Deubiquitinase Activity Profiling Identifies UCHL1 as a Candidate Oncoprotein That Promotes TGFβ-Induced Breast Cancer Metastasis

Abstract

Purpose: Therapies directed to specific molecular targets are still unmet for patients with triple-negative breast cancer (TNBC). Deubiquitinases (DUB) are emerging drug targets. The identification of highly active DUBs in TNBC may lead to novel therapies. Experimental Design: Using DUB activity probes, we profiled global DUB activities in 52 breast cancer cell lines and 52 patients' tumor tissues. To validate our findings in vivo, we employed both zebrafish and murine breast cancer xenograft models. Cellular and molecular mechanisms were elucidated using in vivo and in vitro biochemical methods. A specific inhibitor was synthesized, and its biochemical and biological functions were assessed in a range of assays. Finally, we used patient sera samples to investigate clinical correlations. Results: Two DUB activity profiling approaches identified UCHL1 as being highly active in TNBC cell lines and aggressive tumors. Functionally, UCHL1 promoted metastasis in zebrafish and murine breast cancer xenograft models. Mechanistically, UCHL1 facilitates TGFb signaling-induced metastasis by protecting TGFb type I receptor and SMAD2 from ubiquitination. We found that these responses are potently suppressed by the specific UCHL1 inhibitor, 6RK73. Furthermore, UCHL1 levels were significantly increased in sera of patients with TNBC, and highly enriched in sera exosomes as well as TNBC cell-conditioned media. UCHL1enriched exosomes stimulated breast cancer migration and extravasation, suggesting that UCHL1 may act in a paracrine manner to promote tumor progression. Conclusions: Our DUB activity profiling identified UCHL1 as a candidate oncoprotein that promotes TGFb-induced breast cancer metastasis and may provide a potential target for TNBC treatment.

Discovery of a Potent and Selective Covalent Inhibitor and Activity-Based Probe for the Deubiquitylating Enzyme UCHL1, with Antifibrotic Activity

“…Finally, we show that selective UCHL1 inhibitors can suppress fibrotic phenotypes in IPF cellular models without substantial cytotoxicity. While the precise role of UCHL1 in fibrosis remains to be determined, UCHL1 was recently identified as a potential target in triple-negative breast cancer (TNBC), where it promotes TGF-β1 signaling 8 and suppresses estrogen receptor expression. 30 1 and 2 represent powerful and selective probes to explore UCHL1 activity with potential application to substrate identification, mode of action studies, and cellular target profiling, which can accelerate future development of UCHL1 inhibitors as potential therapeutics.…”

Circulating miR-1246 Targeting UBE2C, TNNI3, TRAIP, UCHL1 Genes and Key Pathways as a Potential Biomarker for Lung Adenocarcinoma: Integrated Biological Network Analysis

“…Jin et al [50] reported that the miR-548e-5p, together with UBE-2C and zinc finger E-box binding homeobox (ZEB1/2), acts as a potential diagnostic biomarker and target for NSCLC. Moreover, another ubiquitin-protein UCHL1 was reported to promote uterine serous cancer cell proliferation, cell cycle progression [51], and TGFβ-induced breast cancer metastasis [52]. So far, no study has mentioned the relationship between miR-1246 and UBE-2C and UCHL1 in LUAD.…”

A Small-Molecule Activity-Based Probe for Monitoring Ubiquitin C-terminal Hydrolase L1 (UCHL1) Activity in Live Cells and Zebrafish Embryos

“…In fact, we recently showed that 6RK73 decreases UCHL1 activity and thereby inhibits TGFβ/SMAD2 and SMAD3 signaling and breast cancer migration and extravasation. 48 We are convinced that the here reported strategy of small-molecule cyanimide-based probes can be expanded to other cysteine proteases and specifically DUBs. With the rising importance of the Ub system as source of practical drug targets we believe that these ABP tools will fill an unmet need allowing us to study active DUBs in their native environment in live cells or animals and as such aid in the development of future therapeutics that target diseases associated with ubiquitination.…”