Quantitative Proteomic Analysis to Discover Potential Diagnostic Markers and Therapeutic Targets in Human Renal Cell Carcinoma

Abstract

Renal cell carcinoma (RCC) is relatively resistant to chemotherapy and radiotherapy. Recent advances in drug development are providing novel agents for the treatment of RCC, but the effects are still minimal. In addition, there is an urgent need to identify diagnostic markers for RCC. In this report, to discover potential diagnostic markers and therapeutic targets, we subjected RCC samples to a quantitative proteomic analysis utilizing 2-nitrobenzenesulfenyl (NBS) reagent. Proteins were extracted from RCC and adjacent normal tissue, obtained surgically from patients, and labeled with NBS reagent containing six 12C or 13C. This was followed by trypsin digestion and the enrichment of labeled peptides. Samples were then subjected to analysis by MALDI-TOF MS. Peaks with altered signal intensities (> 2.0-fold) and that occurred frequently in samples (> 60%) were selected, and 92 proteins were identified by MS/MS analyses as differentially expressed proteins in RCC tissues. Thirty-four proteins were up-regulated in RCC samples of which some were previously known, and some were novel. The up-regulation of two proteins (Galectin-1, CNDP2) and their mRNA expression in RCC was confirmed by Western blotting and quantitative real time RT-PCR. The results suggest that NBS-based quantitative proteomic analysis is useful for discovering diagnostic markers and therapeutic targets for RCC.