A cell-permeable inhibitor and activity-based probe for the caspase-like activity of the proteasome

• Published in: Bioorganic & medicinal chemistry letters Vol. 17; no. 12; pp. 3402 - 3405
• Main Authors: van Swieten, Paul F., Samuel, Emlyn, Hernández, Rosa Orient, van den Nieuwendijk, Adrianus M.C.H., Leeuwenburgh, Michiel A., van der Marel, Gijsbert A., Kessler, Benedikt M., Overkleeft, Herman S., Kisselev, Alexei F.
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 15.06.2007; Elsevier
• Subjects: Antigen presentation; Antineoplastic agents; Antineoplastic Agents - chemical synthesis; Antineoplastic Agents - pharmacology; Azide; Azides - chemistry; Binding Sites; Biological and medical sciences; Caspases - chemistry; Caspases - metabolism; Cell Membrane Permeability - drug effects; Cell Membrane Permeability - physiology; Cysteine Proteinase Inhibitors - chemical synthesis; Cysteine Proteinase Inhibitors - pharmacology; General aspects; Humans; Medical sciences; Models, Chemical; Pharmacology. Drug treatments; Protease inhibitor; Proteasome; Proteasome Endopeptidase Complex - chemistry; Proteasome Endopeptidase Complex - immunology; Proteasome Inhibitors; Ubiquitin; Ubiquitin; Antigen presentation; Azide; Protease inhibitor; Proteasome; Multicatalytic endopeptidase complex; Peptides; Sulfone; Cysteine endopeptidases; Active site; Tripeptide; Selectivity; Subunit; Functionalization; Vinylic compound; Chemical synthesis; Serine endopeptidases; Enzyme; Organic azide; Caspase; Enzyme inhibitor; Biological activity; Protein; Peptidases; Vertebrata; Trypsin; Mammalia; Phenols; Hydrolases
• ISSN: 0960-894X; 1464-3405
• DOI: 10.1016/j.bmcl.2007.03.092

A subunit-specific proteasome inhibitor was developed and its specificity was assayed both by fluorescent substrate hydrolysis and by visualization of the targeted subunits via an activity-based probe approach. The ubiquitin–proteasome pathway degrades the majority of proteins in mammalian cells and plays an essential role in the generation of antigenic peptides presented by major histocompatibility class I molecules. Proteasome inhibitors are of great interest as research tools and drug candidates. Most work on proteasome inhibitors has focused on the inhibition of the chymotryptic-like (β5) sites; little attention has been paid to the inhibition of two other types of active sites, the trypsin-like (β2) and the caspase-like (β1). We report here the development of the first cell-permeable and highly selective inhibitors ( 4 and 5) of the proteasome’s caspase-like site. The selectivity of the compounds is directly and unambiguously established by Staudinger–Bertozzi labeling of proteasome subunits covalently modified with azide-functionalized inhibitor 5. This labeling reveals that the caspase-like site of the immunoproteasome (β1i) is a preferred target of this compound. These compounds can be used as tools to study roles of β1 and β1i sites in generation of specific antigenic peptides and their potential role as co-targets of anti-cancer drugs.