NOTCH signaling is a highly conserved short-range signaling pathway involved in virtually every aspect of embryonic development and controls homeostatic self-renewal in many adult tissues. Germ line mutations in the NOTCH pathway cause several hereditary diseases, and somatic mutations are found in cancer. NOTCH pathway activity is governed by ligand induced proteolytic cleavage of the receptor in the extracellular domain by a metalloprotease. This cleavage is required for the consecutive transmembrane cleavage by the γ-secretase complex leading to the release of the intracellular domain that mediates target gene activation. The precise mechanism that confers ligand regulated NOTCH proteolysis is unresolved but involves a ligand-induced conformational change. Virtually nothing is known on how conformational changes render the NOTCH receptor susceptible to proteolysis and which proteases are involved. Human cancer prone NOTCH1 receptor mutations are characterized by increased metalloprotease dependent cleavage and signaling activity. Here I hypothesize that these oncogenic mutations lead to exposure of buried scissile bonds that facilitate proteolysis. Since metalloprotease cleavage of NOTCH receptors is the regulatory event in NOTCH cascade activation, inhibition of this protease activity may find therapeutic application in diseases with deregulated NOTCH signaling such as cancer. Novel technologies and reagents are needed to gain further insight into the NOTCH-protease interface to accelerate drug target discovery and validate their use in pre-clinical models for cancer where deregulated protease activity plays crucial roles in disease progression.