WO2022075773A1 - Biomarker for diagnosing pancreatic cancer and use thereof - Google Patents

Biomarker for diagnosing pancreatic cancer and use thereof Download PDF

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WO2022075773A1
WO2022075773A1 PCT/KR2021/013796 KR2021013796W WO2022075773A1 WO 2022075773 A1 WO2022075773 A1 WO 2022075773A1 KR 2021013796 W KR2021013796 W KR 2021013796W WO 2022075773 A1 WO2022075773 A1 WO 2022075773A1
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pancreatic cancer
alpha
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isoform
protein
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윤승배
송미영
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가톨릭대학교 산학협력단
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    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
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    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
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    • 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
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
    • 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
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
    • G01N33/57407—Specifically defined cancers
    • G01N33/57438—Specifically defined cancers of liver, pancreas or kidney
    • 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
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
    • G01N33/57484—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving compounds serving as markers for tumor, cancer, neoplasia, e.g. cellular determinants, receptors, heat shock/stress proteins, A-protein, oligosaccharides, metabolites
    • 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
    • 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
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/158—Expression markers

Definitions

  • the present invention relates to a biomarker for diagnosing pancreatic cancer and its use, and more particularly, to a marker composition for diagnosing pancreatic cancer, a composition for diagnosing pancreatic cancer, a diagnostic kit, and a method for providing information for diagnosing pancreatic cancer.
  • Pancreatic cancer is a malignant tumor originating from the pancreas, and it is known that the 5-year survival rate is less than 10% because most patients are found to have advanced cancer. Although the incidence of pancreatic cancer in Korea ranks eighth, cancer-related deaths account for just after lung cancer, liver cancer, stomach cancer, and colorectal cancer. Symptoms of pancreatic cancer include symptoms found in various pancreatic diseases, and abdominal pain, anorexia, weight loss, and jaundice are the most common symptoms. Jaundice occurs in most cases.
  • Korean Patent Application Laid-Open No. 2009-0003308 discloses a method for diagnosing pancreatic cancer by detecting the expression level of REG4 protein in an individual's blood sample
  • Korean Patent Publication No. 2012-0009781 provides information necessary for diagnosing pancreatic cancer in an individual
  • an analysis method for measuring the expression level of XIST RNA in cancer tissues isolated from individuals is disclosed in Korean Patent Publication No. 2007-0119250, a novel gene LBFL313 family that is differently expressed in human pancreatic cancer tissues compared to normal human pancreatic tissues.
  • US Patent Publication No. 2011/0294136 discloses a method for diagnosing pancreatic cancer using biomarkers such as keratin 8 protein.
  • biomarkers such as keratin 8 protein
  • the present inventors collected plasma from normal persons and pancreatic cancer patients in order to discover a genetic marker capable of diagnosing pancreatic cancer, and separated exosomes therefrom to analyze exosome proteins,
  • the present invention was completed based on the discovery of a gene whose expression was significantly increased only in pancreatic cancer patients compared to normal persons.
  • the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4) , NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) containing It provides a marker composition for diagnosing pancreatic cancer, comprising the mRNA of the gene or the protein encoded by the gene.
  • the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591).
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4
  • ENPEP Glutamyl aminopeptidase; NM_001977.4
  • CD14 Monocyte differentiation antigen CD14; NM_000591.
  • NM_001040021.3 NM_001174104.2, NM_001174105.2
  • MARF1 Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2
  • PIGR Polymeric immunoglobulin receptor
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte Differentiation antigen CD14; NM_002644.4) provides an information providing method for diagnosing pancreatic cancer, including measuring the level of mRNA of a gene or a protein encoded from the gene.
  • the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 ( Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor) ; NM_002644.4) provides a marker composition for diagnosing pancreatic cancer, including mRNA of a gene or a protein encoded by the gene.
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_00
  • the marker composition is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277) .2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX) ; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_00135
  • NAP1L1 Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2
  • AMY2A Pancreatic alpha-amylase; NM_000699.3
  • TPM1 Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411.
  • NM_001286665.2 group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2
  • ANPEP Aminopeptidase N; NM_001150.3
  • LAMP2 Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1
  • It may further include mRNA of one or more genes selected from or a protein encoded by the gene.
  • the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591).
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4
  • ENPEP Glutamyl aminopeptidase; NM_001977.4
  • CD14 Monocyte differentiation antigen CD14; NM_000591.
  • NM_001040021.3 including NM_001174104.2, NM_001174105.2
  • MARF1 Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2
  • PIGR Polymeric immunoglobulin receptor; NM_002644.4
  • the composition for diagnosing pancreatic cancer is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein) IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM
  • NAP1L1 Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2
  • AMY2A Pancreatic alpha-amylase; NM_000699.3
  • TPM1 Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411.
  • NM_001286665.2 group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2
  • ANPEP Aminopeptidase N; NM_001150.3
  • LAMP2 Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1
  • It may further comprise an agent for measuring the mRNA level of one or more genes selected from or a protein encoded by the gene.
  • the composition can diagnose pancreatic cancer early.
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte Differentiation antigen CD14; NM_002644.4) provides an information providing method for diagnosing pancreatic cancer, including measuring the level of mRNA of a gene or a protein encoded from the gene.
  • the method comprises HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.
  • LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1) ), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-L
  • NAP1L1 Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2
  • AMY2A Pancreatic alpha-amylase; NM_000699.3
  • TPM1 Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411.
  • NM_001286665.2 group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2
  • ANPEP Aminopeptidase N; NM_001150.3
  • LAMP2 Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1
  • It may further comprise the step of measuring the mRNA level of one or more genes selected from or a protein encoded from the gene.
  • the biological sample may be exosomes derived from blood or plasma.
  • the mRNA level is determined by next generation sequencing (NGS), polymerase chain reaction (PCR), reverse transcription polymerase chain reaction (RT-PCR), real-time polymerase chain reaction ( Real-time PCR), RNase protection assay (RPA), microarray, and northern blotting (northern blotting) can be measured through one or more methods selected from the group consisting of.
  • NGS next generation sequencing
  • PCR polymerase chain reaction
  • RT-PCR reverse transcription polymerase chain reaction
  • Real-time PCR real-time polymerase chain reaction
  • RNase protection assay RNase protection assay
  • microarray and northern blotting (northern blotting) can be measured through one or more methods selected from the group consisting of.
  • the protein level is determined by western blotting, radioimmunoassay (RIA), radioimmunodiffusion, enzyme immunoassay (ELISA), immunoprecipitation. , flow cytometry, immunofluorescence, ouchterlony, complement fixation assay, and at least one method selected from the group consisting of a protein chip can be measured through
  • the present inventors analyzed the protein in exosomes isolated from plasma of normal people and pancreatic cancer patients, and discovered a gene with significantly increased expression in pancreatic cancer patients compared to normal people as biomarkers for diagnosing pancreatic cancer. It is expected to be used effectively in clinical practice.
  • 1 to 35 are analysis of the expression level of proteins in exosomes isolated from plasma of normal people and pancreatic cancer patients, and the bars shown in the figure represent the quantitative protein amounts of each sample, among which the blue bar is the average value of each group sample. and standard deviation (x-axis of all graphs is Quan Channels, left is PC or EPC, right is control, y-axis is Abundance [a.u.], 10 per division).
  • FIG. 1 shows the results of confirming the expression level of MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. That is, Figure 1a shows the results of normal people and pan-pancreatic cancer patients, and Figure 1b shows the results of normal people and early-pancreatic cancer patients.
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA
  • Figure 2 shows the results of confirming the expression level of ENPEP (Glutamyl aminopeptidase; NCBI accession number: NM_001977.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients
  • Figure 2a shows normal people
  • Figure 2b shows the results of patients with pancreatic cancer (Pan-pancreatic cancer), normal people and early-shows the results of patients with early pancreatic cancer.
  • FIG. 3 shows the results of confirming the expression level of CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients
  • Figure 3a shows the results of normal people and pan-pancreatic cancer (Pan-pancreatic cancer) patients
  • Figure 3b shows the results of normal people and early-pancreatic cancer (Early pancreatic cancer) patients.
  • Figure 4 shows the expression level of MARF1 (Meiosis regulator and mRNA stability factor 1; NCBI accession number: NM_014647.4, NM_001184998.2, NM_001184999.2) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. The confirmed results are shown.
  • MARF1 Meiosis regulator and mRNA stability factor 1; NCBI accession number: NM_014647.4, NM_001184998.2, NM_001184999.2
  • PIGR Polymeric immunoglobulin receptor
  • HIST1H2AB Histone H2A type 1-B/E; NM_003513.3
  • FIG. 7 shows the results of confirming the expression level of CLCA1 (Calcium-activated chloride channel regulator 1; NCBI accession number: NM_001285.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients
  • FIG. 7a is a normal person and pan-pancreatic cancer (Pan-pancreatic cancer) patient results
  • Figure 7b is a normal person and early - shows the results of pancreatic cancer (Early pancreatic cancer) patients.
  • LACRT Extracellular glycoprotein lacritin; NCBI accession number: NM_033277.2
  • LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients; is shown.
  • OSTF1 Ostoclast-stimulating factor 1; NCBI accession number: NM_012383.5
  • GP9 Platinum glycoprotein IX; NCBI accession number: NM_000174.5
  • exosome proteins isolated from plasma of normal people and pancreatic cancer patients show the results of confirming the expression level of GP9 (Platelet glycoprotein IX; NCBI accession number: NM_000174.5) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
  • FIG. 12 shows the results of confirming the expression level of PSMA3 (Proteasome subunit alpha type-3; NCBI accession number: NM_002788.4, NM_152132.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
  • Figure 12a shows the results of normal people and pan-pancreatic cancer patients
  • Figure 12b shows the results of normal people and early-pancreatic cancer patients.
  • MME (Neprilysin; NCBI accession number: NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643) proteins in exosomes isolated from plasma of normal people and pancreatic cancer patients. 1) shows the result of confirming the expression level.
  • Figure 14 shows the results of confirming the expression level of UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NCBI accession number: NM_006759.4, NM_001001521.2) in the protein in the exosome isolated from the plasma of normal people and pancreatic cancer patients. it has been shown
  • ANK1 Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14
  • FIG. 16 shows ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NCBI accession number: NM_001995.5, NM_001286708.2, NM_001381877.1, Expression of NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2) expression
  • FIG. 16a shows the results of normal people and pan-pancreatic cancer patients
  • FIG. 16b shows the results of normal people and early-pancreatic cancer patients.
  • FIG. 17 shows the results of confirming the expression level of AHSG (Alpha-2-HS-glycoprotein; NCBI accession number: NM_001622.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients
  • FIG. 17a Figure 17b shows the results of normal people and pan-pancreatic cancer patients
  • Fig. 17b shows the results of normal people and early-pancreatic cancer patients.
  • VASP Vasodilator-stimulated phosphoprotein
  • Figure 19 shows the result of confirming the expression level of F10 (Coagulation factor X; NCBI accession number: NM_000504.4, NM_001312674.2, NM_001312675.2) in the protein in the exosome isolated from the plasma of normal people and pancreatic cancer patients will be.
