CRISPR/Cas9 screens unravel miR-3689a-3p regulating sorafenib resistance in hepatocellular carcinoma via suppressing CCS/SOD1-dependent mitochondrial oxidative stress

Abstract

<h3>Aims</h3><p>Therapeutic outcome of <a href="https://www.sciencedirect.com/topics/medicine-and-dentistry/sorafenib" title="Learn more about sorafenib from ScienceDirect's AI-generated Topic Pages">sorafenib</a> in <a href="https://www.sciencedirect.com/topics/pharmacology-toxicology-and-pharmaceutical-science/liver-cell-carcinoma" title="Learn more about hepatocellular carcinoma from ScienceDirect's AI-generated Topic Pages">hepatocellular carcinoma</a> (HCC) is undermined by the development of drug resistance. This study aimed to identify the critical <a href="https://www.sciencedirect.com/topics/medicine-and-dentistry/microrna" title="Learn more about microRNA from ScienceDirect's AI-generated Topic Pages">microRNA</a> (miRNA) which is responsible for sorafenib resistance at the genomic level.</p><h3>Methods</h3><p>CRISPR/Cas9 screen followed by gain- and loss-of-function assays both in vitro and in vivo were applied to identify the role of miR-3689a-3p in mediating sorafenib response in HCC. The upstream and downstream molecules of miR-3689a-3p and their mechanism of action were investigated.</p><h3>Results</h3><p>CRISPR/Cas9 screening identified miR-3689a-3p was the most up-regulated miRNA in sorafenib sensitive HCC. Knockdown of miR-3689a-3p significantly increased sorafenib resistance, while its overexpression sensitized HCC response to sorafenib <a href="https://www.sciencedirect.com/topics/medicine-and-dentistry/therapeutic-procedure" title="Learn more about treatment from ScienceDirect's AI-generated Topic Pages">treatment</a>. <a href="https://www.sciencedirect.com/topics/medicine-and-dentistry/proteomics" title="Learn more about Proteomic from ScienceDirect's AI-generated Topic Pages">Proteomic</a> analysis revealed that the effect of miR-3689a-3p was related to the copper-dependent mitochondrial <a href="https://www.sciencedirect.com/topics/pharmacology-toxicology-and-pharmaceutical-science/superoxide-dismutase" title="Learn more about superoxide dismutase from ScienceDirect's AI-generated Topic Pages">superoxide dismutase</a> type 1 (SOD1) activity. Mechanistically, miR-3689a-3p targeted the 3′UTR of the intracellular copper chaperone for superoxide dismutase (CCS) and suppressed its expression. As a result, miR-3689a-3p disrupted the intracellular copper trafficking and reduced SOD1-mediated scavenge of mitochondrial <a href="https://www.sciencedirect.com/topics/medicine-and-dentistry/oxidative-stress" title="Learn more about oxidative stress from ScienceDirect's AI-generated Topic Pages">oxidative stress</a> that eventually caused HCC cell death in response to sorafenib treatment. CCS overexpression blunted sorafenib response in HCC. Clinically, miR-3689a-3p was down-regulated in HCC and predicted favorable prognosis for HCC patients.</p><h3>Conclusion</h3><p>Our findings provide comprehensive evidence for miR-3689a-3p as a positive regulator and potential druggable target for improving sorafenib treatment in HCC.</p>