Overcoming the compensatory elevation of NRF2 renders hepatocellular carcinoma cells more vulnerable to disulfiram/copper-induced ferroptosis

Abstract

Hepatocellular carcinoma (HCC) is one of the paramount causes of cancer-related death worldwide. Despite recent advances have been made in clinical treatments of HCC, the general prognosis of patients remains poor. Therefore, it is imperative to develop a less toxic and more effective therapeutic strategy. Currently, series of cellular, molecular, and pharmacological experimental approaches were utilized to address the unrecognized characteristics of disulfiram (DSF), pursuing the goal of repurposing DSF for cancer therapy. We found that DSF/Cu selectively exerted an efficient cytotoxic effect on HCC cell lines, and potently inhibited migration, invasion, and angiogenesis of HCC cells. Importantly, we confirmed that DSF/Cu could intensively impair mitochondrial homeostasis, increase free iron pool, enhance lipid peroxidation, and eventually result in ferroptotic cell death. Of note, a compensatory elevation of NRF2 accompanies the process of ferroptosis, and contributes to the resistance to DSF/Cu. Mechanically, we found that DSF/Cu dramatically activated the phosphorylation of p62, which facilitates competitive binding of Keap1, thus prolonging the half-life of NRF2. Notably, inhibition of NRF2 expression via RNA interference or pharmacological inhibitors significantly facilitated the accumulation of lipid peroxidation, and rendered HCC cells more sensitive to DSF/Cu induced ferroptosis. Conversely, fostering NRF2 expression was capable of ameliorating the cell death activated by DSF/Cu. Additionally, DSF/Cu could strengthen the cytotoxicity of sorafenib, and arrest tumor growth both in vitro and in vivo , by simultaneously inhibiting the signal pathway of NRF2 and MAPK kinase. In summary, these results provide experimental evidence that inhibition of the compensatory NRF2 elevation strengthens HCC cells more vulnerable to DSF/Cu induced ferroptosis, which facilitates the synergistic cytotoxicity of DSF/Cu and sorafenib.

Construction and Validation of a 6-Ferroptosis Related Gene Signature for Prognosis and Immune Landscape Prediction in Melanoma

“…Combining Chemotherapeutic drugs and ICPs inhibitors might lead to increased tumor immunogenicity and immune cell infiltration by triggering ferroptosis of tumor cells (J. Du et al, 2021 ; Ren et al, 2021 ), and in this way, improving the efficacy of immunotherapy ( Xu et al, 2021 ). Some ferroptosis related small-molecules, such as statins, have also been valued in anti-tumor combination drugs ( Tang et al, 2020 ).…”

Downregulated Copper Homeostasis-Related Gene FOXO1 as a Novel Indicator for the Prognosis and Immune Response of Breast Cancer

“…Its typical examples are clioquinol and disulfiram [ 24 ]. Mechanistically, these compounds can function by producing intracellular reactive oxygen species (ROS) and inhibiting proteasome activities, causing the apoptosis of cancer cells [ 31 ]. Overall, Cu homeostasis has a significant effect on various cancers and suggests new and potential anticancer approaches.…”

The Disulfiram/Copper Complex Induces Autophagic Cell Death in Colorectal Cancer by Targeting ULK1

“…The key mechanism of the antitumor role played by DSF is that it can combine with copper to form complexes ( Guo et al, 2010 ; Skrott et al, 2017 ); DSF/Cu incidentally induces apoptotic cell death in breast cancer via inhibition of the proteasome activity ( Chen et al, 2006 ). Moreover, DSF/Cu could intensively impair mitochondrial homeostasis, enhance lipid peroxidation, and eventually result in ferroptotic cell death in hepatocellular carcinoma ( Ren et al, 2021 ). In our study, we further confirm the Cu-dependent cytotoxic effect of DSF on CRC cells.…”