Serious adverse effects and low selectivity to cancer cells are the main obstacles of long term therapy with Tamoxifen (Tmx). This study aimed to develop Tmx-loaded span-based nano-vesicles for delivery to malignant tissues with maximum efficacy. The effect of three variables on vesicle size (Y), zeta potential (Y), entrapment efficiency (Y) and the cumulative percent release after 24 h (Y) were optimized using Box-Behnken design. The optimized formula was prepared and tested for its stability in different storage conditions. The observed values for the optimized formula were 310.2 nm, - 42.09 mV, 75.45 and 71.70% for Y, Y, Y, and Y, respectively. The examination using electron microscopy confirmed the formation of rounded vesicles with distinctive bilayer structure. Moreover, the cytotoxic activity of the optimized formula on both breast cancer cells (MCF-7) and normal cells (BHK) showed enhanced selectivity (9.4 folds) on cancerous cells with IC values 4.7 ± 1.5 and 44.3 ± 1.3 μg/ml on cancer and normal cells, respectively. While, free Tmx exhibited lower selectivity (2.5 folds) than optimized nano-vesicles on cancer cells with IC values of 9.0 ± 1.1 μg/ml and 22.5 ± 5.3 μg/ml on MCF-7 and BHK cells, respectively. The promising prepared vesicular system, with greater efficacy and selectivity, provides a marvelous tool to overcome breast cancer treatment challenges.