  • F10 Coagulation factor X; NCBI accession number: NM_000504.4, NM_001312674.2, NM_001312675.2
  • Figure 20 shows the result of confirming the expression level of PSMB3 (Proteasome subunit beta type-3; NCBI accession number: NM_002795.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients
  • Figure 20a is Figure 20b shows the results of normal people and pan-pancreatic cancer patients, and normal people and early-pancreatic cancer patients.
  • MMRN1 Multimerin-1; NCBI accession number: NM_007351.3
  • HABP2 Hyaluronan-binding protein 2; NCBI accession number: NM_004132.5, NM_001177660.3
  • LAMB1 Laminin subunit beta-1; NCBI accession number: NM_002291.3
  • MMP9 Microx metalloproteinase-9; NCBI accession number: NM_004994.3
  • NAP1L1 Nucleosome assembly protein 1-like 1; NCBI accession number: NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2 shows the results of confirming the expression level
  • FIG. 25a shows the results of normal people and pan-pancreatic cancer patients
  • FIG. 25b shows the results of normal people and early-pancreatic cancer patients. will be.
  • Figure 26 shows the results of confirming the expression level of AMY2A (Pancreatic alpha-amylase; NCBI accession number: NM_000699.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
  • Figure 26b shows the results of normal people and early-pancreatic cancer (Early pancreatic cancer) patients.
  • TPM1 Tropomyosin alpha-1 chain; NCBI accession number: NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366 in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
  • FIG. 27a shows the results of normal people and pan-pancreatic cancer patients.
  • FIG. 27b shows normal people and early-pancreatic cancer patients. shows the results of
  • Figure 28 shows the results of confirming the expression level of CANX (Isoform 2 of Calnexin; NCBI accession number: NM_001363994.1) in the protein in the exosomes isolated from the plasma of normal people and pancreatic cancer patients
  • Figure 28a is normal and Pan-pancreatic cancer (Pan-pancreatic cancer) patient results
  • Figure 28b shows the results of normal people and early-pancreatic cancer (Early pancreatic cancer) patients.
  • FIG. 29 shows GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NCBI accession number: NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. Shows the results of confirming the expression level, and FIG. 29a shows the results of normal people and pan-pancreatic cancer patients, and FIG. 29b shows the results of normal people and early-pancreatic cancer patients.
  • HLA-C HLA class I histocompatibility antigen, C alpha chain; NCBI accession number: NM_002117.6, NM_001243042.1
  • Figure 31 shows the expression level of PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NCBI accession number: NM_000282.4, NM_001127692.3, NM_001178004.2) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. shows the results of checking .
  • PCCA Propionyl-CoA carboxylase alpha chain, mitochondrial; NCBI accession number: NM_000282.4, NM_001127692.3, NM_001178004.2
  • Figure 32 shows the results of confirming the expression level of CHDR2 (Cadherin-related family member 2; NCBI accession number: NM_001171976.2, NM_017675.5) in the protein in the exosome isolated from the plasma of normal people and pancreatic cancer patients .
  • Figure 33 shows POSTN (Periostin; NCBI accession) number: NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666 in exosome proteins isolated from plasma of normal persons and pancreatic cancer patients. 2, NM_001286667.2, NM_001330517.2) shows the results of checking the expression level.
  • ANPEP Aminopeptidase N; NCBI accession number: NM_001150.3
  • LAMP2 immunoglobulin-2C of Lysosome-associated membrane glycoprotein 2; NCBI accession number: NM_001122606.1
  • the present inventors collected plasma from normal persons and pancreatic cancer patients, separated exosomes from them, and analyzed the exosome proteins. As a result, compared to normal persons, expression was significantly expressed only in pancreatic cancer patients. The present invention was completed based on the discovery of this increasing gene.
  • the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591).
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4
  • ENPEP Glutamyl aminopeptidase; NM_001977.4
  • CD14 Monocyte differentiation antigen CD14; NM_000591.
  • NM_001040021.3 NM_001174104.2, NM_001174105.2
  • MARF1 Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2
  • PIGR Polymeric immunoglobulin receptor
  • the marker composition is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2) , LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.
  • HIST1H2AB Histone H2A type 1-B/E; NM_003513.3
  • CLCA1 Calcium-activated chloride channel regulator 1; NM_001285.4
  • LACRT Extracellular glycoprotein lacritin; NM_033277.2
  • LTBP1 Isoform 4 of Latent-transforming growth factor beta-binding protein 1; Uni
  • PSMA3 Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3
  • MME Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1
  • UGP2 UGP2
  • UGP2 UGP2
  • UGP2 UGP2
  • UGP2 UGP2
  • UGP2 UGP2
  • UGP2 UGP2
  • UGP2 UGP2
  • ANK1 Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14
  • ACSL1 Long-chain-fatty-acid -CoA ligase 1;NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_00
  • NAP1L1 Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2
  • AMY2A Pancreatic alpha-amylase; NM_000699.3
  • TPM1 Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411.
  • NM_001286665.2 group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2
  • ANPEP Aminopeptidase N; NM_001150.3
  • LAMP2 Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1
  • It may further include mRNA of one or more genes selected from or a protein encoded by the gene.
  • pancreatic cancer refers to an aggressive characteristic in which pancreatic cells divide and grow ignoring normal growth limits, an invasive characteristic that penetrates into surrounding tissues, and a metastatic (metastatic) characteristic that spreads to other parts of the body.
  • Metastatic refers to diseases caused by cells with characteristics.
  • Diagnosis in a broad sense means judging the actual condition of a patient's disease in all aspects. The content of the judgment is the name of the disease, etiology, type, severity, detailed mode of the disease, and the presence or absence of complications. Diagnosis in the present invention is to determine whether or not cancer is onset and the level of progression.
  • the present inventors identified the use of a specific gene according to the present invention as a novel marker for diagnosing pancreatic cancer through specific examples.
  • AHSG, VASP, F10, PSMB3, MMRN1, HABP2, LAMB1, MMP9, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP and LAMP2 was significantly increased.
  • the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation) antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644) .4) provides a composition for diagnosing pancreatic cancer and a kit for diagnosing pancreatic cancer comprising the composition, comprising an agent for measuring the level of mRNA or protein encoded by the gene comprising the gene.
  • MAN1A1 Mannosyl-oligosaccharide 1,2-
  • the composition for diagnosing pancreatic cancer is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2) ), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174) .5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM
  • NAP1L1 Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2
  • AMY2A Pancreatic alpha-amylase; NM_000699.3
  • TPM1 Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411.
  • NM_001286665.2 group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2
  • ANPEP Aminopeptidase N; NM_001150.3
  • LAMP2 Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1
  • It may further comprise an agent for measuring the mRNA level of one or more genes selected from or a protein encoded by the gene.
  • the composition can diagnose pancreatic cancer early.
  • the kit for prediction of the present invention consists of one or more other component compositions, solutions or devices suitable for the analysis method.
  • the kit of the present invention contains genomic DNA derived from a sample to be analyzed, a primer set specific for the marker gene of the present invention, an appropriate amount of a DNA polymerase, a dNTP mixture, a PCR buffer, and water to perform PCR. It may be a kit comprising
  • the PCR buffer solution may contain KCl, Tris-HCl and MgCl 2 .
  • components necessary for performing electrophoresis that can confirm whether or not the PCR product is amplified may be additionally included in the kit of the present invention.
  • the kit of the present invention may be a kit including essential elements necessary for performing RT-PCR.
  • the RT-PCR kit includes a test tube or other suitable container, reaction buffer, deoxynucleotides (dNTPs), enzymes such as Taq-polymerase and reverse transcriptase, DNase, RNase inhibitors, DEPC -Water (DEPC-water), sterile water, etc. may be included.
  • dNTPs deoxynucleotides
  • enzymes such as Taq-polymerase and reverse transcriptase
  • DNase DNase
  • RNase inhibitors DEPC -Water
  • sterile water etc.
  • a primer pair specific for a gene used as a quantitative control may be included.
  • the kit of the present invention may be a kit including essential elements necessary for performing a DNA chip.
  • the DNA chip kit may include a substrate to which cDNA corresponding to a gene or fragment thereof is attached as a probe, and the substrate may include cDNA corresponding to a quantitative structural gene or fragment thereof.
  • the kit of the present invention may be in the form of a microarray having a substrate on which the marker gene of the present invention is immobilized.
  • the kit of the present invention may be a kit characterized in that it includes essential elements necessary for performing ELISA.
  • the ELISA kit includes an antibody specific for a marker protein, and an agent for measuring the protein level.
  • the ELISA kit may include a reagent capable of detecting an antibody that has formed an "antigen-antibody complex", for example, a labeled secondary antibody, chromopores, an enzyme, and a substrate thereof.
  • an antibody specific for the quantitative control protein may be included.
  • MAN1A1 Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4
  • ENPEP Glutamyl aminopeptidase; NM_001977.
  • CD14 Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2
  • MARF1 Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2
  • PIGR Polymeric immunoglobulin receptor; NM_002644.4
  • the method is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5) ), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP
  • NAP1L1 Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2
  • AMY2A Pancreatic alpha-amylase; NM_000699.3
  • TPM1 Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411.
  • NM_001286665.2 group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2
  • ANPEP Aminopeptidase N; NM_001150.3
  • LAMP2 Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1
  • It may further comprise the step of measuring the mRNA level of one or more genes selected from or a protein encoded from the gene.
  • the term “information providing method for diagnosing pancreatic cancer” used in the present invention provides objective basic information necessary for diagnosing pancreatic cancer as a preliminary step for diagnosis, and the clinical judgment or opinion of a doctor is excluded.
  • the subject-derived biological sample may include tissues, cells, whole blood, blood, saliva, sputum, cerebrospinal fluid and urine, preferably blood or plasma, and more preferably plasma-derived exosomes.
  • the present invention is not limited thereto.
  • the mRNA level is determined by next generation sequencing (NGS), polymerase chain reaction (PCR), reverse transcription polymerase chain reaction (RT-PCR), real-time polymerase chain reaction (Real-time PCR), RNase protection assay (RNase protection assay; RPA), microarray (microarray), and may be measured through one or more methods selected from the group consisting of northern blotting (northern blotting), but is not limited thereto.
  • NGS next generation sequencing
  • PCR polymerase chain reaction
  • RT-PCR reverse transcription polymerase chain reaction
  • Real-time PCR real-time polymerase chain reaction
  • RPA RNase protection assay
  • microarray microarray
  • the protein level was determined by western blotting, radioimmunoassay (RIA), radioimmunodiffusion, enzyme immunoassay (ELISA), immunoprecipitation, flow cytometry, and immunity. It may be measured by at least one method selected from the group consisting of immunofluorescence, ouchterlony, complement fixation assay, and protein chip, but is limited thereto it's not going to be
  • Plasma samples from normal and pancreatic cancer patients are collected in BD Vacutainer Plastic K2EDTA Tube (BD, Cat.no 367525), and plasma is separated by centrifugation within 2 hours at 2,500 rpm and 4°C for 20 minutes. The supernatant plasma was aliquoted into Protein LoBind Tubes (Eppendorf, Cat.no 0030108116) by 500 ⁇ l and stored at -80°C.
