Identification and Validation of Ovarian Cancer-Associated Proteins

Abstract

The proteomic analysis of plasma and tissues in patients has been a major approach to determining biomarkers essential for early disease diagnoses and drug discoveries. Recently, we detected and identified 40 proteins associated wirh ovarian clear cell carcinoma by two-dimensional difference gel electrophoresis and tandem mass spectrometry (MS/MS), and also isobaric tag for relative and quantitative analysis (iTRAQ) and MS/MS. Among the identified proteins, the expression of several proteins such as annexin IV was validated. In most cases, the expression is regulated at transcriptional level in the cancer cells. We found that the annexin IV gene has a transcription regulatory region containing a similar sequence to the NF-κB binding motif. When the expression of the genes encoding these proteins was suppressed with the siRNAs, the proliferation of the cancer cells was inhibited at different levels depending on the protein. On the other hand, we enriched proteins of which phosphorylation is stimulated in the ovarian cancer cells by immunoaffinity chromatography, and identified several proteins including Stat3 by iTRAQ and MS/MS. Finally, we investigated if we can detect the ovarian cancer-associated proteins in the plasma. The detection of the plasma proteins, however, is analytically challenging because the dynamic concentration range of them is extremely wide. We established a novel technique to analyze plasma proteins. In this technique, an originally developed gabundant protein depletion deviceh and a sequentially linked three-dimensional liquid chromatography-MS/MS (3D-LC-MS/MS) system were used. By this technique, we can identify nearly 3,000 low abundant proteins in the plasma. However, among the ovarian cancer-associated proteins identified in the cancer tissues and cultured cells, we detected only annexin IV in the S2/July 2008/Special Issue