Chemically stable diazo peptides: a new class of cellularly active hydrolase inhibitors

Our research has been focused on developing novel methods for fragment-based drug discovery such as protein-templated fragment ligation.(1) The method has been applied for the identification and optimization of protein ligands for viral and human proteases,(2) lactamases,(3) protein tyrosine phosphatases,(4) as well as for protein-protein interactions.(5) The design of and screening for non-cleavable, strongly binding biomimetics has been a successful strategy to obtain specific and potent protein ligands.(6) Diazo compounds have shown selective reactivity toward proteins and nucleic acids and, thus, have been investigated for versatile applications in medicinal chemistry and chemical biology.(7) Here, we will present unpublished work on diazo peptides as a novel class of highly specific, covalent, and irreversible protease inhibitors. Synthesis and chemical stability of this class of molecules has been investigated and then applied to the design of activity-based probes of the human hydrolase caspase 3. Nanomolar inhibitors have been obtained with a remarkable inactivation rate (kinact/KI) of 255,000 M-1s-1 enabling the monitoring of apoptosis in living cells. Current investigations cover the transfer of the inhibitory motif on other hydrolases as protein targets.