  • Plasma samples were centrifuged at 3,000 g and 4 °C for 15 minutes to remove cells and cell debris in the plasma. Then, 250 ⁇ l of Exo2D TM for Protein assay (EXOSOMEplus) was added to 500 ⁇ l of plasma sample at a 2:1 ratio to separate exosomes based on the Aqueous Two-Phase System, and then, HaltTM Protease and Phosphatase Inhibitor Cocktail, EDTA-Free ( The exosome samples were prepared by resuspending the exosomes in 200 ⁇ l of PBS with the addition of Thermo scientific). Samples were stored at -80 °C until analysis.
  • EXOSOMEplus Exo2D TM for Protein assay
  • exosome protein 20 ⁇ g was mixed with SDS loading buffer and then electrophoresed using 10% Sodium Dodecyl Sulfate Polyacrylamide Gel Eletrophoresis (SDS-PAGE) technique to separate proteins according to molecular weight size. After immersing the electrophoresed gel in InstantBlue (Coomassie blue stanning) reagent and staining at room temperature for 15 minutes, protein detection and distribution were first checked.
  • InstantBlue Concentrassie blue stanning
  • 80 ⁇ l of 50 mM TEAB buffer was added and centrifuged at 1,000 g for 60 seconds.
  • 80 ⁇ l of 0.2% FA was added and centrifuged at 1,000 g for 60 seconds.
  • 80 ⁇ l of elution buffer (50% ACN, 0.2% FA) was added, centrifuged at 4,000 g for 60 seconds, and the eluted sample was freeze-dried.
  • Protein peptide samples prepared through the digestion process were analyzed for proteomic bodies using Q Exactive Plus LC-MS/MS System mass spectrometer (Thermo Scientific). Each sample was measured in duplicate, and the data generated by the instrument was analyzed using Proteome Discoverer TM 2.2 (Thermo Scientific) software for database search, comprehensive protein and peptide identification, characterization and quantitative analysis and statistical analysis. was carried out.

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Abstract

The present invention relates to a biomarker for diagnosing pancreatic cancer and use thereof and, more particularly, to a marker composition for diagnosing pancreatic cancer, a composition and diagnostic kit for diagnosing pancreatic cancer, and a method for providing information for diagnosing pancreatic cancer. As a result of analysis of a protein in exosomes isolated from plasma of normal individuals and pancreatic cancer patients, the inventors of the present invention discovered genes of which the expression was significantly increased in pancreatic cancer patients compared to normal individuals. Thus, the genes are expected to be efficiently used in clinical practice as biomarkers useful in the diagnosis of pancreatic cancer.

Description

췌장암 진단용 바이오마커 및 이의 용도Biomarkers for diagnosing pancreatic cancer and uses thereof
본 발명은 췌장암 진단용 바이오마커 및 이의 용도에 관한 것으로, 보다 구체적으로는 췌장암 진단용 마커 조성물, 췌장암 진단용 조성물, 진단 키트, 및 췌장암 진단을 위한 정보제공방법에 관한 것이다.The present invention relates to a biomarker for diagnosing pancreatic cancer and its use, and more particularly, to a marker composition for diagnosing pancreatic cancer, a composition for diagnosing pancreatic cancer, a diagnostic kit, and a method for providing information for diagnosing pancreatic cancer.
췌장암은 췌장에서 기원한 악성 종양으로, 대부분의 환자가 암이 진행된 상태로 발견되기 때문에 5년 생존율이 10%도 되지 않는 암으로 알려져있다. 췌장암의 국내 발생률은 8위에 이르지만, 암에 의한 사망에는 폐암, 간암, 위암, 대장암 바로 다음을 차지하고 있다. 췌장암의 증상으로는 여러 가지 췌장 질환에서 볼 수 있는 증상이 나타날 수 있으며, 복통, 식욕부진, 체중감소, 황달 등이 가장 흔한 증상으로 췌장암 환자의 대부분에서 복통과 체중감소가 나타나고, 췌두부암 환자의 대부분에서 황달이 나타난다.Pancreatic cancer is a malignant tumor originating from the pancreas, and it is known that the 5-year survival rate is less than 10% because most patients are found to have advanced cancer. Although the incidence of pancreatic cancer in Korea ranks eighth, cancer-related deaths account for just after lung cancer, liver cancer, stomach cancer, and colorectal cancer. Symptoms of pancreatic cancer include symptoms found in various pancreatic diseases, and abdominal pain, anorexia, weight loss, and jaundice are the most common symptoms. Jaundice occurs in most cases.
현재까지 증상이 나타나기 전에 췌장암을 조기에 발견할 수 있는 공인된 선별검사 방법이 없는 실정이며, 복부 초음파, 복부 전산화 단층촬영(CT), 자기공명영상(MRI), 내시경적 역행성 담췌관 조영술(ERCP), 내시경 초음파(EUS), 양성자방출 단층촬영(PET), 혈청종양 표지자(CA19-9) 검사에 대한 연구가 활발히 이루어지고 있으나, 아직 유효성이 입증된 진단 방법은 제시되지 못하였다. 따라서 췌장암을 조기에 진단하여 치료 효율을 높일 수 있는 방법의 개발이 시급한 실정이며, 이에 앞서 췌장암의 발병 여부를 미리 예측 가능하게 함으로써 조기진단 및 치료에 대한 대응방법을 차별화하는 것은 매우 중요하므로, 이에 대한 연구 및 기술개발이 요구된다.To date, there is no approved screening method for early detection of pancreatic cancer before symptoms appear. Abdominal ultrasound, abdominal computed tomography (CT), magnetic resonance imaging (MRI), endoscopic retrograde cholangiopancreatography ( ERCP), endoscopic ultrasound (EUS), proton emission tomography (PET), and serum tumor marker (CA19-9) tests are being actively studied, but a diagnostic method with proven effectiveness has not yet been presented. Therefore, there is an urgent need to develop a method for diagnosing pancreatic cancer early and improving treatment efficiency. Research and technology development are required.
대한민국 공개특허 제2009-0003308호는 개체의 혈액 시료에서 REG4 단백질의 발현량을 검출하여 췌장암을 진단하는 방법을 개시하고 있으며, 대한민국 공개특허 제2012-0009781호는 개체의 췌장암 진단에 필요한 정보를 제공하기 위하여 개체로부터 분리한 암 조직 중 XIST RNA의 발현량을 측정하는 분석방법을, 대한민국 공개특허 제2007-0119250호는 정상 인간의 췌장 조직과 비교하여 인간 췌장암 조직에서 다르게 발현된 신규 유전자 LBFL313 패밀리를 개시하고 있고, 미국 공개특허 제2011/0294136호는 케라틴 8 단백질 등의 바이오 마커들을 이용한 췌장암 진단방법을 개시하고 있다. 하지만, 마커마다 그 진단 효율 및 정확성에서 큰 차이를 나타내므로, 효과가 더 우수한 마커를 발굴하고 이를 이용한 진단방법을 개발할 필요성이 있다.Korean Patent Application Laid-Open No. 2009-0003308 discloses a method for diagnosing pancreatic cancer by detecting the expression level of REG4 protein in an individual's blood sample, and Korean Patent Publication No. 2012-0009781 provides information necessary for diagnosing pancreatic cancer in an individual In order to do this, an analysis method for measuring the expression level of XIST RNA in cancer tissues isolated from individuals is disclosed in Korean Patent Publication No. 2007-0119250, a novel gene LBFL313 family that is differently expressed in human pancreatic cancer tissues compared to normal human pancreatic tissues. and US Patent Publication No. 2011/0294136 discloses a method for diagnosing pancreatic cancer using biomarkers such as keratin 8 protein. However, since each marker shows a large difference in its diagnostic efficiency and accuracy, there is a need to discover a marker with better effect and develop a diagnostic method using the marker.
상기와 같은 문제점을 해결하기 위하여, 본 발명자들은 췌장암을 진단할 수 있는 유전자 마커를 발굴하고자 정상인 및 췌장암 환자에서 혈장을 채취하고 이로부터 엑소좀(exosome)을 분리하여 엑소좀 단백질을 분석한 결과, 정상인에 비해 췌장암 환자에서만 유의하게 발현이 증가하는 유전자를 발견하였는 바, 이에 기초하여 본 발명을 완성하였다.In order to solve the above problems, the present inventors collected plasma from normal persons and pancreatic cancer patients in order to discover a genetic marker capable of diagnosing pancreatic cancer, and separated exosomes therefrom to analyze exosome proteins, The present invention was completed based on the discovery of a gene whose expression was significantly increased only in pancreatic cancer patients compared to normal persons.
이에 본 발명은 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질을 포함하는, 췌장암 진단용 마커 조성물을 제공한다.Accordingly, the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4) , NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) containing It provides a marker composition for diagnosing pancreatic cancer, comprising the mRNA of the gene or the protein encoded by the gene.
또한, 본 발명은 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질 수준을 측정하는 제제를 포함하는, 췌장암 진단용 조성물 및 상기 조성물을 포함하는 췌장암 진단용 키트를 제공한다.In addition, the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591). 4, including NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) It provides a composition for diagnosing pancreatic cancer, and a kit for diagnosing pancreatic cancer comprising the composition, comprising an agent for measuring the level of mRNA or protein encoded by the gene.
또한, 본 발명은 피검체 유래 생물학적 시료에서 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4) 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질의 수준을 측정하는 단계를 포함하는, 췌장암 진단을 위한 정보제공방법을 제공한다.In addition, in the present invention, MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte Differentiation antigen CD14; NM_002644.4) provides an information providing method for diagnosing pancreatic cancer, including measuring the level of mRNA of a gene or a protein encoded from the gene.
그러나 본 발명이 이루고자 하는 기술적 과제는 이상에서 언급한 과제에 제한되지 않으며, 언급되지 않은 또 다른 과제들은 아래의 기재로부터 당업자에게 명확하게 이해될 수 있을 것이다.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 skilled in the art from the following description.
본 발명의 목적을 달성하기 위하여, 본 발명은 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질을 포함하는, 췌장암 진단용 마커 조성물을 제공한다.In order to achieve the object of the present invention, the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 ( Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor) ; NM_002644.4) provides a marker composition for diagnosing pancreatic cancer, including mRNA of a gene or a protein encoded by the gene.
본 발명의 일 구현예에서, 상기 마커 조성물은 HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질을 더 포함할 수 있다.In one embodiment of the present invention, the marker composition is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277) .2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX) ; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643. 1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain- fatty-acid-CoA ligase 1; 1, NM_001381885.1, NM_001381886.1, NM_00138188 7.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) It may further include mRNA of one or more genes selected from or a protein encoded by the gene.
또한, 본 발명은 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질 수준을 측정하는 제제를 포함하는, 췌장암 진단용 조성물 및 이를 포함하는 췌장암 진단용 키트를 제공한다. In addition, the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591). 4, including NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) To provide a composition for diagnosing pancreatic cancer and a kit for diagnosing pancreatic cancer comprising the same, comprising an agent for measuring the level of mRNA or protein encoded by the gene.
