BACKGROUND

Breast cancer is a heterogenous disease composed of multiple clonal populations and the mechanism by which the tumor microenvironment induces cancer stem cell plasticity is not fully understood.

METHODS

MCF7 breast cancer cells were treated with macrophage conditioned medium (MɸCM). PD98059 and SB431542 were used for ERK and TGF-βR inhibition respectively. Epithelial-mesenchymal transition (EMT) and cancer stem cell markers (CSC) were studied using qRT-PCR and flowcytometry. SCID mice were used for animal experiments.

RESULTS

MɸCM- induced ERK/TGF-β1 signaling led to enrichment of CSC and EMT in MCF7 cells and mammospheres. These effects were abrogated by both MEK inhibitor PD98059 (TGF-β1 synthesis) and SB431542 (TGF-β1 signaling). The increase in CSC was both hybrid (ALDH1) and mesenchymal (CD44 CD24 cells). Increase in hybrid E/M state was at a single cell level as confirmed by the increase in both claudin-1 (E) and vimentin (M). This did not have any growth advantage in SCID mice and monitoring of CSC and EMT markers before and after growth in SCID mice indicated reversal of these markers in tumor cells recovered from mice. Removal of MɸCM and neutralization of TNF-α, IL-6 and IL-1β in MɸCM abrogated ERK phosphorylation, TGF-β and CSC enrichment indicating the requirement of continuous signaling for maintenance.

CONCLUSIONS

ERK signaling plays an important role in MɸCM- induced EMT and CSC plasticity which is completely reversible upon withdrawal of signals.

GENERAL SIGNIFICANCE

Our experimental observations support the semi-independent nature of EMT-stemness connection which is very dynamic and reversible depending on the microenvironment.