The Molecular Basis for Inhibition of Stemlike Cancer Cells by Salinomycin

Tumors
are phenotypically heterogeneous and include subpopulations
of cancer cells with stemlike properties. The natural product salinomycin,
a K+-selective ionophore, was recently found to exert selectivity
against such cancer stem cells. This selective effect is thought to
be due to inhibition of the Wnt signaling pathway, but the mechanistic
basis remains unclear. Here, we develop a functionally competent fluorescent
conjugate of salinomycin to investigate the molecular mechanism of
this compound. By subcellular imaging, we demonstrate a rapid cellular
uptake of the conjugate and accumulation in the endoplasmic reticulum
(ER). This localization is connected to induction of Ca2+ release from the ER into the cytosol. Depletion of Ca2+ from the ER induces the unfolded protein response as shown by global
mRNA analysis and Western blot analysis of proteins in the pathway.
In particular, salinomycin-induced ER Ca2+ depletion up-regulates
C/EBP homologous protein (CHOP), which inhibits Wnt signaling by down-regulating
β-catenin. The increased cytosolic Ca2+ also activates
protein kinase C, which has been shown to inhibit Wnt signaling. These
results reveal that salinomycin acts in the ER membrane of breast
cancer cells to cause enhanced Ca2+ release into the cytosol,
presumably by mediating a counter-flux of K+ ions. The
clarified mechanistic picture highlights the importance of ion fluxes
in the ER as an entry to inducing phenotypic effects and should facilitate
rational development of cancer treatments.