본 발명의 일 구현예에서, 상기 췌장암 진단용 조성물은 HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질 수준을 측정하는 제제를 더 포함할 수 있다.In one embodiment of the present invention, the composition for diagnosing pancreatic cancer is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein) IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643 .1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain) -fatty-acid-CoA ligase 1;NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM .1, NM_001381885.1, NM_001381886.1, NM_001 381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) It may further comprise an agent for measuring the mRNA level of one or more genes selected from or a protein encoded by the gene.
본 발명의 다른 구현예에서, 상기 조성물은 췌장암을 조기에 진단할 수 있다.In another embodiment of the present invention, the composition can diagnose pancreatic cancer early.
또한, 본 발명은 피검체 유래 생물학적 시료에서 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4) 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질의 수준을 측정하는 단계를 포함하는, 췌장암 진단을 위한 정보제공방법을 제공한다.In addition, in the present invention, MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte Differentiation antigen CD14; NM_002644.4) provides an information providing method for diagnosing pancreatic cancer, including measuring the level of mRNA of a gene or a protein encoded from the gene.
본 발명의 일 구현예에서, 상기 방법은 HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질 수준을 측정하는 단계를 더 포함할 수 있다.In one embodiment of the present invention, the method comprises HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277. 2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1) ), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain-fatty) -acid-CoA ligase 1;NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1 , NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) It may further comprise the step of measuring the mRNA level of one or more genes selected from or a protein encoded from the gene.
본 발명의 다른 구현예에서, 상기 생물학적 시료는 혈액 또는 혈장 유래 엑소좀일 수 있다.In another embodiment of the present invention, the biological sample may be exosomes derived from blood or plasma.
본 발명의 또 다른 구현예에서, 상기 mRNA 수준은 차세대 염기서열 분석(Next generation sequencing; NGS), 중합효소연쇄반응(PCR), 역전사 중합효소연쇄반응(RT-PCR), 실시간 중합효소연쇄반응(Real-time PCR), RNase 보호 분석법(RNase protection assay; RPA), 마이크로어레이(microarray), 및 노던 블롯팅(northern blotting)으로 이루어진 군으로부터 선택되는 1종 이상의 방법을 통해 측정될 수 있다.In another embodiment of the present invention, the mRNA level is determined by next generation sequencing (NGS), polymerase chain reaction (PCR), reverse transcription polymerase chain reaction (RT-PCR), real-time polymerase chain reaction ( Real-time PCR), RNase protection assay (RPA), microarray, and northern blotting (northern blotting) can be measured through one or more methods selected from the group consisting of.
본 발명의 또 다른 구현예에서, 상기 단백질 수준은 웨스턴 블롯팅(western blotting), 방사선면역분석법(radioimmunoassay; RIA), 방사 면역 확산법(radioimmunodiffusion), 효소면역분석법(ELISA), 면역침강법(immunoprecipitation), 유세포분석법(flow cytometry), 면역형광염색법(immunofluorescence), 오우크테로니(ouchterlony), 보체 고정 분석법(complement fixation assay), 및 단백질 칩(protein chip)으로 이루어진 군으로부터 선택되는 1종 이상의 방법을 통해 측정될 수 있다.In another embodiment of the present invention, the protein level is determined by western blotting, radioimmunoassay (RIA), radioimmunodiffusion, enzyme immunoassay (ELISA), immunoprecipitation. , flow cytometry, immunofluorescence, ouchterlony, complement fixation assay, and at least one method selected from the group consisting of a protein chip can be measured through
본 발명자들은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질을 분석하여, 정상인에 비해 췌장암 환자에서 유의적으로 발현이 증가한 유전자를 췌장암 진단용 바이오마커로 발굴하였는바, 상기 유전자 마커들은 췌장암을 조기에 진단할 수 있어 임상에 효율적으로 이용될 것으로 기대된다.The present inventors analyzed the protein in exosomes isolated from plasma of normal people and pancreatic cancer patients, and discovered a gene with significantly increased expression in pancreatic cancer patients compared to normal people as biomarkers for diagnosing pancreatic cancer. It is expected to be used effectively in clinical practice.
도 1 내지 35는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질의 발현 수준을 분석한 것으로, 도면에 나타난 막대는 각 샘플의 정량적 단백질 양을 나타내며, 그 중 파란색 막대는 각 그룹 샘플의 평균값과 표준편차를 나타낸 것이다(모든 그래프의 x축은 Quan Channels로서 좌측은 PC 또는 EPC, 우측은 control을 나타내며, y축은 Abundance [a.u.]로서 눈금당 10을 나타낸다).1 to 35 are analysis of the expression level of proteins in exosomes isolated from plasma of normal people and pancreatic cancer patients, and the bars shown in the figure represent the quantitative protein amounts of each sample, among which the blue bar is the average value of each group sample. and standard deviation (x-axis of all graphs is Quan Channels, left is PC or EPC, right is control, y-axis is Abundance [a.u.], 10 per division).
도 1은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 1a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 1b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.1 shows the results of confirming the expression level of MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. That is, Figure 1a shows the results of normal people and pan-pancreatic cancer patients, and Figure 1b shows the results of normal people and early-pancreatic cancer patients.
도 2는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 ENPEP (Glutamyl aminopeptidase; NCBI 접근(accession)번호 : NM_001977.4)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 2a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 2b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.Figure 2 shows the results of confirming the expression level of ENPEP (Glutamyl aminopeptidase; NCBI accession number: NM_001977.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients, Figure 2a shows normal people and plate- Figure 2b shows the results of patients with pancreatic cancer (Pan-pancreatic cancer), normal people and early-shows the results of patients with early pancreatic cancer.
도 3은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 3a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 3b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.3 shows the results of confirming the expression level of CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients, Figure 3a shows the results of normal people and pan-pancreatic cancer (Pan-pancreatic cancer) patients, Figure 3b shows the results of normal people and early-pancreatic cancer (Early pancreatic cancer) patients.
도 4는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 MARF1 (Meiosis regulator and mRNA stability factor 1; NCBI 접근(accession)번호 : NM_014647.4, NM_001184998.2, NM_001184999.2)의 발현 수준을 확인한 결과를 나타낸 것이다. Figure 4 shows the expression level of MARF1 (Meiosis regulator and mRNA stability factor 1; NCBI accession number: NM_014647.4, NM_001184998.2, NM_001184999.2) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. The confirmed results are shown.
도 5는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 PIGR (Polymeric immunoglobulin receptor; NCBI 접근(accession)번호 : NM_002644.4)의 발현 수준을 확인한 결과를 나타낸 것이다.5 shows the results of confirming the expression level of PIGR (Polymeric immunoglobulin receptor; NCBI accession number: NM_002644.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 6은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3)의 발현 수준을 확인한 결과를 나타낸 것이다. 6 shows the results of confirming the expression level of HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 7은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 CLCA1 (Calcium-activated chloride channel regulator 1; NCBI 접근(accession)번호 : NM_001285.4)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 7a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 7b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.7 shows the results of confirming the expression level of CLCA1 (Calcium-activated chloride channel regulator 1; NCBI accession number: NM_001285.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients, FIG. 7a is a normal person and pan-pancreatic cancer (Pan-pancreatic cancer) patient results, Figure 7b is a normal person and early - shows the results of pancreatic cancer (Early pancreatic cancer) patients.
도 8은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 LACRT (Extracellular glycoprotein lacritin; NCBI 접근(accession)번호 : NM_033277.2)의 발현 수준을 확인한 결과를 나타낸 것이다. 8 shows the results of confirming the expression level of LACRT (Extracellular glycoprotein lacritin; NCBI accession number: NM_033277.2) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 9는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4)의 발현 수준을 확인한 결과를 나타낸 것이다. 9 is a result of confirming the expression level of LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients; is shown.
도 10은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 OSTF1 (Osteoclast-stimulating factor 1; NCBI 접근(accession)번호 : NM_012383.5)의 발현 수준을 확인한 결과를 나타낸 것이다. 10 shows the results of confirming the expression level of OSTF1 (Osteoclast-stimulating factor 1; NCBI accession number: NM_012383.5) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 11은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 GP9 (Platelet glycoprotein IX; NCBI 접근(accession)번호 : NM_000174.5)의 발현 수준을 확인한 결과를 나타낸 것이다. 11 shows the results of confirming the expression level of GP9 (Platelet glycoprotein IX; NCBI accession number: NM_000174.5) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 12는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 PSMA3 (Proteasome subunit alpha type-3; NCBI 접근(accession)번호 : NM_002788.4, NM_152132.3)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 12a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 12b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.12 shows the results of confirming the expression level of PSMA3 (Proteasome subunit alpha type-3; NCBI accession number: NM_002788.4, NM_152132.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. , Figure 12a shows the results of normal people and pan-pancreatic cancer patients, and Figure 12b shows the results of normal people and early-pancreatic cancer patients.
도 13은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 MME (Neprilysin; NCBI 접근(accession)번호 : NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1)의 발현 수준을 확인한 결과를 나타낸 것이다. 13 shows MME (Neprilysin; NCBI accession number: NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643) proteins in exosomes isolated from plasma of normal people and pancreatic cancer patients. 1) shows the result of confirming the expression level.
도 14는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NCBI 접근(accession)번호 : NM_006759.4, NM_001001521.2)의 발현 수준을 확인한 결과를 나타낸 것이다. Figure 14 shows the results of confirming the expression level of UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NCBI accession number: NM_006759.4, NM_001001521.2) in the protein in the exosome isolated from the plasma of normal people and pancreatic cancer patients. it has been shown
도 15는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14)의 발현 수준을 확인한 결과를 나타낸 것이다. 15 shows the results of confirming the expression level of ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 16은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NCBI 접근(accession)번호 : NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 16a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 16b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.16 shows ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NCBI accession number: NM_001995.5, NM_001286708.2, NM_001381877.1, Expression of NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2) expression As a result of confirming the level, FIG. 16a shows the results of normal people and pan-pancreatic cancer patients, and FIG. 16b shows the results of normal people and early-pancreatic cancer patients.
도 17은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 AHSG (Alpha-2-HS-glycoprotein; NCBI 접근(accession)번호 : NM_001622.4)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 17a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 17b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.17 shows the results of confirming the expression level of AHSG (Alpha-2-HS-glycoprotein; NCBI accession number: NM_001622.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients, FIG. 17a Figure 17b shows the results of normal people and pan-pancreatic cancer patients, and Fig. 17b shows the results of normal people and early-pancreatic cancer patients.
도 18은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 VASP (Vasodilator-stimulated phosphoprotein; NCBI 접근(accession)번호 : NM_003370.4)의 발현 수준을 확인한 결과를 나타낸 것이다. 18 shows the results of confirming the expression level of VASP (Vasodilator-stimulated phosphoprotein; NCBI accession number: NM_003370.4) in exosome proteins isolated from plasma of normal persons and pancreatic cancer patients.
