This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Successful treatment of breast cancer is hampered by the occurrence of drug resistance in approximately 50% of patients with advanced disease. Currently available markers cannot effectively predict the response to the therapy, and therefore there is a need for new and better biomarkers. The ultimate goal of the proposed work is to unravel functional pathways that lead to clinical resistance to the anti-estrogen tamoxifen by identifying responsible key proteins with their specific modifications, and their interacting partners. These pathways may be targets for newly designed drugs and may lead to individual treatment strategies for patients with (advanced) breast cancer. The responsible key proteins will be identified from laser capture microdissection (LCM) derived tumor samples using nanoLC-FTICR mass spectrometry in conjunction with Accurate Mass and Time (AMT) tag approach and targeted MS/MS strategies. Laser microdissection techniques enable isolation of selected subpopulations of cells, thereby effectively addressing the issue of cell heterogeneity within the tumor. However, the minimal quantity of available sample (<5,000 cells corresponding to only few micrograms of total protein) has seriously limited the ability to perform comprehensive analysis using contemporary proteomics tools. Initial experiments demonstrated that a substantial number of proteins can be detected and identified from minute samples using ultrasensitive LC-FTICR technology and AMT tag approach, thus offering a unique opportunity to study protein profiles within tissue specimens procured by microdissection techniques.
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