Role of Nrf2 in drug resistance of cancer stem-like cells

抄録

Cancer stem cells (CSCs) are known to express high levels of antioxidant proteins and drug efflux transporters, and are therefore believed to be responsible for tumor recurrence after chemotherapy. Several CSC markers such as cluster of differentiation 44 (CD44) and aldehyde dehydrogenase (ALDH) have been used to isolate CSC population from cancer cell lines and tumor masses. In this study, we investigated the role of NF-E2-related factor 2 (NFE2L2; NRF2), a key regulator of the expression of antioxidant and detoxifying genes, in CSCs resistance. First, we isolated the CD44-high cell population (ADR44P) from doxorubicin-resistance breast cancer MCF7 and utilized it as a CSC-like cell line system. The ADR44P cells expressed the standard isoform of CD44 (CD44s) and displayed relatively high resistance to doxorubicin and oxidative stress compared to the CD44-low MCF7. Particularly, NRF2 and its target genes expression were substantially high in ADR44P. The treatment of ADR44P cells with hyaluronic acid (HA), a CD44 ligand, further activated NRF2 expression, whereas the inhibition of HA synthesis attenuated NRF2 activation, implying the direct activating role of CD44 in NRF2 signaling. In line with these, overexpression of CD44s in MCF7 led to NRF2 activation. As an underlying mechanism of CD44s-meidated NRF2 activation, it was observed that p62 levels were increased and its suppression resulted in the blockade of NRF2 activation in ADR44P cells. As functional implications of NRF2 activation in CD44+ CSC-like cells, the silencing of NRF2 rendered ADR44P cells sensitive to anticancer drugs, and NRF2-knockdown ADR44P cells displayed a significant tumor growth retardation in mouse xenografts. In addition to CD44, we observed that the aldehyde dehydrogenase (ALDH)-enriched ovarian cancer stem-like cells expressed high levels of NRF2 and its target genes, and stable silencing of NRF2 led to the reductions in CSC markers expression, colony formation, sphere formation, doxorubicin resistance, and tumor growth in xenografts model. Similarly to CD44+ CSCs-like cells, ALDH+ CSC-like cells showed p62 elevation, and the inhibition of p62 suppressed NRF2 signaling. Collectively, these results showed that CSCs-like cells activated NRF2 signaling, which can endow CSCs with stress resistance, aggressive phenotypes, and tumor growth.