Abstract 4573: Discovery of candidate biomarkers for colorectal cancer using mass spectrometry-based proteomic analysis of cell line conditioned media

Abstract

Abstract Colorectal cancer (CRC) represents the third leading cause of cancer-related deaths with approximately 655,000 deaths annually, worldwide. Specific biomarkers for early detection of CRC are needed, mainly because the currently applied serum biomarkers (CEA, CA 19.9) lack adequate sensitivity and specificity for screening purposes, while colonoscopy is invasive, with considerable cost, risks and inconvenience. In the present study, we utilized a mass spectrometry-based platform to analyze the proteome of the conditioned media from 12 CRC cell lines with diverse genetic profiles, to capture the disease heterogeneity. Proteins were trypsin-digested and fractionated by strong cation-exchange and reverse-phase liquid chromatography prior to mass spectrometric analysis with an LTQ Orbitrap instrument. The samples were run in triplicate and results were searched against the IPI human forward and reverse database, using MASCOT and X!Tandem search engines. We were able to identify approximately 2,000 unique proteins for each cell line, with reproducibility ranging from 71 to 82% among the triplicates. The efficiency of this approach was signified by the large number of known biomarkers, such as CEA, successfully identified in our list of candidates, as well as by the identification of internal control proteins, such as kallikreins 6, 7, 10, and 11, which were also quantified by ELISA in cancer cell supernatants. The acquired protein datasets were further subjected to bioinformatic analysis using Gene Ontology. The proteins were assigned a subcellular localization and 35-40% were classified as extracellular or plasma membrane proteins. Further mining of this vast database by using bioinformatics tools and experimental approaches (such as multiple reaction monitoring and ELISA) could lead to shortlists of proteins that may have utility as circulating CRC biomarkers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4573.