Disulfiram: A Novel Repurposed Drug for Cancer Therapy
Cancer remains one of the most significant global health challenges, with its prevalence and mortality rates continuing to rise. Effective therapeutic strategies are crucial to prolong patient survival and reduce treatment costs. Drug repurposing, which involves identifying new therapeutic uses for approved drugs, has emerged as a promising approach. This strategy offers several advantages, including reduced research costs, shorter development timelines, and increased safety profiles. Disulfiram (DSF), a drug approved by the Food and Drug Administration (FDA) for the treatment of chronic alcoholism, has shown significant potential as an anticancer agent. This review explores the anticancer mechanisms of DSF, particularly in combination with copper (DSF/Cu), and its potential to target various human malignancies.
Introduction
Cancer is a leading cause of death worldwide, and its incidence is expected to increase in the coming years. Traditional cancer treatments include radical surgery, radiotherapy, immunotherapy, and chemotherapy. While chemotherapy remains one of the most effective treatments, it is often associated with significant side effects. The development of new anticancer drugs is challenging due to high costs and lengthy timelines. Drug repurposing offers a practical alternative by utilizing approved drugs with known toxicological and pharmacokinetic profiles for new indications.
Disulfiram (DSF) has been used since 1951 to treat chronic alcoholism and is well-tolerated with minimal side effects. DSF works by irreversibly inhibiting aldehyde dehydrogenase (ALDH), leading to the accumulation of acetaldehyde in the body, which establishes an alcohol aversion reflex. Recent evidence suggests that DSF has potential therapeutic applications beyond alcoholism, including inflammation, Lyme disease, metabolic disorders, and cancer. Mechanistic studies have revealed that DSF exhibits anticancer effects by triggering oxidative stress, inhibiting proteasome activity, reducing angiogenesis, arresting the cell cycle, reducing cancer stem cell (CSC) stemness, reversing drug resistance, constraining tumor metastasis, and regulating the immune microenvironment.
Anticancer Mechanisms of DSF/Cu
The anticancer effects of DSF are significantly enhanced when combined with copper (Cu). Copper is an essential trace metal that participates in various cellular processes, including mitochondrial respiration, reactive oxygen species (ROS) generation, and antioxidant/detoxification processes. Elevated levels of copper are often observed in cancer cells, where it promotes cell proliferation, angiogenesis, and metastasis. DSF binds to copper in tumor cells, impairing the activities of copper-dependent enzymes and inhibiting cuproplasia (copper-dependent cellular proliferation). Additionally, high concentrations of copper in cancer cells can induce cytotoxicity through oxidative stress or by inhibiting enzyme activity, leading to a specific form of copper-dependent cell death known as cuproptosis.
The major metabolite of DSF combined with copper, Cu(DDC)2 (bis-diethyldithiocarbamate-copper, also known as CuET), is the active form responsible for its tumor-suppressing effects. Cu(DDC)2 exhibits potent anticancer activity by altering ROS levels, activating the mitogen-activated protein kinase (MAPK) pathway, and inhibiting ubiquitin proteasome activity. Furthermore, DSF/Cu inhibits nuclear factor-kappa B (NF-kB) signaling, which plays a critical role in cancer cell survival and drug resistance.
Effects of DSF on ROS
Oxidative stress occurs when the accumulation of ROS exceeds the body’s antioxidant capacity. Increased ROS levels can be toxic, leading to the destruction of cellular structures and cell death. DSF-mediated cytotoxicity is partially caused by increased ROS production. DSF, DDC, and its copper complex Cu(DDC)2 accumulate in cancer cells, promoting ROS generation and triggering apoptosis. DSF/Cu-induced metallothionein expression results in oxidative stress and inhibits DNA replication in prostate cancer cells. Additionally, the reaction between DDC and Cu2+ reduces Cu2+ to Cu+, a more toxic form of copper ion, which further reacts with O2 and Fe2+ to produce highly cytotoxic hydroxyl radicals (·OH) through a Fenton-like reaction.
DSF also inhibits the scavenging of ROS. It downregulates glutathione peroxidase 4 (GPX4) expression, preventing ROS clearance and inducing ferroptosis in glioblastoma (GBM). DSF/Cu treatment leads to hepatocellular carcinoma (HCC) cell death via induction of ferroptosis, associated with a compensatory activation of the transcription factor nuclear factor erythroid 2-related factor 2 (NRF2), which plays a key role in counteracting oxidative stress. DSF, as a specific inhibitor of ALDH, prevents ROS scavenging and detoxification mediated by ALDH isozymes.
Effects of DSF on Proteasome Inhibition
The proteasome complex, which comprises a catalytic 20S core and a 19S regulator, selectively regulates and degrades ubiquitinated proteins. The ubiquitin proteasome system (UPS) is critical for maintaining protein degradation balance and cellular physiological functions. Cancer cells are more dependent on UPS than normal cells, making it an attractive pharmacological target for cancer therapy. DSF/Cu or Cu(DDC)2 blocks the upstream p97 pathway of the proteasome, inducing the accumulation of polyubiquitinated proteins and leading to cell death. Cu(DDC)2 induces higher cytotoxicity in various cancer cells compared to DSF or DDC alone.
