The involvement of the MAPK signalling pathway in the adaptation of MCF-7 cells to long-term oestrogen deprivation

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The molecular mechanism associated with the transition of breast tumours to steroid hormone-independent growth is poorly understood. However, a number of studies have demonstrated the potential role of the mitogen-activated protein kinase (MAPK) signalling pathway in the initiation and pathogenesis of breast cancer.

In an attempt to study the transition to oestrogen-independent growth, wild-type (wt) MCF-7 cells were cultured in oestrogen (E2)-deficient medium for over 100 weeks. During this time the cells were characterised and shown to pass through three distinct phases. Quiescent (LTED-Q), followed by an increase in basal growth rate paralleled by hypersensitivity to E2 (LTED-H), and finally transition to an E2-independent phase (LTED-I). Western blot analysis of the LTED cells showed elevated levels of ERα compared to the wt MCF-7 cells. The ERα was phosphorylated on serine 118 despite absence of E2.

Assessment of the level of activated MAPK showed a mean 7-fold increase in LTED-Q, 4- fold in LTED-H and 6-fold in LTED-I compared to the wt MCF-7 cells. As the LTED-I phase progressed MAPK levels fell, but after 90 weeks remained 30% higher compared to wt MCF-7. Suppression of MAPK activity in LTED-I cells, using a MEK inhibitor, significantly reduced but did not block ERα phosphorylation. Similarly transfection of LTED-I cells with an E2 responsive reporter construct (pEREtkCAT), followed by treatment of the cells with a MEK inhibitor, resulted in a 50% decrease in basal ERα transcription. However, a combination of E2 and the MEK inhibitor suppressed ERα-directed transcription by only 30% compared to E2 alone.

These data support previous findings that elevated MAPK levels are found during ligand-independent cell proliferation. However, this is unlikely to be the sole pathway operating to achieve this adaptation; rather a complex network of kinases and molecular switches may operate at different temporal stages during long-term oestrogen deprivation.