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Chem Sci. 2019 Jul 31;10(36):8461-8477. doi: 10.1039/c9sc00997c. eCollection 2019 Sep 28.

Fluorescent probes towards selective cathepsin B detection and visualization in cancer cells and patient samples.

Author information

1
Sanford Burnham Prebys Medical Discovery Institute , 10901 North Torrey Pines Road , La Jolla , CA 92037 , USA . Email: marcin.poreba@pwr.edu.pl ; Email: marcin.drag@pwr.edu.pl ; Email: gsalvesen@sbpdiscovery.org.
2
Department of Bioorganic Chemistry , Faculty of Chemistry , Wroclaw University of Technology , Wyb. Wyspianskiego 27 , 50-370 Wroclaw , Poland.
3
Department of Biochemistry and Molecular and Structural Biology , Jožef Stefan Institute , SI-1000 Ljubljana , Slovenia.
4
Faculty of Chemistry and Chemical Technology , University of Ljubljana , SI-1000 Ljubljana , Slovenia.

Abstract

Human cysteine cathepsins constitute an 11-membered family of proteases responsible for degradation of proteins in cellular endosomal-lysosomal compartments as such, they play important roles in antigen processing, cellular stress signaling, autophagy, and senescence. Moreover, for many years these enzymes were also linked to tumor growth, invasion, angiogenesis and metastasis when upregulated. Individual biological roles of each cathepsin are difficult to establish, because of their redundancy and similar substrate specificities. Selective chemical tools that enable imaging of individual cathepsin activities in living cells, tumors, and the tumor microenvironment may provide a better insight into their functions. In this work, we used HyCoSuL technology to profile the substrate specificity of human cathepsin B. The use of unnatural amino acids in the substrate library enabled us to uncover the broad cathepsin B preferences that we utilized to design highly-selective substrates and fluorescent activity-based probes (ABPs). We further demonstrated that Cy5-labeled MP-CB-2 probe can selectively label cathepsin B in eighteen cancer cell lines tested, making this ABP highly suitable for other biological setups. Moreover, using Cy5-labelled MP-CB-2 we were able to demonstrate by fluorescence microscopy that in cancer cells cathepsins B and L share overlapping, but not identical subcellular localization.

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