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Background: Breast cancer is one of the most frequent cancer types within female populations. Silibinin is a chemotherapeutic agent active against cancer. Niosomes are biodegradable, biocompatible, safe and effective carriers for drug delivery., Objective: To prepare nanoniosomal silibinin and evaluate its cytotoxicity in the T-47D breast cancer cell line., Materials and Methods: Niosomes were prepared by reverse phase evaporation of a mixture of span 20, silibinin, PEG-2000 and cholesterol in chloroform and methanol solvent (1:2 v/v). The solvent phase was evaporated using a rotary evaporator and the remaining gel phase was hydrated in phosphate buffer saline. Mean size, size distribution and zeta potential of niosomes were measured with a Zetasizer instrument and then nanoparticles underwent scanning electron microscopy. The drug releasing pattern was evaluated by dialysis and the cytotoxicity of nanoniosomes in T-47D cells was assessed by MTT assay., Results: Particle size, size variation and zeta potential of the niosomal nanoparticles were measured as 178.4 ± 5.4 nm, 0.38 ± 0.09 and -15.3 ± 1.3 mV, respectively. The amount of encapsulated drug and the level of drug loading were determined 98.6 ± 2.7% and 22.3 ±1.8%, respectively; released drug was estimated about 18.6±2.5% after 37 hours. The cytotoxic effects of nanoniosome were significantly increased when compared with the free drug., Conclusions: This study findings suggest that silibinin nanoniosomes could serve as a new drug formulation for breast cancer therapy.