peroxidase detection system (Vector Labs Inc.).
  • Streptavidin-biotin-peroxidase detection system Vector Labs Inc.
  • Mx1 was expressed in Clara cells in bronchial epithelium and macrophages in alveoli in normal lung tissue.
  • high expression was observed in type II alveolar epithelial hyperplasia and aggregated alveolar macrophages in the lung tissue of NSIP patients. That is, it is predicted that the localization of Mx1 in the lung tissue of NSIP patients is recognized as an antigen and autoantibodies (anti-Mx1 antibodies) are produced.
  • Mx1 recombinant protein obtained from Invitrogen, Carlsbad, CA expressed in Sf9 insect cells was immobilized on an ELISA plate (Thermo Fisher Scientific Inc., Waltham, MA). The immobilization was performed by incubating the ELISA plate and Mx1 recombinant protein overnight at 4 ° C. The concentration of the Mx1 recombinant protein used for the solid phase was 1 ⁇ g / ml, 2 ⁇ g / ml, 4 ⁇ g / ml or 8 ⁇ g / ml.
  • Serum to be measured is diluted 100-fold, allowed to react with Mx1 recombinant protein immobilized on an ELISA plate at room temperature for 1 hour, washed with phosphate buffered saline, and then HRP-labeled anti-human IgG ( ⁇ chain) antibody ( Goat polyclonal, MBL Co., Ltd., Nagoya, Japan) and allowed to stand at room temperature for 1 hour. After the color was developed using TMB, the absorbance at 450 nm was measured with an ARVO MX1420 multilabel counter (PerkinElmer). The results are shown in the graph of FIG.
  • Mx1 cDNA was obtained from the spleen tissue cDNA library of the MTC multi-tissue cDNA panel (Clontech Laboratories Inc.) using the following primer set. 5'-TTTAAGCTTATGGTTGTTTCCGAAGTG (SEQ ID NO: 1) 5'-TTTTCTAGATTAACCGGGGAACTGGGCAAG (SEQ ID NO: 2)
  • Mx1 cDNA was cloned into pFLAG-CMV TM -4 Expression Vector (Sigma-Aldrich, St. Louis, MO) to obtain an expression vector.
  • This expression vector was transfected into 293 cells, and cell lysates were collected after 16 hours. Cell lysate and serum were mixed and rotated at 4 ° C. overnight, and the immune complex was precipitated using Protein G sepharose (GE Healthcare, Buckinghamshire, UK). The precipitate was separated by SDS-PAGE, and the protein in the gel (precipitate) was transferred to a nitrocellulose membrane. The nitrocellulose membrane was blocked overnight at 4 ° C. in 5% skim milk solution.
  • the membrane was washed with TBS, and the nitrocellulose membrane was reacted with an anti-FLAG antibody (Sigma-Aldrich, St. Louis, MO) at room temperature for 1 hour. Then, it wash
  • Mx1-presented MHC class II molecules Detection of autoantibodies (anti-Mx1 antibodies) using Mx1 presented on MHC class II molecules (1) Preparation of cells that present Mx1-presented MHC class II molecules on the cell surface First, Mx1-presented MHC class is as follows. Cells presenting II molecules on the cell surface were prepared.
  • HLA-DR ⁇ expression vector and HLA-DR ⁇ expression vector HLA-DR ⁇ chain (HLA-DRA * 01: 01) from human peripheral blood mononuclear cells (3H Biomedical) or human cell line cDNA
  • HLA-DRB1 * 03: 04 polynucleotide encoding the ⁇ chain
  • the sequence information of the HLA-DR cDNA was referred to the IMGT / HLA Database (http://www.ebi.ac.uk/imgt/hla/index/html).
  • Mx1 Expression Vector A polynucleotide encoding Mx1c was amplified from a human spleen cDNA library using the following primers by PCR, cloned into a pME18SFL3 vector after restriction enzyme treatment (EcoRI, XhoI).
  • Sense primer 5'-AATAATGAATTCATGGTTGTTTCCGAAGTG-3 '(SEQ ID NO: 3)
  • Anti-sense primer 5'- TAATAACTCGAGTTAACCGGGGAACTGGG-3 '(SEQ ID NO: 4)
  • the expression vector was introduced using the PEI max solution instead of Lipofectamine (trademark) 2000. After 24 hours of culturing, the cells were transferred to GFP. Was confirmed with a fluorescence microscope.
  • Mx1 is complexed with HLA-DR under the above conditions and is displayed on the cell surface because the analysis by flow cytometry using labeled anti-Mx1 antibody and anti-HLA-DR antibody or cell lysate is immunized. After sedimentation, the immunoprecipitated sample has been confirmed by Western blotting using labeled anti-Mx1 antibody and anti-HLA-DR antibody.
  • FIG. 7A is a diagram schematically showing the cell surface of APC positive cells in this test
  • FIG. 7B shows the result of FACS analysis (vertical axis is APC fluorescence intensity).
  • FIG. 7B also shows the results of measuring the serum anti-Mx1 antibody titer using a protein array. From this result, the measurement result of the autoantibody (anti-Mx1 antibody) using the cells presenting the Mx1-HLA-DR complex corresponds well with the serum anti-Mx1 antibody titer by the protein array, and Mx1-HLA It was confirmed that autoantibodies (anti-Mx1 antibodies) can be efficiently and accurately measured by using the -DR complex.
  • anti-Mx1 antibody, anti-NINJ2 antibody, anti-DCX antibody, anti-C10orf49 antibody, anti-ZMAT4 antibody, anti-CDC42SE1 antibody and anti-GPT2 antibody are biomarkers that can perform NSIP testing easily and accurately. These biomarkers have been shown to be useful in the differentiation between IPF / UIP and NSIP. In particular, it was shown that NSIP diagnosis and discrimination between IPF / UIP and NSIP can be performed with higher accuracy by using anti-Mx1 antibody and / or anti-NINJ2 antibody as biomarkers.
  • SEQ ID NO: 1 shows the base sequence of the primer used for cloning of Mx1 cDNA.
  • SEQ ID NO: 2 shows the base sequence of a primer used for cloning of Mx1 cDNA.
  • SEQ ID NO: 3 shows the nucleotide sequence of a primer used for cloning of Mx1 cDNA.
  • SEQ ID NO: 4 shows the base sequence of a primer used for cloning of Mx1 cDNA.
  • SEQ ID NO: 5 shows the base sequence of GFP cDNA.

