1. Inhibition of the ubiquitin-proteasome system by an NQO1-activatable compound
- Author
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Tatiana A. Giovannucci, Anna-Lena Gustavsson, John Inge Johnsen, Patrick Young, Florian A. Salomons, Jürgen Eirich, Rozbeh Jafari, Nico P. Dantuma, Malin Wickström, Martin Haraldsson, Mikael Altun, Lotta Elfman, and Thomas Lundbäck
- Subjects
Cancer Research ,Programmed cell death ,Proteasome Endopeptidase Complex ,Ubiquitylation ,Proteolysis ,Immunology ,Mice, Nude ,Article ,Substrate Specificity ,Small Molecule Libraries ,Cellular and Molecular Neuroscience ,Ubiquitin ,Cell Line, Tumor ,medicine ,NAD(P)H Dehydrogenase (Quinone) ,Animals ,Humans ,chemistry.chemical_classification ,medicine.diagnostic_test ,biology ,QH573-671 ,Cell Death ,Deubiquitinating Enzymes ,Cell Biology ,Xenograft Model Antitumor Assays ,In vitro ,Recombinant Proteins ,Cell biology ,High-Throughput Screening Assays ,Enzyme ,Phenotype ,chemistry ,Proteasome ,Caspases ,biology.protein ,Screening ,Female ,NAD+ kinase ,Cytology ,Proteasome Inhibitors ,Intracellular - Abstract
Malignant cells display an increased sensitivity towards drugs that reduce the function of the ubiquitin-proteasome system (UPS), which is the primary proteolytic system for destruction of aberrant proteins. Here, we report on the discovery of the bioactivatable compound CBK77, which causes an irreversible collapse of the UPS, accompanied by a general accumulation of ubiquitylated proteins and caspase-dependent cell death. CBK77 caused accumulation of ubiquitin-dependent, but not ubiquitin-independent, reporter substrates of the UPS, suggesting a selective effect on ubiquitin-dependent proteolysis. In a genome-wide CRISPR interference screen, we identified the redox enzyme NAD(P)H:quinone oxidoreductase 1 (NQO1) as a critical mediator of CBK77 activity, and further demonstrated its role as the compound bioactivator. Through affinity-based proteomics, we found that CBK77 covalently interacts with ubiquitin. In vitro experiments showed that CBK77-treated ubiquitin conjugates were less susceptible to disassembly by deubiquitylating enzymes. In vivo efficacy of CBK77 was validated by reduced growth of NQO1-proficient human adenocarcinoma cells in nude mice treated with CBK77. This first-in-class NQO1-activatable UPS inhibitor suggests that it may be possible to exploit the intracellular environment in malignant cells for leveraging the impact of compounds that impair the UPS.
- Published
- 2021