도 19는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 F10 (Coagulation factor X; NCBI 접근(accession)번호 : NM_000504.4, NM_001312674.2, NM_001312675.2)의 발현 수준을 확인한 결과를 나타낸 것이다. Figure 19 shows the result of confirming the expression level of F10 (Coagulation factor X; NCBI accession number: NM_000504.4, NM_001312674.2, NM_001312675.2) in the protein in the exosome isolated from the plasma of normal people and pancreatic cancer patients will be.
도 20은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 PSMB3 (Proteasome subunit beta type-3; NCBI 접근(accession)번호 : NM_002795.4)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 20a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 20b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.Figure 20 shows the result of confirming the expression level of PSMB3 (Proteasome subunit beta type-3; NCBI accession number: NM_002795.4) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients, Figure 20a is Figure 20b shows the results of normal people and pan-pancreatic cancer patients, and normal people and early-pancreatic cancer patients.
도 21은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 MMRN1 (Multimerin-1; NCBI 접근(accession)번호 : NM_007351.3)의 발현 수준을 확인한 결과를 나타낸 것이다. 21 shows the results of confirming the expression level of MMRN1 (Multimerin-1; NCBI accession number: NM_007351.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 22는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 HABP2 (Hyaluronan-binding protein 2; NCBI 접근(accession)번호 : NM_004132.5, NM_001177660.3)의 발현 수준을 확인한 결과를 나타낸 것이다. 22 shows the results of confirming the expression level of HABP2 (Hyaluronan-binding protein 2; NCBI accession number: NM_004132.5, NM_001177660.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 23은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 LAMB1 (Laminin subunit beta-1; NCBI 접근(accession)번호 : NM_002291.3)의 발현 수준을 확인한 결과를 나타낸 것이다. 23 shows the results of confirming the expression level of LAMB1 (Laminin subunit beta-1; NCBI accession number: NM_002291.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 24는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 MMP9 (Matrix metalloproteinase-9; NCBI 접근(accession)번호 : NM_004994.3)의 발현 수준을 확인한 결과를 나타낸 것이다. 24 shows the results of confirming the expression level of MMP9 (Matrix metalloproteinase-9; NCBI accession number: NM_004994.3) in the protein in the exosome isolated from the plasma of normal people and pancreatic cancer patients.
도 25는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 NAP1L1 (Nucleosome assembly protein 1-like 1; NCBI 접근(accession)번호 : NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 25a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 25b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.25 is NAP1L1 (Nucleosome assembly protein 1-like 1; NCBI accession number: NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2) shows the results of confirming the expression level, and FIG. 25a shows the results of normal people and pan-pancreatic cancer patients, and FIG. 25b shows the results of normal people and early-pancreatic cancer patients. will be.
도 26은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 AMY2A (Pancreatic alpha-amylase; NCBI 접근(accession)번호 : NM_000699.3)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 26a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 26b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.Figure 26 shows the results of confirming the expression level of AMY2A (Pancreatic alpha-amylase; NCBI accession number: NM_000699.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. Pan-pancreatic cancer (Pan-pancreatic cancer) patient results, Figure 26b shows the results of normal people and early-pancreatic cancer (Early pancreatic cancer) patients.
도 27은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 TPM1 (Tropomyosin alpha-1 chain; NCBI 접근(accession)번호 : NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 27a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 27b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.27 is TPM1 (Tropomyosin alpha-1 chain; NCBI accession number: NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366 in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. 6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1 NM_001365781.2, NM_001365782.1) shows the results of confirming the expression level, and FIG. 27a shows the results of normal people and pan-pancreatic cancer patients. FIG. 27b shows normal people and early-pancreatic cancer patients. shows the results of
도 28은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 CANX (Isoform 2 of Calnexin; NCBI 접근(accession)번호 : NM_001363994.1)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 28a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 28b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.Figure 28 shows the results of confirming the expression level of CANX (Isoform 2 of Calnexin; NCBI accession number: NM_001363994.1) in the protein in the exosomes isolated from the plasma of normal people and pancreatic cancer patients, Figure 28a is normal and Pan-pancreatic cancer (Pan-pancreatic cancer) patient results, Figure 28b shows the results of normal people and early-pancreatic cancer (Early pancreatic cancer) patients.
도 29는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NCBI 접근(accession)번호 : NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3)의 발현 수준을 확인한 결과를 나타낸 것으로, 도 29a는 정상인 및 판-췌장암(Pan-pancreatic cancer) 환자의 결과를 도 29b는 정상인 및 초기-췌장암(Early pancreatic cancer) 환자의 결과를 나타낸 것이다.29 shows GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NCBI accession number: NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. Shows the results of confirming the expression level, and FIG. 29a shows the results of normal people and pan-pancreatic cancer patients, and FIG. 29b shows the results of normal people and early-pancreatic cancer patients.
도 30은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 HLA-C (HLA class I histocompatibility antigen, C alpha chain; NCBI 접근(accession)번호 : NM_002117.6, NM_001243042.1)의 발현 수준을 확인한 결과를 나타낸 것이다. 30 shows the expression level of HLA-C (HLA class I histocompatibility antigen, C alpha chain; NCBI accession number: NM_002117.6, NM_001243042.1) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. The confirmed results are shown.
도 31은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NCBI 접근(accession)번호 : NM_000282.4, NM_001127692.3, NM_001178004.2)의 발현 수준을 확인한 결과를 나타낸 것이다. Figure 31 shows the expression level of PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NCBI accession number: NM_000282.4, NM_001127692.3, NM_001178004.2) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. shows the results of checking .
도 32는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 CHDR2 (Cadherin-related family member 2; NCBI 접근(accession)번호 : NM_001171976.2, NM_017675.5)의 발현 수준을 확인한 결과를 나타낸 것이다. Figure 32 shows the results of confirming the expression level of CHDR2 (Cadherin-related family member 2; NCBI accession number: NM_001171976.2, NM_017675.5) in the protein in the exosome isolated from the plasma of normal people and pancreatic cancer patients .
도 33은 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 POSTN (Periostin; NCBI 접근(accession)번호 : NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2)의 발현 수준을 확인한 결과를 나타낸 것이다. Figure 33 shows POSTN (Periostin; NCBI accession) number: NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666 in exosome proteins isolated from plasma of normal persons and pancreatic cancer patients. 2, NM_001286667.2, NM_001330517.2) shows the results of checking the expression level.
도 34는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 ANPEP (Aminopeptidase N; NCBI 접근(accession)번호 : NM_001150.3)의 발현 수준을 확인한 결과를 나타낸 것이다. 34 shows the results of confirming the expression level of ANPEP (Aminopeptidase N; NCBI accession number: NM_001150.3) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients.
도 35는 정상인 및 췌장암 환자의 혈장에서 분리한 엑소좀 내 단백질에서 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NCBI 접근(accession)번호 : NM_001122606.1)의 발현 수준을 확인한 결과를 나타낸 것이다.35 shows the results of confirming the expression level of LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NCBI accession number: NM_001122606.1) in exosome proteins isolated from plasma of normal people and pancreatic cancer patients. will be.
본 발명자들은 췌장암을 진단하기 위한 유전자 마커를 발굴하기 위해 정상인 및 췌장암 환자에서 혈장을 채취하고 이로부터 엑소좀(exosome)을 분리하여 엑소좀 단백질을 분석한 결과, 정상인에 비해 췌장암 환자에서만 유의하게 발현이 증가하는 유전자를 발견하였는 바, 이에 기초하여 본 발명을 완성하였다.In order to discover a gene marker for diagnosing pancreatic cancer, the present inventors collected plasma from normal persons and pancreatic cancer patients, separated exosomes from them, and analyzed the exosome proteins. As a result, compared to normal persons, expression was significantly expressed only in pancreatic cancer patients. The present invention was completed based on the discovery of this increasing gene.
이에, 본 발명은 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질을 포함하는, 췌장암 진단용 마커 조성물을 제공한다.Accordingly, the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591). 4, including NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) It provides a marker composition for diagnosing pancreatic cancer, comprising the mRNA of a gene or a protein encoded by the gene.
본 발명에 있어서, 상기 마커 조성물은 HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질을 더 포함할 수 있다.In the present invention, the marker composition is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2) , LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174. 5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain-fatty-acid -CoA ligase 1;NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001288586710.2, NM_0013818814.1 .1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) It may further include mRNA of one or more genes selected from or a protein encoded by the gene.
본 발명에서 사용되는 용어 “췌장암”은 췌장 세포가 정상적인 성장한계를 무시하고 분열 및 성장하는 공격(aggressive) 특성, 주위 조직에 침투하는 침투적(invasive) 특성 및 체내의 다른 부위로 퍼지는 전이적(metastatic) 특성을 갖는 세포에 의한 질병을 총칭하는 의미이다. As used herein, the term “pancreatic cancer” refers to an aggressive characteristic in which pancreatic cells divide and grow ignoring normal growth limits, an invasive characteristic that penetrates into surrounding tissues, and a metastatic (metastatic) characteristic that spreads to other parts of the body. Metastatic) refers to diseases caused by cells with characteristics.
본 발명에서 사용되는 용어 “진단(diagnosis)”이란 넓은 의미로는 환자의 병의 실태를 모든 면에 걸쳐서 판단하는 것을 의미한다. 판단의 내용은 병명, 병인, 병형, 경중, 병상의 상세한 양태 및 합병증의 유무 등이다. 본 발명에서 진단은 암의 발병 여부 및 진행단계 수준 등을 판단하는 것이다. As used herein, the term “diagnosis” in a broad sense means judging the actual condition of a patient's disease in all aspects. The content of the judgment is the name of the disease, etiology, type, severity, detailed mode of the disease, and the presence or absence of complications. Diagnosis in the present invention is to determine whether or not cancer is onset and the level of progression.
본 발명자들은 구체적인 실시예를 통해 췌장암 진단을 위한 신규 마커로써 본 발명에 따른 특정 유전자의 용도를 규명하였다.The present inventors identified the use of a specific gene according to the present invention as a novel marker for diagnosing pancreatic cancer through specific examples.
본 발명의 일 실시예에서는 정상인에 비해 췌장암 환자의 검체에서 특정 유전자(MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, AHSG, VASP, F10, PSMB3, MMRN1, HABP2, LAMB1, MMP9, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP 및 LAMP2)의 발현이 유의하게 증가되는 것을 확인하였다(실시예 2 참조).In one embodiment of the present invention, specific genes (MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, It was confirmed that the expression of AHSG, VASP, F10, PSMB3, MMRN1, HABP2, LAMB1, MMP9, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP and LAMP2) was significantly increased. (See Example 2).
상기 결과를 통해 특정 유전자(MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, AHSG, VASP, F10, PSMB3, MMRN1, HABP2, LAMB1, MMP9, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP 및 LAMP2) 또는 상기 유전자가 암호화하는 단백질이 췌장암 진단을 위한 마커로 이용될 수 있음을 유추할 수 있다.Through the above results, specific genes (MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, AHSG, VASP, F10, PSMB3, MMRN1, HABP2 , LAMB1, MMP9, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP and LAMP2) or the protein encoded by the gene can be inferred that it can be used as a marker for pancreatic cancer diagnosis can
본 발명의 다른 양태로서, 본 발명은 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질 수준을 측정하는 제제를 포함하는, 췌장암 진단용 조성물 및 상기 조성물을 포함하는 췌장암 진단용 키트를 제공한다.As another aspect of the present invention, the present invention provides MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation) antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644) .4) provides a composition for diagnosing pancreatic cancer and a kit for diagnosing pancreatic cancer comprising the composition, comprising an agent for measuring the level of mRNA or protein encoded by the gene comprising the gene.
