Proteomics technologies are revolutionizing the way in which cancer pathways and drug targets can be identified, validated and assessed. To understand the roles of proteases in metastasis and in tumor associated macrophages we applied quantitative proteomics in multiplex analyses to identify the proteome, the N-terminome, and cleaved protease substrates on a global scale. In the 4T1 syngeneic mouse model of breast cancer we analyzed protease expression by our CLIP-CHIPTM, a complete dedicated human and murine protease and inhibitor oligonucleotide microarray. Matrix metalloproteinases (MMPs) 10 and 13 and matriptase-2 were highly expressed. MMP proteases were imaged spatially in 3D and temporally by F18 coupled to Marimastat, a reversible nM MMP inhibitor drug. To elucidate the function of these proteases in vivo by identifying their cleaved substrates we developed a new technique called Terminal Amine Isotope Labeling of Substrates (TAILS) (Nature Biotechnology 28, 281-288, 2010). MS/MS both identifies the substrate and sequence of the cleavage site in the same experiment. We identified numerous novel MMP-10 and 13 substrate candidates including members of the insulin-like growth factor binding protein family in breast cancer secretomes. Proteomic evidence of proteolytic processing of novel substrates was found including Insulin-like growth factor binding protein-4 and 6, follistatin-like 1 and cystatin C, heparin affin regulatory peptide (HARP/pleiotrophin) and connective tissue growth factor (CTGF), which released vascular endothelial growth factor (VEGF) from angiogenic inhibitory complexes. Hence the unmasking of cytokines, such as VEGF, by metalloproteinase processing of their binding proteins is a new mechanism in the control of cytokine activation and angiogenesis. This underscores the potential of TAILS to quantitatively analyze the proteolytic fingerprint of noninvasive 67NR, the metastatic 66cl and the highly malignant 4T1 tumors in vivo in ongoing studies.