Project Summary The Proteomics and Metabolomics Shared Resource (PMSR) provides a comprehensive set of proteomics and metabolomics assays to the Wistar Cancer Center, Fox Chase Cancer Center at Temple University, and Sidney Kimmel Cancer Center at Thomas Jefferson University as a primary goal. Resources of the PMSR are also available to investigators in other Cancer Centers and academic institutions as a secondary goal. Dr. Tang’s role as the PMSR Managing Director is to support NCI-funded cancer research projects by providing expert consultation and state-of-the-art technologies that operate at maximum performance at affordable costs to cancer investigators and other biomedical researchers. The Managing Director will assist in experimental design, perform MS data analyses as needed, and assist in the biological interpretation of results. The Managing Director will also devote substantial effort to optimizing and implementing new methods, update analytical and data analyses methods, and update instrumentation to ensure each project is performed using state-of-the-art methodologies. This is critical because instrumentation, software, and analytical strategies continue to evolve rapidly, and most current and anticipated future projects involve very challenging proteomics and metabolomics problems. Proteomics projects will include: 1) in-depth, global quantitative comparisons of exosomes, secretomes, cell lysates, tissues and biological fluids; 2) quantitative comparisons of post-translational modifications; and 3) LC-MS/MS analysis of isolated protein complexes with and without chemical crosslinking. Quantitative data will be obtained either using label-free quantitation of integrated MS ion currents, or using stable isotopes such as SILAC or TMT isobaric tag labeling. Metabolomics projects will include quantifying the steady state levels of polar metabolites, lipids and fatty acids, and 13C isotope tracer analysis. Representative specific plans for future development of new analytical approaches include: 1) improving characterization of protein palmitoylation with identification of specific modification sites; 2) de novo peptide sequencing for identification of HLA peptides; 3) an alternate approach to ubiquitome enrichment; and 4) absolute quantitation of gut metabolites. In addition, Dr. Tang will implement additional new or improved methods that are likely to be needed in future cancer projects. These proteomics and metabolomics analyses will contribute to critical data required to identify biomolecular targets, as well as generate hypotheses that are vital for the success of the cancer-related projects described in this application.

Project Narrative The application is aimed to support NCI-funded cancer research projects from investigators at the Wistar Institute Cancer Center, the Fox Chase Cancer Center at Temple University, the Sidney Kimmel Cancer Center at Thomas Jefferson University, and other academic institutions using proteomics and metabolomics technologies offered by the Wistar Proteomics and Metabolomics Shared Resource. Supported research projects are diverse with focus on many different types of cancer, such as prostate, ovarian, and melanoma, and experimental samples include immortalized or primary cells, biological fluids, and tissues. The analytical assays will provide insight into perturbed biological pathways and identify key biomolecules associated with disease states, with the final goal of developing methods for cancer prevention, diagnosis and treatment.