The association between superparamagnetic iron oxide nanoparticles (SPION), carrying small interfering RNA (siRNA) as therapeutic agents and humanized anti- human epidermal growth factor receptor-2 (HER2) single-chain antibody fragments (scFv) for the active delivery into HER2-overexpressing cells appears as an interesting approach for patients with HER2-overexpressing advanced breast cancer. The obtained Targeted Stealth Magnetic siRNA Nanovectors (TS-MSN) are formulated by combining: (i) the synthesis protocol of Targeted Stealth Fluorescent Particles (T-SFP) which form the core of TS-MSN and (ii) the formulation protocol allowing the loading of T-SFP with polyplexes (siRNA and cationic polymers). TS-MSN have suitable physico-chemical characteristics for intravenous administration and protect siRNA against enzymatic degradation up to 24 h. The presence of HER2-targeting scFv on TS-MSN allowed an improved internalization (3-4 times more compared to untargeted S-MSN) in HER2-overexpressing breast cancer cells (BT-474). Furthermore, anti-survivin siRNA delivered by TS-MSN in HER2-negative breast-cancer control cells (MDA-MB-231) allowed significant down-regulation of the targeted anti-apoptotic protein of about 70%. This protein down-regulation increased in HER2 + cells to about 90% (compared to 70% with S-MSN in both cell lines) indicating the contribution of the HER2-active targeting. In conclusion, TS-MSN are promising nanocarriers for the specific and efficient delivery of siRNA to HER2-overexpressing breast cancer cells.