본 발명에 있어서, 상기 췌장암 진단용 조성물은 HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)으로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질 수준을 측정하는 제제를 더 포함할 수 있다.In the present invention, the composition for diagnosing pancreatic cancer is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2) ), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174) .5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1) , UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain-fatty- acid-CoA ligase 1; NM_001381885.1, NM_001381886.1, NM_0013818 87.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) It may further comprise an agent for measuring the mRNA level of one or more genes selected from or a protein encoded by the gene.
본 발명에 있어서, 상기 조성물은 췌장암을 조기에 진단할 수 있다.In the present invention, the composition can diagnose pancreatic cancer early.
본 발명의 예측용 키트는 분석 방법에 적합한 한 종류 또는 그 이상의 다른 구성성분 조성물, 용액 또는 장치로 구성된다.The kit for prediction of the present invention consists of one or more other component compositions, solutions or devices suitable for the analysis method.
예컨대, 본 발명의 키트는 PCR을 수행하기 위해, 분석하고자 하는 시료로부터 유래된 게놈 DNA, 본 발명의 마커 유전자에 대해 특이적인 프라이머 세트, 적당량의 DNA 중합 효소, dNTP 혼합물, PCR 완충용액 및 물을 포함하는 키트일 수 있다. 상기 PCR 완충용액은 KCl, Tris-HCl 및 MgCl2를 함유할 수 있다. 이외에 PCR 산물의 증폭 여부를 확인할 수 있는 전기영동 수행에 필요한 구성 성분들이 본 발명의 키트에 추가로 포함될 수 있다.For example, the kit of the present invention contains genomic DNA derived from a sample to be analyzed, a primer set specific for the marker gene of the present invention, an appropriate amount of a DNA polymerase, a dNTP mixture, a PCR buffer, and water to perform PCR. It may be a kit comprising The PCR buffer solution may contain KCl, Tris-HCl and MgCl 2 . In addition, components necessary for performing electrophoresis that can confirm whether or not the PCR product is amplified may be additionally included in the kit of the present invention.
또한, 본 발명의 키트는 RT-PCR을 수행하기 위해 필요한 필수 요소를 포함하는 키트일 수 있다. RT-PCR 키트는 마커 유전자에 대한 특이적인 각각의 프라이머 쌍 외에도 테스트 튜브 또는 다른 적절한 컨테이너, 반응 완충액, 데옥시뉴클레오티드(dNTPs), Taq-폴리머레이즈 및 역전사 효소와 같은 효소, DNase, RNase 억제제, DEPC-수(DEPC-water), 멸균수 등을 포함할 수 있다. 또한 정량 대조군으로 사용되는 유전자에 특이적인 프라이머 쌍을 포함할 수 있다.In addition, the kit of the present invention may be a kit including essential elements necessary for performing RT-PCR. In addition to each primer pair specific for a marker gene, the RT-PCR kit includes a test tube or other suitable container, reaction buffer, deoxynucleotides (dNTPs), enzymes such as Taq-polymerase and reverse transcriptase, DNase, RNase inhibitors, DEPC -Water (DEPC-water), sterile water, etc. may be included. In addition, a primer pair specific for a gene used as a quantitative control may be included.
또한, 본 발명의 키트는 DNA 칩을 수행하기 위해 필요한 필수 요소를 포함하는 키트일 수 있다. DNA 칩 키트는, 유전자 또는 그의 단편에 해당하는 cDNA가 프로브로 부착되어 있는 기판을 포함하고, 기판은 정량구조 유전자 또는 그의 단편에 해당하는 cDNA를 포함할 수 있다. 또한, 본 발명의 키트는 본 발명의 마커 유전자가 고정화되어 있는 기판을 갖는 마이크로어레이 형태일 수 있다.In addition, the kit of the present invention may be a kit including essential elements necessary for performing a DNA chip. The DNA chip kit may include a substrate to which cDNA corresponding to a gene or fragment thereof is attached as a probe, and the substrate may include cDNA corresponding to a quantitative structural gene or fragment thereof. In addition, the kit of the present invention may be in the form of a microarray having a substrate on which the marker gene of the present invention is immobilized.
또한, 본 발명의 키트는 ELISA를 수행하기 위해 필요한 필수 요소를 포함하는 것을 특징으로 하는 키트일 수 있다. ELISA 키트는 마커 단백질에 대한 특이적인 항체를 포함하며, 상기 단백질 수준을 측정하는 제제를 포함한다. 상기 ELISA 키트는 "항원-항체 복합체"를 형성한 항체를 검출할 수 있는 시약, 예를 들면 표지된 2차 항체, 발색단(chromopores), 효소, 및 그의 기질을 포함할 수 있다. 또한, 정량 대조군 단백질에 특이적인 항체를 포함할 수 있다.In addition, the kit of the present invention may be a kit characterized in that it includes essential elements necessary for performing ELISA. The ELISA kit includes an antibody specific for a marker protein, and an agent for measuring the protein level. The ELISA kit may include a reagent capable of detecting an antibody that has formed an "antigen-antibody complex", for example, a labeled secondary antibody, chromopores, an enzyme, and a substrate thereof. In addition, an antibody specific for the quantitative control protein may be included.
본 발명의 또 다른 양태로서, 본 발명은 피검체 유래 생물학적 시료에서 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질의 수준을 측정하는 단계를 포함하는, 췌장암 진단을 위한 정보제공방법을 제공한다.As another aspect of the present invention, in the biological sample derived from the subject, MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977. 4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) provides an information providing method for diagnosing pancreatic cancer, including measuring the level of mRNA of a gene or a protein encoded from the gene.
본 발명에 있어서, 상기 방법은 HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)으로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질 수준을 측정하는 단계를 더 포함할 수 있다.In the present invention, the method is HIST1H2AB (Histone H2A type 1-B/E; NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5) ), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain-fatty-acid- CoA ligase 1 ; 1, NM_001381886.1, NM_001381887.1, NM_ 001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) It may further comprise the step of measuring the mRNA level of one or more genes selected from or a protein encoded from the gene.
본 발명에서 사용되는 용어 “췌장암 진단을 위한 정보제공방법”은 진단을 위한 예비적 단계로써 췌장암의 진단을 위하여 필요한 객관적인 기초정보를 제공하는 것이며 의사의 임상학적 판단 또는 소견은 제외된다. The term “information providing method for diagnosing pancreatic cancer” used in the present invention provides objective basic information necessary for diagnosing pancreatic cancer as a preliminary step for diagnosis, and the clinical judgment or opinion of a doctor is excluded.
상기 피검체 유래의 생물학적 시료는 조직, 세포, 전혈, 혈액, 타액, 객담, 뇌척수액 및 뇨 등을 포함할 수 있으며, 바람직하게는 혈액 또는 혈장일 수 있고, 더욱 바람직하게는 혈장 유래 엑소좀일 수 있으나 이에 제한되는 것은 아니다.The subject-derived biological sample may include tissues, cells, whole blood, blood, saliva, sputum, cerebrospinal fluid and urine, preferably blood or plasma, and more preferably plasma-derived exosomes. However, the present invention is not limited thereto.
상기 mRNA 수준은 차세대 염기서열 분석(Next generation sequencing; NGS), 중합효소연쇄반응(PCR), 역전사 중합효소연쇄반응(RT-PCR), 실시간 중합효소연쇄반응(Real-time PCR), RNase 보호 분석법(RNase protection assay; RPA), 마이크로어레이(microarray), 및 노던 블롯팅(northern blotting)으로 이루어진 군으로부터 선택되는 1종 이상의 방법을 통해 측정되는 것일 수 있으나, 이에 제한되는 것은 아니다. The mRNA level is determined by next generation sequencing (NGS), polymerase chain reaction (PCR), reverse transcription polymerase chain reaction (RT-PCR), real-time polymerase chain reaction (Real-time PCR), RNase protection assay (RNase protection assay; RPA), microarray (microarray), and may be measured through one or more methods selected from the group consisting of northern blotting (northern blotting), but is not limited thereto.
상기 단백질 수준은 웨스턴 블롯팅(western blotting), 방사선면역분석법(radioimmunoassay; RIA), 방사 면역 확산법(radioimmunodiffusion), 효소면역분석법(ELISA), 면역침강법(immunoprecipitation), 유세포분석법(flow cytometry), 면역형광염색법(immunofluorescence), 오우크테로니(ouchterlony), 보체 고정 분석법(complement fixation assay), 및 단백질 칩(protein chip)으로 이루어진 군으로부터 선택되는 1종 이상의 방법을 통해 측정되는 것일 수 있으나, 이에 제한되는 것은 아니다. The protein level was determined by western blotting, radioimmunoassay (RIA), radioimmunodiffusion, enzyme immunoassay (ELISA), immunoprecipitation, flow cytometry, and immunity. It may be measured by at least one method selected from the group consisting of immunofluorescence, ouchterlony, complement fixation assay, and protein chip, but is limited thereto it's not going to be
이하, 본 발명의 이해를 돕기 위하여 바람직한 실시예를 제시한다. 그러나 하기의 실시예는 본 발명을 보다 쉽게 이해하기 위하여 제공되는 것일 뿐, 하기 실시예에 의해 본 발명의 내용이 한정되는 것은 아니다.Hereinafter, preferred examples are presented to help the understanding of the present invention. However, the following examples are only provided for easier understanding of the present invention, and the contents of the present invention are not limited by the following examples.
[실시예][Example]
실시예 1. 혈장 내 엑소좀의 특이 단백질 분석Example 1. Analysis of specific proteins of exosomes in plasma
1-1. 엑소좀 샘플 준비1-1. Exosome Sample Preparation
정상인 및 췌장암 환자의 혈액을 BD Vacutainer Plastic K2EDTA Tube (BD, Cat.no 367525)에 채취한 후 2시간 이내 2,500 rpm, 4 ℃ 조건에서 20분간 원심 분리하여 혈장을 분리한다. 상층 혈장을 Protein LoBind Tube (Eppendorf, Cat.no 0030108116)에 500 ㎕씩 분주하고 -80 ℃에 보관하였다. Blood from normal and pancreatic cancer patients is collected in BD Vacutainer Plastic K2EDTA Tube (BD, Cat.no 367525), and plasma is separated by centrifugation within 2 hours at 2,500 rpm and 4°C for 20 minutes. The supernatant plasma was aliquoted into Protein LoBind Tubes (Eppendorf, Cat.no 0030108116) by 500 μl and stored at -80°C.
