Nanomedicine is one of the many therapies currently being studied for treating cancer, one of the most challenging medical problems. The current study's objective was to investigate the potential anticancer effects of CuO NPs employing Ehrlich Ascitic carcinoma (EC) model. To accomplish the objective, a recent synthetic method along with the characterization of CuO NPs were exploited. 90 female albino mice were involved and randomly divided into 6 equal groups: divided into Control, untreated group (EC), and treated Groups. The Ascitic fluid from cancer groups and tissues from the control group were used for the assay of cytosolic GADPH and mitochondrial F1 beta subunit of ATP synthase level, Forkhead transcription factor O3a (FOXO3a) gene expression, and nitric oxide level. The results obtained showed a notable improvement in mean survival time and a decrease in cell viability, F1 beta subunit, and FOXO3a expression in the groups treated with CuO NPs, especially those receiving 10mg/kg every other day. Meanwhile, GAPDH and NO levels were decreased compared to the EC group. In conclusion, CuO NPs possess an antitumor effect by improving the bioenergetic signature.