Article

SILAC-based quantitative proteomic analysis of gastric cancer secretome

Institute of Bioinformatics, International Technology Park, Bangalore, 560066, India.
PROTEOMICS - CLINICAL APPLICATIONS (Impact Factor: 2.68). 06/2013; 7(5-6). DOI: 10.1002/prca.201200069
Source: PubMed

ABSTRACT PURPOSE: Gastric cancer is a commonly occurring cancer in Asia and one of the leading causes of cancer deaths. However, there is no reliable blood-based screening test for this cancer. Identifying proteins secreted from tumor cells could lead to the discovery of clinically useful biomarkers for early detection of gastric cancer. EXPERIMENTAL DESIGN: A SILAC-based quantitative proteomic approach was employed to identify secreted proteins that were differentially expressed between neoplastic and non-neoplastic gastric epithelial cells. Proteins from the secretome were subjected to SDS-PAGE and SCX-based fractionation, followed by mass spectrometric analysis on an LTQ-Orbitrap Velos mass spectrometer. Immunohistochemical labeling was employed to validate a subset of candidates using tissue microarrays. RESULTS: We identified 2,205 proteins in the gastric cancer secretome of which 263 proteins were overexpressed >4-fold in gastric cancer-derived cell lines as compared to non-neoplastic gastric epithelial cells. Three candidate proteins, proprotein convertase subtilisin/kexin type 9 (PCSK9), lectin mannose binding 2 (LMAN2) and PDGFA associated protein 1 (PDAP1), were validated by immunohistochemical labeling. CONCLUSIONS AND CLINICAL RELEVANCE: We report here the largest cancer secretome described to date. The novel biomarkers identified in the current study are excellent candidates for further testing as early detection biomarkers for gastric adenocarcinoma.

2 Followers
 · 
199 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: Owing to recent advances in proteomics analytical methods coupled with bioinformatics capabilities there is a growing trend towards using these capabilities for the development of drugs to treat human disease, including target and drug evaluation, understanding mechanisms of action, and biomarker discovery. Currently the genetic sequences of many major organisms are available, which has helped greatly in characterizing proteomes in model animal systems and in humans. Through proteomics, global profiles of different disease states can be characterized (e.g. changes in types and relative levels as well as post-translational modifications such as glycosylation or phosphorylation). Although intracellular proteomics can provide a broad overview of physiology of cells and tissues, it has been difficult to quantify the low abundance proteins which can be important for understanding the diseased states and treatment progression. For this reason, there is increasing interest in coupling comparative proteomics methods with subcellular fractionation and enrichment techniques for membranes, nucleus, phosphoproteome, glycoproteome as well as low abundance serum proteins. In this review, we will provide examples of where the utilization of different proteomics-coupled enrichment techniques has aided target and biomarker discovery, understanding the targeting mechanism, and mAb discovery. Taken together, these improvements will help to provide a better understanding of the pathophysiology of various diseases including cancer, autoimmunity, inflammation, cardiovascular, and neurological conditions, and in the design and development of better medicines for treating these afflictions. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    PROTEOMICS - CLINICAL APPLICATIONS 02/2015; 9(1-2). DOI:10.1002/prca.201400097 · 2.68 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Amino acids play essential roles in both metabolism and the proteome. Many studies have profiled free amino acids (FAAs) or proteins; however, few have connected the measurement of FAA with individual amino acids in the proteome. In this study, we developed a metabolomics method to comprehensively analyze amino acids in different domains, using two examples of different sample types and disease models. We first examined the responses of FAAs and insoluble-proteome amino acids (IPAAs) to the Myc oncogene in Tet21N human neuroblastoma cells. The metabolic and proteomic amino acid profiles were quite different, even under the same Myc-induced transfection, and their combination provided a better understanding of the biological status. In addition, amino acids were measured in 3 domains (FAAs, free and soluble-proteome amino acids (FSPAAs), and IPAAs) to study changes in serum amino acid profiles related to colon cancer. A penalized logistic regression model based on the amino acids from the three domains had better sensitivity and specificity than that from each individual domain. To the best of our knowledge, this is the first study to perform a combined analysis of amino acids in different domains, and indicates the useful biological information available from a metabolomics analysis of the protein pellet. This study lays the foundation for further quantitative tracking of the distribution of amino acids in different domains, with opportunities for better diagnosis and mechanistic studies of various diseases.
    The Analyst 02/2015; 140(8). DOI:10.1039/C4AN02386B · 3.91 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Obesity is a major public health threat for many industrialised countries. Bariatric surgery is the most effective treatment against obesity, suggesting that gut derived signals are crucial for energy balance regulation. Several descriptive studies have proven the presence of gastric endogenous systems that modulate energy homeostasis; however, these systems and the interactions between them are still not well known. In the present study, we show for the first time the comparative 2-DE gastric secretome analysis under different nutritional status. We have identified 38 differently secreted proteins by comparing stomach secretomes from tissue explant cultures of rats under feeding, fasting and re-feeding conditions. Among the proteins identified, glyceraldehyde-3-phosphate dehydrogenase was found to be more abundant in gastric secretome and plasma after re-feeding, and downregulated in obesity. Additionally, two calponin-1 species were decreased in feeding state, and other were modulated by nutritional and metabolic conditions. These and other secreted proteins identified in this work may be considered as potential gastrokines implicated in food intake regulation.
    Journal of Proteomics 01/2015; DOI:10.1016/j.jprot.2015.01.001 · 3.93 Impact Factor