Due to its potent anti-tumor activity, well-investigated pharmacokinetic properties and safety profile, disulfiram (DSF) has emerged as a promising candidate for drug repurposing in cancer therapy. Although several molecular mechanisms have been proposed for its anti-cancer effects, the precise underlying mechanisms remain unclear. In the present study, we showed that DSF inhibited proliferation of cancer cells by inducing reactive oxygen species (ROS) production, a G1 cell cycle arrest and autophagy. Moreover, DSF triggered apoptosis via suppression of the anti-apoptotic protein survivin.

To elucidate the mechanisms for the anti-proliferative activities of DSF, we applied a 2-DE combined with MALDI-TOF-MS/MS analysis to identify differentially expressed proteins in breast cancer cells upon treatment with DSF. Nine differentially expressed proteins were identified among which, three candidates including calmodulin (CaM), peroxiredoxin 1 (PRDX1) and collagen type I alpha 1 (COL1A1) are involved in the regulation of the AKT signaling pathway. The results of western blot analysis confirmed that DSF inhibited p-AKT, suggesting that DSF induces its anti-tumor effects via AKT blockade. Moreover, we found that DSF increased the mRNA levels of FOXO1, FOXO3 and FOXO4, and upregulated the expression of their target genes involved in G1 cell cycle arrest, apoptosis and autophagy. Finally, DSF potentiated the anti-proliferative effects of well-known chemotherapeutic agents such as arsenic trioxide (ATO), doxorubicin, paclitaxel and cisplatin. Altogether, these findings provide mechanistic insights into the anti-growth activities of DSF.

由于其强大的抗肿瘤活性、经过充分研究的药代动力学特性和安全性，双硫仑 (DSF) 已成为癌症治疗中药物再利用的有希望的候选者。尽管已经提出了几种抗癌作用的分子机制，但确切的潜在机制仍不清楚。在本研究中，我们表明 DSF 通过诱导活性氧 (ROS) 产生、G1 细胞周期停滞和自噬来抑制癌细胞的增殖。此外，DSF 通过抑制抗凋亡蛋白存活蛋白触发细胞凋亡。

为了阐明 DSF 抗增殖活性的机制，我们应用 2-DE 结合 MALDI-TOF-MS/MS 分析来鉴定用 DSF 治疗的乳腺癌细胞中差异表达的蛋白质。确定了九种差异表达的蛋白质，其中包括钙调蛋白 (CaM)、过氧还蛋白 1 (PRDX1) 和 I 型胶原蛋白 1 (COL1A1) 在内的三种候选蛋白参与了 AKT 信号通路的调节。Western印迹分析结果证实DSF抑制p-AKT，表明DSF通过AKT阻断诱导其抗肿瘤作用。此外，我们发现 DSF 增加了FOXO1、FOXO3和FOXO4的 mRNA 水平，并上调其参与 G1 细胞周期停滞、细胞凋亡和自噬的靶基因的表达。最后，DSF 增强了众所周知的化疗药物的抗增殖作用，如三氧化二砷 (ATO)、多柔比星、紫杉醇和顺铂。总而言之，这些发现为 DSF 的抗生长活动提供了机制上的见解。