**1. Introduction**

Melanoma is one of the most aggressive forms of malignant skin neoplasms and one of the main causes of cancer mortality. Although it represents only 4% of dermatological cancers, it is responsible for 80% of skin cancer deaths due to its high metastatic capacity and high refractoriness to chemotherapy [1–4]. The 5-year survival rate of patients with metastatic melanoma is less than 20% [5]. Dacarbazine, an alkylating agent, is the main treatment for advanced melanoma. However, serious side effects have been observed, and the therapeutic response rate is approximately 10% [5,6]. These results indicate that new therapies for melanoma are urgently required.

Oxidative stress plays a central role in the pathophysiology of melanoma since the generation of melanin leads to the generation of hydrogen peroxide and consumption of reduced glutathione (GSH) [7,8]. Consequently, melanoma maintains high baseline

**Citation:** Fontes, S.S.; Nogueira, M.L.; Dias, R.B.; Rocha, C.A.G.; Soares, M.B.P.; Vannier-Santos, M.A.; Bezerra, D.P. Combination Therapy of Curcumin and Disulfiram Synergistically Inhibits the Growth of B16-F10 Melanoma Cells by Inducing Oxidative Stress. *Biomolecules* **2022**, *12*, 1600. https://doi.org/10.3390/ biom12111600

Academic Editor: Vladimir N. Uversky

Received: 27 September 2022 Accepted: 16 October 2022 Published: 31 October 2022

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ROS levels compared to normal cells, which makes melanoma cells more susceptible to oxidative stress [7,9].

Curcumin (CUR) is a polyphenolic phytochemical isolated from turmeric, a food spice made from the rhizome of *Curcuma longa* L. [10]. Turmeric is traditionally used in many South Asian countries, both in traditional medicine and cooking, and has been used for over 2000 years as a medicine in China and India [11]. Curiously, many studies have reported that CUR causes cell death by stimulating reactive oxygen species (ROS) production in fibroblasts [12], leukemia [13], lymphoma [14], melanoma [15], and colon cancer [16].

Disulfiram (DSS) is a thiocarbamate drug approved by the United States Food and Drug Administration (US-FDA) for the clinical treatment of alcoholism and has been used for over 60 years [17]. DSS is an irreversible inhibitor of aldehyde dehydrogenase (ALDH). Interestingly, ALDH can decrease intracellular oxidative stress due to its ROS scavenger action. Consequently, DSS can act by inhibiting the intracellular antioxidant system [18,19].

Importantly, both DSS and CUR not only have antitumor activities but can also potentiate the action of anticancer chemotherapy drugs by blocking drug efflux pumps [20–26]. Furthermore, these compounds have been reported as agents capable of reducing the adverse reactions of highly toxic chemotherapeutic agents [27,28].

Therefore, we hypothesized that CUR, an oxidative stress inducer [29], combined with DSS, which acts as an antagonist of the intracellular antioxidant system [22], will have a synergistic effect, providing greater sensitization of cancer cells and resulting in cell death. Herein, we aimed to study the antitumor potential of the combination of CUR with DSS in B16-F10 melanoma cells using in vitro and in vivo models.
