TY - JOUR
T1 - SILAC-based quantitative proteomic approach to identify potential biomarkers from the esophageal squamous cell carcinoma secretome
AU - Kashyap, Manoj Kumar
AU - Harsha, H. C.
AU - Renuse, Santosh
AU - Pawar, Harsh
AU - Sahasrabuddhe, Nandini A.
AU - Kim, Min Sik
AU - Marimuthu, Arivusudar
AU - Keerthikumar, Shivakumar
AU - Muthusamy, Babylakshmi
AU - Kandasamy, Kumaran
AU - Subbannayya, Yashwanth
AU - Prasad, Thottethodi Subrahmanya Keshava
AU - Mahmood, Riaz
AU - Chaerkady, Raghothama
AU - Meltzer, Stephen J.
AU - Kumar, Rekha V.
AU - Rustgi, Anil K.
AU - Pandey, Akhilesh
N1 - Funding Information:
We thank the Department of Biotechnology (DBT), Government of India for research support to the Institute of Bioinformatics, Bangalore. M.K.K. is a recipient of an independent Senior Research Fellowship award (IRIS ID# 2006-02010) from the Indian Council of Medical Research (ICMR), New Delhi, India. We thank the Council for Scientific and Industrial Research (CSIR), India for the research support to N.P. and H.P. and the University Grants Commission (UGC), India for the research support to S.R. and Y.S. The work was supported in part by grant CA146799, DK087454 and CA85069 to S.J.M. Also, this work was supported in part by grant NCI P01-CA098101 (Mechanisms of Esophageal Carcinogenesis and its Cell Culture Core) and an American Cancer Society Research Professorship to A.K.R. We thank Agilent Technologies for access to instrumentation.
PY - 2010/10/15
Y1 - 2010/10/15
N2 - 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.
AB - 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.
KW - Het-1A
KW - Mass spectrometry
KW - Metastasis
KW - Multiple reaction monitoring
KW - Prognostication
KW - Tumor differentiation
KW - Tumor grade
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U2 - 10.4161/cbt.10.8.12914
DO - 10.4161/cbt.10.8.12914
M3 - Article
C2 - 20686364
AN - SCOPUS:78049240516
SN - 1538-4047
VL - 10
SP - 796
EP - 810
JO - Cancer Biology and Therapy
JF - Cancer Biology and Therapy
IS - 8
ER -