Abstract The overarching goal of the Proteomics Core (Core 3) is to provide state-of-the-art, mass spectrometry-based- proteomics and phosphoproteomics data and analyses in support of Projects 1-3. Genetic alterations in human cancer have been systematically mapped by genomics landscape studies in the past decade, however, the direct consequences of these alterations on the functional proteome are poorly understood. Deep scale, mass spectrometry (MS)-based proteomic data when integrated with genomic data (`proteogenomics') have been shown to improve specificity for identifying cancer-relevant pathways triggered by somatic DNA variants or DNA copy number alterations (CNAs) compared to genomic characterization alone, and help to narrow target selection for potential therapeutic intervention. Proteomics alone, especially with deep, quantitative profiling of posttranslational modifications (PTM) provides information on signaling related to disease pathophysiology that are largely opaque to genomics. Core 3 will apply micro-scaled mass spectrometry-based proteomics technologies we have developed that utilize highly multiplexed stable-isotope mass tagging (TMT 16-plex) for precise relative quantification of the proteome and phosphoproteome of very small amounts with very deep coverage for the study of transformation of chronic lymphocytic leukemia (CLL) to Richter's Syndrome (RS). The resulting proteomic data, including the critical quantitative and site-specific modification information, will be integrated with personalized genomic data using bioinformatics tools that have been integrated into the cloud-based pipeline PANOPLY. Multi-omics clustering and analysis will be done to define the intrinsic structure of the integrated proteogenomes across baseline and treated samples. We will extract proteogenomic features that drive the underlying cluster structure and will perform pathway-level analysis to further characterize each cluster in CLL and RS samples. Copy number to mRNA, protein, and phosphoprotein correlations will be done to determine cis- and trans-regulated genes. Pathways and molecular mechanisms underlying treatment response in patient and mouse models will be explored using single sample Gene Set Enrichment Analysis (ssGSEA), and PTM Signature Enrichment Analysis (PTM-SEA) will be used to perform pathway analysis on phosphorylation data generated. To enable more rapid and specific analyses of proteins and phosphopeptides targets of interest emerging from the discovery experiments, the proteomics core will develop high sensitivity targeted MS assays, to be utilized in Projects 1-3. Assays developed will use stable isotope-labeled standards for unambiguous identification and quantification and applied to human biospecimens and preclinical samples in native and drug-perturbed states.