Cancer stem cells (CSCs) express high levels of drug efflux transporters and 
antioxidant genes, and are therefore believed to be responsible for cancer 
recurrence following chemo/radiotherapy intervention. In this study, we 
investigated the role of NF-E2-related factor 2 (NRF2), a master regulator of 
antioxidant gene expression, in the growth and stress resistance of CSC-enriched 
mammosphere. The MCF7 mammospheres expressed significantly higher levels of the 
NRF2 protein and target gene expression compared to the monolayer. As underlying 
mechanisms, we observed that proteolytic activity and expression of the 
proteasome catalytic subunits were decreased in the mammospheres. Additionally, 
mammospheres retained a high level of p62 and the silencing of p62 was observed 
to attenuate NRF2 activation. NRF2 increase was confirmed in sphere-cultures of 
the colon and ovarian cancer cells. The functional implication of NRF2 was 
demonstrated in NRF2-knockdown mammospheres. NRF2-silenced mammospheres 
demonstrated increased cell death and retarded sphere growth as a result of 
target gene repression. Moreover, unlike the control mammospheres, 
NRF2-knockdown mammospheres did not develop anticancer drug resistance. 
Collectively, these results indicated that altered proteasome function and p62 
expression caused NRF2 activation in CSC-enriched mammospheres. In addition, 
NRF2 appeared to play a role in CSC survival and anticancer drug resistance.