Aberrant proteolysis by cysteine cathepsins is implicated in carcinogenesis, but knowledge of cathepsin substrates mediating tumor-promoting or suppressing effects is limited. Here we characterize tumor proteome and in vivo cathepsin substrates using cathepsin knockout mice and the RIP1-Tag2 model of pancreatic islet carcinogenesis. Applying an unbiased systems-level proteomics approach, Terminal Amine Isotopic Labeling of Substrates (TAILS), we identified cysteine cathepsin B, H, L, S, Z substrates and their cleavage sites. Among 1,935 proteins and 1,114 N-termini identified by TAILS using 8-plex iTRAQ protein labeling, 145 neo-N-termini were significantly changed in one (55%) or more knockouts suggesting a lack of direct compensation at substrate level by other cathepsins. Most affected N-termini (56-83% for different cathepsins) represented degradative cathepsin activity, whereas 17-44% of neo-N termini represented stable proteolytic products in the tumors and were enriched for extracellular proteins. We identified candidate substrates for mediating signaling roles of cysteine cathepsins in tumorigenesis.