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.
Highlights
Proteotoxic stress is a condition that is observed in many cancer types and can be capitalized in anticancer regimens [1]
Cell-based, high-content screen for small molecule inhibitors of the ubiquitin-proteasome system (UPS) With the aim of finding novel UPS inhibitors, we performed a phenotypic screen for small molecules that cause a general accumulation of a UPS reporter substrate (Fig. 1A; Supplementary Table S1)
In line with a general inhibition of ubiquitin-dependent degrada- We found a clear enrichment of guides targeting the oxidortion, CBK77 treatment increased the levels of Hsp70, which is a eductase NAD(P)H:quinone oxidoreductase 1 (NQO1) in the CBK77-treated population (Fig. 5C, common stress response elicited by inhibition of protein Supplementary Table S3)
Summary
Proteotoxic stress is a condition that is observed in many cancer types and can be capitalized in anticancer regimens [1]. The underlying causes of proteotoxic stress in malignant cells are multifactorial and inherently linked to their hyperactive state, such as an overall increase in protein synthesis and accumulation of aberrant and orphan proteins due to the loss of genome integrity. The ubiquitin-proteasome system (UPS) is the primary proteolytic system in charge of the timely and efficient degradation of aberrant and excessive proteins that are at risk of polluting the intracellular milieu [2]. The increased demand for clearance of misfolded proteins generates a therapeutic window at which reduction of UPS activity is lethal for cancer cells without severely affecting protein homeostasis in healthy cells [4]. Despite the overall success of proteasome inhibitors in the treatment of hematological malignancies, several remaining challenges, such as adverse side effects, drug resistance, and poor activity towards solid tumors, motivate the exploration of alternative approaches for inhibiting UPS activity
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
