Estudo do potencial antineoplásico do ácido rosmarínico em células de melanoma cutâneo

Abstract

Introduction: cutaneous melanoma is a malignant neoplasm that has a poor prognosis in stages III and IV, due to the failure of systemic treatment. With technological advances, several bioactive compounds of natural origin have been studied and have shown promising results as a form of adjuvant therapy for malignant neoplasms. One such compound that stands out is rosmarinic acid (RA). Likewise, the purinergic system has been pointed out by several studies as an interesting target in antineoplastic therapy. On the other hand, oxidative stress also seems to be involved in the pathophysiology of cancer. Objective: to analyze in vitro the antineoplastic potential of rosmarinic acid in cutaneous melanoma cells through parameters of oxidative stress and purinergic signaling. Methodology: SK-MEL-28 cells were grown in DMEM medium under minimal conditions necessary. Neoplastic negative control cells were treated only with culture medium. Next, the cultures were treated with concentrations of 50 μM, 100 μM, 200 μM, 400 μM and 800 μM of AR for 24 hours. They were also used as peripheral blood mononuclear cells to verify cell viability and cytotoxicity in non-neoplastic cells. After treatment, the supernatant and neoplastic cells were collected for the evaluation of cell viability and cytotoxic potential, parameters of oxidative stress and purinergic system. Oxidative stress parameters were evaluated through levels of PSH and NPSH antioxidant defenses, and ROS prooxidants. The gene expression of caspase 8 and 3, inflammasome (NLRP3), the enzymatic activity of caspase 3, and the formation of apoptotic bodies were also evaluated. Finally, regarding the purinergic system, gene and protein expression of CD39 and CD73, enzymatic activity of CD39, CD73 and ADA, and quantification of extracellular levels of ATP were verified. Results: It was found that RA significantly reduced the viability of SK-MEL-28, while it had no cytotoxic effect on non-neoplastic cells. In addition, at high concentrations they also showed antioxidant potential, promoting a reduction in ROS levels and an increase in PSH and NPSH levels. AR increased the gene expression of caspases e 8 and 3, and reduced the expression of NLRP3. Caspase 3 enzymatic activity was also increased. It was possible to detect the formation of apoptotic bodies at concentrations of 400 μM and 800 μM. As for the purinergic parameters, the AR in high concentrations modulated the gene and protein expression of CD73. The enzymatic activity of the CD39, CD73 and ADA ectonucleotidases was also modulated. No change in extracellular ATP levels was detected. Conclusion: RA is a powerful candidate to be study in clinical trials for the treatment of CM. In this study with CM cells of the SK-MEL-28 lineage, the RA showed antineoplastic effect on reduction of cell viability, modulated the gene and protein expression of apoptotic mediators, contributed to the redox balance, as well as modulated the purinergic signaling.