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Molecular Biology (AREA)
  • Urology & Nephrology (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Hematology (AREA)
  • Food Science & Technology (AREA)
  • Analytical Chemistry (AREA)
  • Microbiology (AREA)
  • Cell Biology (AREA)
  • Biotechnology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Peptides Or Proteins (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

Le problème principal résolu par la présente invention est l'apport d'un biomarqueur qui permet un diagnostic clinique ou histopathologique simple, à haute précision, d'une pneumonie interstitielle non spécifique. Au moins un anticorps, contenu dans un échantillon sanguin, choisi parmi un groupe consistant en un anticorps anti-Mx1, un anticorps anti-NINJ2, un anticorps anti-DCX, un anticorps anti-C10orf49, un anticorps anti-ZMAT4, un anticorps anti-CDC42SE1, et un anticorps anti-GPT2 peuvent servir en tant que biomarqueur pour une pneumonie interstitielle non spécifique (NSIP).
PCT/JP2014/057128 2013-03-18 2014-03-17 Biomarqueur pour le diagnostic d'une pneumonie interstitielle non spécifique WO2014148429A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
JP2015506765A JP6315832B2 (ja) 2013-03-18 2014-03-17 非特異的間質性肺炎の診断のためのバイオマーカー

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2013-055358 2013-03-18
JP2013055358 2013-03-18

Publications (1)

Publication Number Publication Date
WO2014148429A1 true WO2014148429A1 (fr) 2014-09-25

Family

ID=51580108

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2014/057128 WO2014148429A1 (fr) 2013-03-18 2014-03-17 Biomarqueur pour le diagnostic d'une pneumonie interstitielle non spécifique

Country Status (2)

Country Link
JP (1) JP6315832B2 (fr)
WO (1) WO2014148429A1 (fr)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP3734285A1 (fr) * 2019-04-25 2020-11-04 Sapporo Medical University Procédé d'acquisition d'informations sur l'état d'un patient atteint de pneumonie interstitielle et son utilisation
WO2021177691A1 (fr) * 2020-03-02 2021-09-10 연세대학교 산학협력단 Composition pour le diagnostic ou le traitement de la résistance aux médicaments anticancéreux

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
H.-P. HAUBER: "Nicht spezifische interstitielle Pneumonie.", PNEUMOLOGIE, vol. 65, no. 8, August 2011 (2011-08-01), pages 477 - 483 *
Y. YANG: "Detection of antivimentin antibody in sera of patients with idiopathic pulmonary fibrosis and non-specific interstitial pneumonia", CLINICAL AND EXPERIMENTAL IMMUNOLOGY, vol. 128, no. 1, April 2002 (2002-04-01), pages 169 - 174 *

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP3734285A1 (fr) * 2019-04-25 2020-11-04 Sapporo Medical University Procédé d'acquisition d'informations sur l'état d'un patient atteint de pneumonie interstitielle et son utilisation
US11719706B2 (en) 2019-04-25 2023-08-08 Sapporo Medical University Method for measuring a biomarker in a biological sample of an IPAF patient
WO2021177691A1 (fr) * 2020-03-02 2021-09-10 연세대학교 산학협력단 Composition pour le diagnostic ou le traitement de la résistance aux médicaments anticancéreux

Also Published As

Publication number Publication date
JPWO2014148429A1 (ja) 2017-02-16
JP6315832B2 (ja) 2018-04-25

Similar Documents

Publication Publication Date Title
US11199549B2 (en) MEl'hods and means for diagnosing spondylarthritis using autoantibody markers
JP5663723B2 (ja) 関節リウマチの診断のための方法およびキット
CA2814023C (fr) Fragments de moesine et ses utilisations
US20150185226A1 (en) Method for Diagnosing Scleroderma
US20150133322A1 (en) Method for the diagnosis of early rheumatoid arthritis
CA2814029C (fr) Fragments de la moesine associes a une thrombocytopenie immunitaire
JP6315832B2 (ja) 非特異的間質性肺炎の診断のためのバイオマーカー
JP5924502B2 (ja) リンパ球性漏斗下垂体後葉炎のバイオマーカー及びその用途
US11460467B2 (en) Diagnosis method for lupus
US9915667B2 (en) Methods and means for diagnosing vasculitis
WO2019111797A1 (fr) Biomarqueur pour diagnostiquer une maladie musculaire
WO2023275235A1 (fr) Procédé et moyens de diagnostic de spondylarthrite
US8313917B2 (en) Methods of diagnosing latent and active malignancies
CN115605758A (zh) 使用蛋白质-精氨酸脱亚胺酶1(pad1)自身抗原诊断和评估类风湿性关节炎的组合物和方法
CN116970693A (zh) 作为早老性痴呆诊断标志物的ddit4l剪切产物

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 14770778

Country of ref document: EP

Kind code of ref document: A1

ENP Entry into the national phase

Ref document number: 2015506765

Country of ref document: JP

Kind code of ref document: A

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 14770778

Country of ref document: EP

Kind code of ref document: A1