Naturally Occurring Isothiocyanates Exert Anticancer Effects by Inhibiting Deubiquitinating Enzymes

Abstract

The anticancer properties of cruciferous vegetables are well known and attributed to an abundance of isothiocyanates (ITCs) such as benzyl ITC (BITC) and phenethyl ITC (PEITC). While many potential targets of ITCs have been proposed, a full understanding of the mechanisms underlying their anticancer activity has remained elusive. Here we report that BITC and PEITC effectively inhibit deubiquitinating enzymes (DUBs), including the enzymes USP9x and UCH37, which are associated with tumorigenesis, at physiologically relevant concentrations and time scales. USP9x protects the anti-apoptotic protein Mcl-1 from degradation, and cells dependent on Mcl-1 were especially sensitive to BITC and PEITC. These ITCs increased Mcl-1 ubiquitination and either ITC treatment or RNAi-mediated silencing of USP9x decreased Mcl-1 levels, consistent with the notion that USP9x is a primary target of ITC activity. These ITCs also increased ubiquitination of the oncogenic fusion protein Bcr-Abl, resulting in degradation under low ITC concentrations and aggregation under high ITC concentrations. USP9x inhibition paralleled the decrease in Bcr-Abl levels induced by ITC treatment, and USP9x silencing was sufficient to decrease Bcr-Abl levels, further suggesting that Bcr-Abl is a USP9x substrate. Overall, our findings suggest that USP9x targeting is critical to the mechanism underpinning the well established anticancer activity of ITC. We propose that the ITC-induced inhibition of DUB may also explain how ITCs affect inflammatory and DNA repair processes, thus offering a unifying theme in understanding the function and useful application of ITCs to treat cancer as well as a variety of other pathological conditions.

Development of a Novel Experimental In Vitro Model of Isothiocyanate-induced Apoptosis in Human Malignant Melanoma Cells

“…The majority of studies have utilized non-melanoma (8)(9)(10)(11)(12)(13)(14) and melanoma (15)(16)(17)(18)(19) experimental models based on a single bolus addition of large ITC concentrations where a wide range of biological effects were documented but without provision of appropriate control (non-tumorigenic) cells. Similarly, we have been able to document such anticancer activity in melanoma cells but also in control keratinocyte cells suggesting a non-specific potency towards any type of cell line (tumorigenic or not).…”

Breaking the Fourth Wall: Modulating Quaternary Associations for Protein Regulation and Drug Discovery

“…To stabilize MDM2, a DUB (USP7) associates with MDM2. Knockdown or inhibition of USP7, in cancers manifesting MDM2 overexpression that suppresses wt‐p53, results in MDM2 degradation and upregulation of p53induced apoptosis (Figure ) . Interesting small‐molecule inhibitors of USP7 have been developed .…”

“…Ultimately, BCR‐ABL degradation may be more efficacious than that of kinase inhibition or PPI blocking alone to elicit a “cure”, rather than disease suppression (which is observed with “most” current treatments). Several strategies to degrade BCR‐ABL are in development …”

“…For instance, ribociclib (Scheme ), which is a dual, ATP‐competitive inhibitor of CDK4/CDK6 approved for the treatment of metastatic breast cancer, and a multi‐kinase inhibitor, midostaurin (Scheme ), have also been approved . Similarly, polypharmacological DUB inhibitors are being investigated . Given the diversity and size of PPI interfaces, it is harder to imagine how a PPI intervention strategy could lead to targeting several clients of a scaffolding protein, unless the scaffold itself is blocked.…”

Subcellular Redox Targeting: Bridging in Vitro and in Vivo Chemical Biology

“…There are also dietary plant-based RES of which the most common are isothiocyanates. 25 Isothiocyanates form thiocarbamylated enzyme adducts that initially form cysteine adducts that may transfer to lysine, depending on conditions/protein. 15 There are also well-known synthetic RES including electrophilic drugs, such as dimethyl fumarate (Tecfidera ® ) approved for the treatment of multiple sclerosis, and bardoxolone methyl in Phase 2/3 trials for various conditions.…”