혈장 시료를 3,000 g, 4 ℃ 조건에서 15분간 원심 분리하여 혈장 내 세포 및 세포 찌꺼기(cell debris)를 제거하였다. 다음 혈장 시료 500 ㎕에 Exo2DTM for Protein assay (EXOSOMEplus) 250 ㎕, 2:1 비율로 첨가하여 Aqueous Two-Phase System을 기반으로 엑소좀을 분리 후, Halt™ Protease and Phosphatase Inhibitor Cocktail, EDTA-Free(Thermo scientific)을 첨가한 200 ㎕ 용량의 PBS에 엑소좀을 재부유(Resuspending)시켜 엑소좀 샘플을 준비하였다. 시료 분석 전까지 -80 ℃에 보관하였다.Plasma samples were centrifuged at 3,000 g and 4 °C for 15 minutes to remove cells and cell debris in the plasma. Then, 250 μl of Exo2D TM for Protein assay (EXOSOMEplus) was added to 500 μl of plasma sample at a 2:1 ratio to separate exosomes based on the Aqueous Two-Phase System, and then, Halt™ Protease and Phosphatase Inhibitor Cocktail, EDTA-Free ( The exosome samples were prepared by resuspending the exosomes in 200 μl of PBS with the addition of Thermo scientific). Samples were stored at -80 °C until analysis.
1-2. 엑소좀 단백질 정량분석1-2. Exosome protein quantitative analysis
상기 실시예 1-1에서 제조된 혈장 엑소좀 샘플 30 ㎕에 프로테아제 및 포스페타아제(1X)를 포함하는 RIPA 버퍼 30 ㎕를 첨가하고 충분히 혼합시킨 후 원심분리 하였다. 추출된 엑소좀 단백질은 Bovine Serum Albumin (BSA) Standard Set (BIO-RAD)와 PierceTM BCA Protein Assay (Thermo scientific)을 이용하여 농도 측정 및 정량분석을 진행하였다.30 μl of RIPA buffer containing protease and phosphatase (1X) was added to 30 μl of the plasma exosome sample prepared in Example 1-1, mixed thoroughly, and then centrifuged. The extracted exosome protein was subjected to concentration measurement and quantitative analysis using Bovine Serum Albumin (BSA) Standard Set (BIO-RAD) and Pierce TM BCA Protein Assay (Thermo scientific).
다음으로, 엑소좀 단백질 20 ㎍을 SDS loading buffer와 섞은 후 10% Sodium Dodecyl Sulfate Polyacrylamide Gel Eletrophoresis (SDS-PAGE) 기법으로 전기영동 하여 단백질들을 분자량 크기에 따라 분리시켰다. 전기영동된 젤을 InstantBlue쪠 (Coomassie blue stanning) 시약에 담가 상온에서 15분간 염색한 후 1차적으로 단백질 검출 및 분포 상황을 확인하였다.Next, 20 μg of exosome protein was mixed with SDS loading buffer and then electrophoresed using 10% Sodium Dodecyl Sulfate Polyacrylamide Gel Eletrophoresis (SDS-PAGE) technique to separate proteins according to molecular weight size. After immersing the electrophoresed gel in InstantBlue (Coomassie blue stanning) reagent and staining at room temperature for 15 minutes, protein detection and distribution were first checked.
1-3. S-Trap쪠 Mini Spin Column (ProtiFi)을 이용한 단백질 digestion1-3. Protein digestion using S-Trap Mini Spin Column (ProtiFi)
(1) 시료에 5% SDS, 50 mM TAEB (pH7.55), 1x PPIC을 추가하여 용해하였다. 인산을 이용하여 pH 값을 맞춰주었다.(1) 5% SDS, 50 mM TAEB (pH7.55), and 1x PPIC were added to the sample and dissolved. The pH value was adjusted using phosphoric acid.
(2) BCA assay을 이용하여 정량한 단백질 시료 300 ㎍을 취하였다. Digestion할 단백질에 최종농도가 20 mM이 되게 DTT를 추가하여 95 ℃에서 10분 동안 알킬화시켰다.(2) 300 ㎍ of a protein sample quantified using BCA assay was taken. DTT was added to the protein to be digested to a final concentration of 20 mM and alkylated at 95 °C for 10 minutes.
(3) 실온에서 식혀준 후 최종농도가 40 mM 되게 IAA을 추가하여 어두운 곳에서 30분간 reduction 시켰다. 다음 12% 인산을 1.2% 인산으로 되게끔 섞어주었다. (3) After cooling at room temperature, IAA was added so that the final concentration was 40 mM and reduced for 30 minutes in the dark. Then, 12% phosphoric acid was mixed to make 1.2% phosphoric acid.
(4) S-Trap binding buffer (90% methanol 100 mM TEAB (pH 7.1)) 350 ㎕ 추가한 후 모두 S-Trap spin column으로 옮겼다. 다음 원심 분리기를 이용하여 4,000 g에서 30초간 원심 분리하였다.(4) 350 μl of S-Trap binding buffer (90% methanol, 100 mM TEAB (pH 7.1)) was added, and all were transferred to an S-Trap spin column. Then, centrifugation was performed at 4,000 g for 30 seconds using a centrifuge.
(5) S-Trap buffer 400 ㎕을 S-Trap spin column에 추가한 후 원심 분리기를 이용하여 4,000 g에서 30초간 원심 분리하였다. 이를 3회 반복하였다.(5) 400 μl of S-Trap buffer was added to the S-Trap spin column and centrifuged at 4,000 g for 30 seconds using a centrifuge. This was repeated 3 times.
(6) S-Trap spin column을 새로운 2 ml sample tube로 옮긴 후, 단백질과1:25 비율의 trypsin을 50 mM TEAB buffer 125 ㎕에 녹여 S-Trap spin column에 추가하여 37 ℃에서 16시간 동안 incubation 하였다.(6) After transferring the S-Trap spin column to a new 2 ml sample tube, dissolve the protein and trypsin in a ratio of 1:25 in 125 μl of 50 mM TEAB buffer, add it to the S-Trap spin column, and incubate at 37°C for 16 hours. did
(7) 50 mM TEAB buffer를 80 ㎕ 추가하여 1,000 g에서 60초간 원심 분리하였다. 다음 0.2% FA를 80 ㎕ 추가하여 1,000 g에서 60초간 원심 분리하였다. 마지막으로 elution buffer (50% ACN, 0.2% FA)를 80 ㎕ 추가하여 4,000 g에서 60초간 원심 분리하고 용리한 시료를 동결 건조하였다.(7) 80 μl of 50 mM TEAB buffer was added and centrifuged at 1,000 g for 60 seconds. Then, 80 μl of 0.2% FA was added and centrifuged at 1,000 g for 60 seconds. Finally, 80 μl of elution buffer (50% ACN, 0.2% FA) was added, centrifuged at 4,000 g for 60 seconds, and the eluted sample was freeze-dried.
1-4. 프로테오믹스 분석1-4. Proteomics analysis
Digestion 과정을 거쳐 준비된 단백질 펩티드 시료를 Q Exactive Plus LC-MS/MS System 질량분석기(Thermo Scientific)를 이용하여 단백질체 분석을 하였다. 각 시료마다 두 번 반복(duplicate)하여 측정하였고, 기기에서 생성된 데이터는 Proteome DiscovererTM 2.2 (Thermo Scientific) 소프트웨어를 이용하여 데이터베이스 검색, 종합적인 단백질 및 펩티드의 식별, 특성 분석과 정량 분석 및 통계분석을 진행하였다.Protein peptide samples prepared through the digestion process were analyzed for proteomic bodies using Q Exactive Plus LC-MS/MS System mass spectrometer (Thermo Scientific). Each sample was measured in duplicate, and the data generated by the instrument was analyzed using Proteome Discoverer TM 2.2 (Thermo Scientific) software for database search, comprehensive protein and peptide identification, characterization and quantitative analysis and statistical analysis. was carried out.
실시예 2. 췌장암 특이 유전자 발현 확인Example 2. Confirmation of pancreatic cancer-specific gene expression
정상인 4명 및 판-췌장암(Pan pancreatic cancer) 환자 10명의 혈장을 수득하고, 실시예 1의 방법을 통해 혈장 엑소좀 단백질을 추출한 후, 췌장암 환자에서 유의적인 발현을 나타내는 유전자를 확인하였다.After obtaining plasma from 4 normal people and 10 patients with pancreatic cancer, and extracting plasma exosome proteins through the method of Example 1, genes showing significant expression in pancreatic cancer patients were identified.
그 결과, 도 1a, 2a, 3a, 4 내지 6, 7a, 8 내지 11, 12a, 13 내지 15, 16a, 17a, 18, 19, 20a, 21 내지 24, 25a, 26a, 27a, 28a 및 29a에 나타낸 바와 같이 MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, AHSG, VASP 및 F10 유전자의 발현이 정상인에 비해 판-췌장암(Pan pancreatic cancer)환자의 검체에서만 유의적으로 증가되어 있는 것을 확인하였다.As a result, in Figures 1a, 2a, 3a, 4-6, 7a, 8-11, 12a, 13-15, 16a, 17a, 18, 19, 20a, 21-24, 25a, 26a, 27a, 28a and 29a As shown, the expression of MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, AHSG, VASP, and F10 genes was higher in pancreatic cancer compared to normal subjects. (Pan pancreatic cancer) was found to be significantly increased only in the sample of the patient.
나아가, 정상인 4명 및 초기-췌장암(Early pancreatic cancer) 환자 3명의 혈장을 수득하고 실시예 1의 방법을 통해 혈장 엑소좀 단백질을 추출한 후, 췌장암 환자에서 유의적인 발현을 나타내는 유전자를 확인하였다.Furthermore, after obtaining plasma from 4 normal people and 3 patients with early pancreatic cancer and extracting plasma exosome proteins through the method of Example 1, genes showing significant expression in pancreatic cancer patients were identified.
그 결과, 도 1b, 2b, 3b, 7b, 12b, 16b, 17b, 20b, 25b, 26b, 27b, 28b, 29b, 30 내지 35에 나타낸 바와 같이 MAN1A1, ENPEP, CD14, CLCA1, PSMA3, ACSL1, AHSG, PSMB3, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP 및 LAMP2 유전자의 발현이 정상인에 비해 초기-췌장암(Early pancreatic cancer)환자의 검체에서만 유의적으로 증가되어 있는 것을 확인하였다.As a result, as shown in FIGS. 1b, 2b, 3b, 7b, 12b, 16b, 17b, 20b, 25b, 26b, 27b, 28b, 29b, 30 to 35, MAN1A1, ENPEP, CD14, CLCA1, PSMA3, ACSL1, AHSG , PSMB3, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP and LAMP2 gene expression was significantly increased only in samples from patients with early pancreatic cancer compared to normal subjects. confirmed that there is.
