Breast cancer is the most common cancer among women worldwide, with 20-30% of patients featuring the more aggressive and hard-to-treat erbB2 high variant of the pathology. Accumulating evidence suggests that breast cancer involves tumour-initiating cells (TICs), which may determine the progress and metastasis of the disease. Mitocans, a group of compounds that induce apoptosis of cancer cells by way of mitochondria destabilisation, are showing their potential in killing TICs. In this project, using the mammosphere model of breast TICs derived from murine NeuTL cells, and the syngenic mouse model with erbB2high breast tumours, we investigated the effect of mitochondrially targeted vitamin E succinate (MitoVES), a mitocan designed by our research team, for its in vitro and in vivo efficacy on breast TICs. We found that mammospheres derived from NeuTL cells were enriched in the level of stemness and that the sphere cells featured altered mitochondrial function. The sphere cultures were resistant to several established anti-cancer agents while they were susceptible to MitoVES. MitoVES is effective in apoptosis induction in NeuTL TICs by increasing the generation of reactive oxygen species and triggering the intrinsic apoptotic pathways. Compared with their differentiated counterparts, NeuTL TICs have higher mitochondrial membrane potential, higher level of mitochondrial complex II expression and higher respiration capacity via complex II, which contributes to their high susceptibility to MitoVES. Importantly, MitoVES inhibited the progression of syngeneic erbB2high tumours derived from breast TICs by inducing apoptosis in tumour cells. This suggests that targeting mitochondria of breast TICs is a promising approach to treat erbB2high breast tumours, which has a potential translational relevance.