Mechanistically, Cu(DDC)2, with high affinity for thiol-containing proteins, induces aggregation and dysfunction of the nuclear protein localization protein 4 (NPL4), an adaptor of the p97 segregase essential for proteasome activity. This leads to the accumulation of misfolded or toxic proteins, inducing endoplasmic reticulum (ER) stress and the heat-shock response (HSR). Inactivated p97 segregase also interferes with DNA replication, causing DNA damage and enhancing replication stress. Proteasome inhibition by DSF/Cu leads to the inhibition of NF-kB signaling, which is well-known for its antiapoptotic role in many malignant tumors.
DSF Targets Cancer Stem Cells (CSCs)
Cancer stem cells (CSCs) are a small population of quiescent cancer cells capable of self-renewal and differentiation, playing a critical role in tumor initiation, progression, relapse, metastasis, and resistance to therapy. Targeting CSCs is a promising strategy to improve cancer therapeutics. ALDH, a typical marker of CSCs, is irreversibly inhibited by DSF. Recent studies show that DSF potently inhibits CSCs in various cancers, including acute myeloid leukemia (AML), breast cancer, and ovarian cancer (OC), by targeting ALDH through diverse mechanisms.
DSF/Cu targets aldehyde dehydrogenase isoform-1A1 (ALDH1A1) to inhibit non-small cell lung cancer (NSCLC) growth and recurrence and to overcome cisplatin resistance in breast cancer by inhibiting stemness-related transcription factor expression in ALDH-positive CSCs. DSF efficiently inhibits ALDH activity and represses sphere formation under CSC-enriching conditions, demonstrating its ability to inhibit CSC formation in OC cells. DSF decreases CSC populations and reduces relapse in in vitro and in vivo models, highlighting its potential to prevent OC recurrence.
However, recent evidence challenges the notion that DSF-induced CSC toxicity is solely attributed to ALDH inhibition. Some studies suggest that DSF/Cu complexes block the formation of breast cancer CSCs by downregulating the NF-kB-stemness gene pathway. DSF combined with radiotherapy significantly inhibits mammary primary tumor growth and spontaneous lung metastasis, increasing DNA damage and inducing apoptosis and autophagy in irradiated CSCs.
DSF Reverses Drug Resistance
Drug resistance, either intrinsic or acquired, is a significant challenge in cancer treatment. Factors contributing to drug resistance include hypoxia, preexisting CSC populations, enhanced drug efflux pumps, and activation of NF-kB. DSF/Cu has been shown to reverse drug resistance in various cancers. For example, DSF/Cu lengthens survival and decreases the progression of NSCLC by inducing superoxide production and mitochondrial stress, mitigating resistance to radiation and chemotherapy.
DSF/Cu reverses Taxol-resistant (A549/Taxol) cells and vincristine-resistant cells (KB/VCR cells) by decreasing the expression of ALDH2 and stem cell transcription factors. DSF also inhibits ATP7A expression, increasing the levels of platinum–DNA adducts and apoptosis in human urothelial carcinoma (UC) cells. Additionally, DSF reverses chemoresistance in breast cancer cells by targeting the NF-kB pathway, enhancing sensitivity to ionizing radiation and chemotherapy.
Improved Drug Delivery Systems for DSF
Despite the promising anticancer effects of DSF/Cu, clinical studies have shown inconsistent outcomes, likely due to the rapid degradation of DSF or the unwanted modification of its metabolite DDC in the liver. To overcome these challenges, various drug delivery systems (DDSs) have been developed, including liposomes, polymers, polymeric micelles, and protein (albumin) particles encapsulating DSF/DDC. These systems protect the functional thiol groups of DDC, increasing drug concentrations at the lesion site and alleviating cytotoxicity.
Nanoparticles are easily captured by tumor cells, enhancing drug delivery and therapeutic efficacy. Codelivery systems for DSF and other chemotherapeutics have also been developed to efficiently overcome drug resistance and promote synergistic effects. These advancements in DSF-based treatment strategies hold significant potential for the future of cancer therapy.
Challenges and Perspectives
While DSF shows great promise as an anticancer agent, several challenges must be addressed for its successful clinical application. The cytotoxicity of DSF depends on copper, and supplementation with copper may be necessary for patients with copper deficiency. Balancing the efficacy of DSF with the strong cytotoxicity of DSF/Cu is crucial for safe and effective treatment. Additionally, DSF/Cu regulates the immune microenvironment and induces immunogenic cell death, but further research is needed to prevent excessive cytokine release and inflammatory responses.
Pharmacological interactions between DSF and comedications should be carefully investigated to ensure the success of future clinical trials. DSF can also be combined with other metals, such as zinc, which also exhibits anticancer activities. Despite these challenges, the potential of DSF as a cancer treatment is promising, and further studies should focus on its in-depth mechanisms and clinical applications.
Conclusion
Disulfiram, particularly in combination with copper, exhibits potent anticancer effects across various cancer types. Its mechanisms of action include the induction of intracellular ROS, inhibition of proteasome activity, and suppression of NF-kB signaling. DSF also targets cancer stem cells and reverses drug resistance, offering a novel approach to cancer treatment. Improved drug delivery systems and further research into its mechanisms and clinical applications will enhance the therapeutic potential of DSF in oncology.
doi.org/10.1097/CM9.0000000000002909
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