title: SILAC-based quantitative proteomic approach to identify potential biomarkers from the esophageal squamous cell carcinoma secretome.
abstract: The identification of secreted proteins that are differentially expressed between non-neoplastic and esophageal squamous cell carcinoma (ESCC) cells can provide potential biomarkers of ESCC. We used a SILAC-based quantitative proteomic approach to compare the secretome of ESCC cells with that of non-neoplastic esophageal squamous epithelial cells. Proteins were resolved by SDS-PAGE, and tandem mass spectrometry analysis (LC-MS/MS) of in-gel trypsin-digested peptides was carried out on a high-accuracy qTOF mass spectrometer. In total, we identified 441 proteins in the combined secretomes, including 120 proteins with > 2-fold upregulation in the ESCC secretome vs. that of non-neoplastic esophageal squamous epithelial cells. In this study, several potential protein biomarkers previously known to be increased in ESCC including matrix metalloproteinase 1, transferrin receptor, and transforming growth factor beta-induced 68 kDa were identified as overexpressed in the ESCC-derived secretome. In addition, we identified several novel proteins that have not been previously reported to be associated with ESCC. Among the novel candidate proteins identified, protein disulfide isomerase family a member 3 (PDIA3), GDP dissociation inhibitor 2 (GDI2), and lectin galactoside binding soluble 3 binding protein (LGALS3BP) were further validated by immunoblot analysis and immunohistochemical labeling using tissue microarrays. This tissue microarray analysis showed overexpression of protein disulfide isomerase family a member 3, GDP dissociation inhibitor 2, and lectin galactoside binding soluble 3 binding protein in 93%, 93% and 87% of 137 ESCC cases, respectively. Hence, we conclude that these potential biomarkers are excellent candidates for further evaluation to test their role and efficacy in the early detection of ESCC.
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore, India.
authors.forename: Manoj Kumar
authors.initials: MK
authors.lastname: Kashyap
authors.forename: H C
authors.initials: HC
authors.lastname: Harsha
authors.forename: Santosh
authors.initials: S
authors.lastname: Renuse
authors.forename: Harsh
authors.initials: H
authors.lastname: Pawar
authors.forename: Nandini A
authors.initials: NA
authors.lastname: Sahasrabuddhe
authors.forename: Min-Sik
authors.initials: MS
authors.lastname: Kim
authors.forename: Arivusudar
authors.initials: A
authors.lastname: Marimuthu
authors.forename: Shivakumar
authors.initials: S
authors.lastname: Keerthikumar
authors.forename: Babylakshmi
authors.initials: B
authors.lastname: Muthusamy
authors.forename: Kumaran
authors.initials: K
authors.lastname: Kandasamy
authors.forename: Yashwanth
authors.initials: Y
authors.lastname: Subbannayya
authors.forename: Thottethodi Subrahmanya Keshava
authors.initials: TS
authors.lastname: Prasad
authors.forename: Riaz
authors.initials: R
authors.lastname: Mahmood
authors.forename: Raghothama
authors.initials: R
authors.lastname: Chaerkady
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Meltzer
authors.forename: Rekha V
authors.initials: RV
authors.lastname: Kumar
authors.forename: Anil K
authors.initials: AK
authors.lastname: Rustgi
authors.forename: Akhilesh
authors.initials: A
authors.lastname: Pandey
mesh_terms: D000328:Adult; D000368:Aged; D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000032:analysis* / Q000378:metabolism; D015153:Blotting, Western; D002247:Carbon Isotopes / Q000737:chemistry; D002294:Carcinoma, Squamous Cell / Q000378:metabolism* / Q000473:pathology; D002460:Cell Line; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004938:Esophageal Neoplasms / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D007150:Immunohistochemistry; D007553:Isotope Labeling / Q000379:methods; D013058:Mass Spectrometry / Q000379:methods; D008875:Middle Aged; D009587:Nitrogen Isotopes / Q000737:chemistry; D020543:Proteome / Q000032:analysis* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D046888:Tissue Array Analysis
chemical_list: D000596:Amino Acids; D014408:Biomarkers, Tumor; D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D020543:Proteome