The mammalian proteasome is estimated to cleave ~70% of all intracellular proteins and is increasingly recognized as a dynamic complex that modulates cellular function in health and disease. However, the regulatory principles targeting specific substrates to proteasomal degradation and their cleavage products are still poorly understood. Recently, we developed mass spectrometry analysis of proteolytic peptides (MAPP), a method for proteasomal profiling that allows capture, isolation and analysis of proteasome-cleaved peptides. Application of MAPP to a clinical samples of cancer patients reveals novel regulatory principles of proteasomal degradation as well as putative targets for cancer immunotherapy.