USP28 and USP25 are downregulated by Vismodegib in vitro and in colorectal cancer cell lines

Abstract

Deubiquitinase USP28 plays a crucial role in tumorigenesis by enhancing the stabilities of multiple cancer‐related proteins including c‐Myc, Notch1, and LSD1, and has become an attractive target for anticancer drug development. However, to date, only a few of USP28‐targeted active compounds have been developed, and the active compound‐binding pocket in USP28 has not been experimentally revealed yet. In this study, bioassay‐based high‐throughput screening was applied to discover USP28‐targeted inhibitors from the commercially available drug library. Vismodegib, an inhibitor of Hedgehog signaling pathway and FDA‐approved drug for the treatment of basal cell carcinoma, was found to exhibit inhibition activity against USP28 (IC50: 4.41 ± 1.08 μm). Multiple biophysical and biochemical techniques including NMR, ITC, thermal shift assay, HDX‐MS, and site‐directed mutagenesis analysis were then used to characterize the interaction between Vismodegib and USP28. The binding pocket in USP28 for Vismodegib, which is mainly composed of two helical structures spanning D255‐N278 and N286‐Y293, was revealed. According to the possible binding pose generated by HDX‐MS data‐defined molecular docking, the binding cavity occupied by Vismodegib in USP28 aligns well with one of the reported‐binding pockets in USP7 for its inhibitors. Furthermore, cellular assays were conducted to confirm that Vismodegib could interact with the evolutionarily related deubiquitinases USP28 and USP25 and downregulate the levels of the two enzymes' substrate proteins c‐Myc, Notch1, and Tankyrase‐1/2.

Targeting the MYC Ubiquitination-Proteasome Degradation Pathway for Cancer Therapy

“…Inhibitors targeting the MYC degradation pathway.Vismodegib, a sonic hedgehog inhibitor used for the treatment of basal cell carcinoma, was recently shown to bind to USP28 and inhibit its DUB activity(101). Vismodegib exhibits selectivity towards USP28 and its evolutionally related USP25.…”

“…Vismodegib exhibits selectivity towards USP28 and its evolutionally related USP25. Treatment of cancer cells with Vismodegib reduces the levels of c-Myc and Notch1 and suppresses cell growth(101). In addition, lanatoside C, a cardiac glycoside, has been shown to reduce MYC levels and suppress gastric cancer cell proliferation by inhibiting USP28 binding to MYC, thereby destabilizing MYC, although it remains to be determined whether lanatoside C directly targets USP28(102).Given that SENP1 positively regulates MYC levels and activity, SENP1 is an interesting cancer therapeutic target.…”

Antiproliferative Activity and Potential Mechanism of Marine-Sourced Streptoglutarimide H against Lung Cancer Cells

“…Benzylaminoethanols (AZ1–AZ4) were the first reported compounds to directly and effectively target USP28 [ 29 ]. More recently, a few synthesized USP28 inhibitors were also reported [ 30 , 31 ]. To the best of our knowledge, streptoglutarimide H is the first USP28 inhibitor found from natural resource to display potent antiproliferative activity against lung cancer cells with unique mechanism of action.…”

Emerging Roles of Ubiquitin-Specific Protease 25 in Diseases

“…Wang et al found that this inhibition suppressed the expression levels of its related substrate proteins, including c-Myc, Notch1 and tankyrase-1/2 in vitro and in colorectal cell lines. The binding pocket for vismodegib in USP25 is within two helices that span Asp262 to Glu285 and Asn293 to Tyr300 ( Wang et al, 2020a ).…”