USP32 confers cancer cell resistance to YM155 via promoting ER-associated degradation of solute carrier protein SLC35F2

Abstract

<b>Background:</b> The most commonly preferred chemotherapeutic agents to treat cancers are small-molecule drugs. However, the differential sensitivity of various cancer cells to small molecules and untargeted delivery narrow the range of potential therapeutic applications. The mechanisms responsible for drug resistance in a variety of cancer cells are also largely unknown. Several deubiquitinating enzymes (DUBs) are the main determinants of drug resistance in cancer cells. <b>Methods:</b> We used CRISPR-Cas9 to perform genome-scale knockout of the entire set of genes encoding ubiquitin-specific proteases (USPs) and systematically screened for DUBs resistant to the clinically evaluated anticancer compound YM155. A series of <i>in vitro</i> and <i>in vivo</i> experiments were conducted to reveal the relationship between USP32 and SLC35F2 on YM155-mediated DNA damage in cancer cells. <b>Results:</b> CRISPR-based dual-screening method identified USP32 as a novel DUB that governs resistance for uptake of YM155 by destabilizing protein levels of SLC35F2, a solute-carrier protein essential for the uptake of YM155. The expression of USP32 and SLC35F2 was negatively correlated across a panel of tested cancer cell lines. YM155-resistant cancer cells in particular exhibited elevated expression of USP32 and low expression of SLC35F2. <b>Conclusion:</b> Collectively, our DUB-screening strategy revealed a resistance mechanism governed by USP32 associated with YM155 resistance in breast cancers, one that presents an attractive molecular target for anti-cancer therapies. Targeted genome knockout verified that USP32 is the main determinant of SLC35F2 protein stability <i>in vitro</i> and <i>in vivo</i>, suggesting a novel way to treat tumors resistant to small-molecule drugs.