상기 결과로부터, 췌장암 환자 유래 혈장 엑소좀에서 검출된 특정 유전자(MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, AHSG, VASP, F10, PSMB3, MMRN1, HABP2, LAMB1, MMP9, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP 및 LAMP2)가 췌장암을 진단하는데 유용하게 이용될 수 있을 것으로 판단된다.From the above results, specific genes (MAN1A1, ENPEP, CD14, MARF1, PIGR, HIST1H2AB, CLCA1, LACRT, LTBP1, OSTF1, GP9, PSMA3, MME, UGP2, ANK1, ACSL1, AHSG, VASP, F10, PSMB3, MMRN1, HABP2, LAMB1, MMP9, NAP1L1, AMY2A, TPM1, CANX, GP2, HLA-C, PCCA, CDHR2, POSTN, ANPEP and LAMP2) could be usefully used to diagnose pancreatic cancer is judged
상기 진술한 본 발명의 설명은 예시를 위한 것이며, 본 발명이 속하는 기술분야의 통상의 지식을 가진 자는 본 발명의 기술적 사상이나 필수적인 특징을 변경하지 않고서 다른 구체적인 형태로 쉽게 변형이 가능하다는 것을 이해할 수 있을 것이다. 그러므로 이상에서 기술한 실시예들은 모든 면에서 예시적인 것이며 한정적이 아닌 것으로 이해해야만 한다.The description of the present invention stated above is for illustration, and those of ordinary skill in the art to which the present invention pertains can understand that it can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. There will be. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive.

Claims (11)

  1. MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질을 포함하는, 췌장암 진단용 마커 조성물.MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3) , NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) mRNA or A marker composition for diagnosing pancreatic cancer, comprising a protein encoded by the gene.
  2. 제1항에 있어서, The method of claim 1,
    상기 마커 조성물은 HIST1H2AB (Histone H2A type 1-B/E; NCBI 접근(accession)번호 : NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질을 더 포함하는 것을 특징으로 하는, 마커 조성물.The marker composition is HIST1H2AB (Histone H2A type 1-B/E; NCBI accession number: NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277). 2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1) ), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain-fatty) -acid-CoA ligase 1;NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1 , NM_001381885.1, NM_001381886.1, NM_ 001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) One or more genes selected from mRNA or a protein encoded by the gene, characterized in that it further comprises a marker composition.
  3. MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4)을 포함하는 유전자의 mRNA 또는 상기 유전자가 암호화하는 단백질 수준을 측정하는 제제를 포함하는, 췌장암 진단용 조성물.MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3) , NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) mRNA or A composition for diagnosing pancreatic cancer, comprising an agent for measuring the level of the protein encoded by the gene.
  4. 제3항에 있어서,4. The method of claim 3,
    상기 조성물은 HIST1H2AB (Histone H2A type 1-B/E; NCBI 접근(accession)번호 : NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질 수준을 측정하는 제제를 더 포함하는 것을 특징으로 하는, 췌장암 진단용 조성물.The composition is HIST1H2AB (Histone H2A type 1-B/E; NCBI accession number: NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2) ), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174) .5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1) , UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain-fatty- acid-CoA ligase 1; NM_001381885.1, NM_001381886.1, NM_001 381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) A composition for diagnosing pancreatic cancer, characterized in that it further comprises an agent for measuring the mRNA or protein level of one or more genes selected from the gene.
  5. 제3항에 있어서,4. The method of claim 3,
    상기 조성물은 췌장암을 조기에 진단할 수 있는 것을 특징으로 하는, 조성물.The composition is characterized in that it can diagnose pancreatic cancer early, the composition.
  6. 제3항의 조성물을 포함하는, 췌장암 진단용 키트.A kit for diagnosing pancreatic cancer, comprising the composition of claim 3.
  7. 피검체 유래 생물학적 시료에서 MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI 접근(accession)번호 : NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591.4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) 및 PIGR (Polymeric immunoglobulin receptor; NM_002644.4) 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질의 수준을 측정하는 단계를 포함하는, 췌장암 진단을 위한 정보제공방법.In biological samples derived from the subject, MAN1A1 (Mannosyl-oligosaccharide 1,2-alpha-mannosidase IA; NCBI accession number: NM_005907.4), ENPEP (Glutamyl aminopeptidase; NM_001977.4), CD14 (Monocyte differentiation antigen CD14; NM_000591) .4, NM_001040021.3, NM_001174104.2, NM_001174105.2), MARF1 (Meiosis regulator and mRNA stability factor 1; NM_014647.4, NM_001184998.2, NM_001184999.2) and PIGR (Polymeric immunoglobulin receptor; NM_002644.4) genes An information providing method for diagnosing pancreatic cancer, comprising the step of measuring the level of the mRNA or protein encoded by the gene.
  8. 제7항에 있어서, 8. The method of claim 7,
    상기 방법은 HIST1H2AB (Histone H2A type 1-B/E; NCBI 접근(accession)번호 : NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB 접근(accession)번호 : Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174.5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1), UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB 접근(accession)번호 : P16157-14), ACSL1 (Long-chain-fatty-acid-CoA ligase 1; NM_001995.5, NM_001286708.2, NM_001381877.1, NM_001381878.1, NM_001381879.1, NM_001381880.1, NM_001381881.1, NM_001381882.1, NM_001381883.1, NM_001286710.2, NM_001381884.1, NM_001381885.1, NM_001381886.1, NM_001381887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994.3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha-1 chain; NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_001330346.2, NM_001330351.2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major glycoprotein GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936.2, NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) 및 LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1)로 이루어진 군에서 선택된 하나 이상의 유전자의 mRNA 또는 상기 유전자로부터 코딩되는 단백질 수준을 측정하는 단계를 더 포함하는 것을 특징으로 하는, 정보제공방법.The method is HIST1H2AB (Histone H2A type 1-B/E; NCBI accession number: NM_003513.3), CLCA1 (Calcium-activated chloride channel regulator 1; NM_001285.4), LACRT (Extracellular glycoprotein lacritin; NM_033277.2) ), LTBP1 (Isoform 4 of Latent-transforming growth factor beta-binding protein 1; UniProtKB accession number: Q14766-4), OSTF1 (Osteoclast-stimulating factor 1; NM_012383.5), GP9 (Platelet glycoprotein IX; NM_000174) .5), PSMA3 (Proteasome subunit alpha type-3; NM_002788.4, NM_152132.3), MME (Neprilysin; NM_000902.5, NM_007288.3, NM_007289.4, NM_007287.4, NM_001354642.2, NM_001354643.1) , UGP2 (UTP-glucose-1-phosphate uridylyltransferase; NM_006759.4, NM_001001521.2), ANK1 (Isoform Er13 of Ankyrin-1; UniProtKB accession number: P16157-14), ACSL1 (Long-chain-fatty- acid-CoA ligase 1; NM_001381885.1, NM_001381886.1, NM_0013 81887.1, NM_001286711.2), AHSG (Alpha-2-HS-glycoprotein; NM_001622.4), VASP (Vasodilator-stimulated phosphoprotein; NM_003370.4), F10 (Coagulation factor X; NM_000504.4, NM_001312674.2, NM_001312675.2), PSMB3 (Proteasome subunit beta type-3; NM_002795.4), MMRN1 (Multimerin-1; NM_007351.3), HABP2 (Hyaluronan-binding protein 2; NM_004132.5, NM_001177660.3), LAMB1 (Laminin subunit beta-1; NM_002291.3), MMP9 (Matrix metalloproteinase-9; NM_004994). 3), NAP1L1 (Nucleosome assembly protein 1-like 1; NM_139207.5, NM_004537.7, NM_001330231.2, NM_001307924.3, NM_001330232.2), AMY2A (Pancreatic alpha-amylase; NM_000699.3), TPM1 (Tropomyosin alpha -1 chain: NM_001018005.2, NM_001018007.2, NM_001018004.2, NM_001018006.2, NM_000366.6, NM_001018008.2, NM_001018020.2, NM_001301244.2, NM_001301289.2, NM_001330344.2, NM_0013303411. 2, NM_001365776.1, NM_001365777.1, NM_001365778.1, NM_001365779.1, NM_001365780.1, NM_001365781.2, NM_001365782.1), CANX (Isoform 2 of Calnexin; NM_001363994.1), GP2 (Pancreatic secretory granule membrane major) glycopro tain GP2; NM_001007240.3, NM_001502.4, NM_001007241.3, NM_001007242.3), HLA-C (HLA class I histocompatibility antigen, C alpha chain; NM_002117.6, NM_001243042.1), PCCA (Propionyl-CoA carboxylase alpha chain, mitochondrial ; NM_000282.4, NM_001127692.3, NM_001178004.2), CHDR2 (Cadherin-related family member 2; NM_001171976.2, NM_017675.5), POSTN (Periostin; NM_006475.3, NM_001135934.2, NM_001135935.2, NM_001135936). 2, group consisting of NM_001286665.2, NM_001286666.2, NM_001286667.2, NM_001330517.2), ANPEP (Aminopeptidase N; NM_001150.3) and LAMP2 (Isoform LAMP-2C of Lysosome-associated membrane glycoprotein 2; NM_001122606.1) Method for providing information, characterized in that it further comprises the step of measuring the mRNA level of one or more genes selected from or a protein encoded from the gene.
  9. 제7항에 있어서,8. The method of claim 7,
    상기 생물학적 시료는 혈액 또는 혈장 유래 엑소좀인 것을 특징으로 하는, 정보제공방법.The biological sample is characterized in that the exosomes derived from blood or plasma, information providing method.
  10. 제7항에 있어서, 8. The method of claim 7,
    상기 mRNA 수준은 차세대 염기서열 분석(Next generation sequencing; NGS), 중합효소연쇄반응(PCR), 역전사 중합효소연쇄반응(RT-PCR), 실시간 중합효소연쇄반응(Real-time PCR), RNase 보호 분석법(RNase protection assay; RPA), 마이크로어레이(microarray), 및 노던 블롯팅(northern blotting)으로 이루어진 군으로부터 선택되는 1종 이상의 방법을 통해 측정되는 것을 특징으로 하는, 정보제공방법.The mRNA level is determined by next generation sequencing (NGS), polymerase chain reaction (PCR), reverse transcription polymerase chain reaction (RT-PCR), real-time polymerase chain reaction (Real-time PCR), RNase protection assay (RNase protection assay; RPA), microarray (microarray), characterized in that measured through one or more methods selected from the group consisting of northern blotting (northern blotting), information providing method.
  11. 제7항에 있어서,8. The method of claim 7,
    상기 단백질 수준은 웨스턴 블롯팅(western blotting), 방사선면역분석법(radioimmunoassay; RIA), 방사 면역 확산법(radioimmunodiffusion), 효소면역분석법(ELISA), 면역침강법(immunoprecipitation), 유세포분석법(flow cytometry), 면역형광염색법(immunofluorescence), 오우크테로니(ouchterlony), 보체 고정 분석법(complement fixation assay), 및 단백질 칩(protein chip)으로 이루어진 군으로부터 선택되는 1종 이상의 방법을 통해 측정되는 것을 특징으로 하는, 정보제공방법.The protein level was determined by western blotting, radioimmunoassay (RIA), radioimmunodiffusion, enzyme immunoassay (ELISA), immunoprecipitation, flow cytometry, and immunity. Information characterized in that it is measured through one or more methods selected from the group consisting of immunofluorescence, ouchterlony, complement fixation assay, and protein chip. How to provide.
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