Ewing's sarcoma is the most aggressive connective tissue tumor, mainly affecting children and adolescents; the 5 year survival rate is only 50%. Current treatments have poor effectiveness, and more efficient treatments are being sought. Silver-based nanoparticles, such as silver chloride nanoparticles (AgCl-NPs) and silver/silver chloride (Ag/AgCl-NPs) nanoparticles, can be biologically produced and can release Ag+ ions into solution; however, their antitumor activity has been minimally investigated. The aim of this study was to evaluate the antitumor potential of AgCl-NPs and Ag/AgCl-NPs against Ewing's sarcoma cells. A673 cells (Ewing's sarcoma) were treated for 72 h with 0-12.5 µg ml-1 of Ag/AgCl-NPs or 0-40 µg ml-1 of AgCl-NPs. Human cells from the RPE-1 cell line (pigmented retinal epithelium) were used as a model of nontumor cells. The RPE-1 cells were less affected by the administration of AgCl-NPs or Ag/AgCl-NPs, with small reductions in the number of cells and viability and a small increase in apoptosis rates, while lysosomal damage, changes in reactive oxygen species (ROS) production, loss of mitochondrial membrane potential and alterations in microfilaments or cell areas were not observed. A673 tumor cells had significantly reduced number and viability levels when treated with AgCl-NPs, with reductions of 65.05% and 99.17%, respectively, whereas with Ag/AgCl-NP treatment, reductions of 65.53% and 92.51% were observed, respectively. When treated with silver-based nanoparticles, A673 cells also showed a significant increase in ROS production and loss of mitochondrial membrane potential, which culminated in an increase in the percentage of apoptosis among the population. Lysosomal damage was also observed when A673 cells were treated with the highest concentration of AgCl-NPs. In conclusion, the results showed that both AgCl-NPs and Ag/AgCl-NPs had some antitumor activity with minimal effects against healthy cells, which demonstrated the possibility of their use in cancer therapy.