Proteomic profiling of breast cancer serum via two-dimensional liquid-chromatography—tandem mass spectrometry

Abstract

542 Background: Discovering biomarkers for early cancer detection is a major goal of translational research. We have applied modern proteomic technology, two-dimensional liquid-chromatography-tandem mass spectrometry (2-D LC-MS/MS), to large-scale breast cancer serum profiling. Methods: 189 human sera from breast disease or cancer patients (14 atypical hyperplasia, 63 benign, 29 DCIS, and 83 invasive CA) were analyzed via 2-D LC-MS/MS method. Patient ages ranged from 19 to 97 years, with a mean of 55. Ethnicities included African American, Asian, Hispanic, Native American, and Caucasian. This methodology utilizes only 10 μL of human serum per specimen. Every sample was run three times to ensure reproducibility and to maximize identification. The protein identification was conducted using the Bioworks Browser 3.1 and the IPI_human protein database (version 6.23.04) and filtered via HUPO criteria to ensure confidence. Results: 230 high-confidence proteins were identified from analyses of 189 sera. 26 common serum proteins were detected in all samples and 17 were found in two- or three- diagnostic stages. 83 proteins such as insulin receptor substrate 4, serine/threonine-protein kinase WNK4, DNA mismatch repair protein MSH6, and signal recognition particle 72 kDa protein were detected only in invasive sera. 30 proteins including breast carcinoma amplified sequence 3, uncharacterized bone marrow protein BM032, fibroblast growth factor receptor 2, and tumor necrosis factor receptor superfamily member 4 were detected only in DCIS sera. The 15 proteins detected only in atypical hyperplasia sera included cholecystokinin type A receptor and cell division cycle 2-related protein kinase 7. 59 proteins including serine/threonine protein phosphatase 6, probable transcription factor CST, insulin-like growth factor IA, and connective tissue growth factor were found in benign samples. Conclusions: Identification of signaling pathway/transcriptions proteins and cell cycle related proteins in breast cancer sera, and identification of breast cancer amplified sequence 3 proteins in the DCIS sera, suggest that high-throughput proteomic technologies like 2-D LC - MS/MS may facilitate the discovery of diagnostically useful breast cancer biomarkers. No significant financial relationships to disclose.