An Activity-Based Probe for Cathepsin K Imaging with Excellent Potency and Selectivity

• Published in: Journal of medicinal chemistry 2021-09, Vol.64 (18), p.13793-13806
• Main Authors: Lemke, Carina, Benýšek, Jakub, Brajtenbach, Dominik, Breuer, Christian, Jílková, Adéla, Horn, Martin, Buša, Michal, Ulrychová, Lenka, Illies, Annika, Kubatzky, Katharina F, Bartz, Ulrike, Mareš, Michael, Gütschow, Michael
• Format: Article
• Language: English
• Subjects: Acrylamides - chemical synthesis; Acrylamides - chemistry; Acrylamides - metabolism; Catalytic Domain; Cathepsin K - antagonists & inhibitors; Cathepsin K - chemistry; Cathepsin K - metabolism; Cell Line, Tumor; Cysteine Proteinase Inhibitors - chemical synthesis; Cysteine Proteinase Inhibitors - chemistry; Cysteine Proteinase Inhibitors - metabolism; Drug Design; Fluorescent Dyes - chemical synthesis; Fluorescent Dyes - chemistry; Fluorescent Dyes - metabolism; Humans; Microscopy, Confocal; Microscopy, Fluorescence; Protein Binding
• ISSN: 0022-2623; 1520-4804
• DOI: 10.1021/acs.jmedchem.1c01178

The cysteine protease cathepsin K is a target for the treatment of diseases associated with high bone turnover. Cathepsin K is mainly expressed in osteoclasts and responsible for the destruction of the proteinaceous components of the bone matrix. We designed various fluorescent activity-based probes (ABPs) and their precursors that bind to and inactivate cathepsin K. ABP 25 exhibited extraordinary potency (k inac/K i = 35,300 M–1s–1) and selectivity for human cathepsin K. Crystal structures of cathepsin K in complex with ABP 25 and its nonfluorescent precursor 21 were determined to characterize the binding mode of this new type of acrylamide-based Michael acceptor with the particular orientation of the dibenzylamine moiety to the primed subsite region. The cyanine-5 containing probe 25 allowed for sensitive detection of cathepsin K, selective visualization in complex proteomes, and live cell imaging of a human osteosarcoma cell line, underlining its applicability in a pathophysiological environment.