Selective potentiation of paclitaxel (taxol)-induced cell death by mitogen-activated protein kinase kinase inhibition in human cancer cell lines

ABSTRACT Activation of the mitogen-activated protein kinase (MAPK) pathway in HeLa and Chinese hamster ovary cells after treatment with paclitaxel (Taxol) and other microtubule interacting agents has been investigated. Using a trans-reporting system, the phosphorylation of the nuclear transcription factors Elk-1 and c-jun was measured. Concentration-and time-dependent activation of the Elk-1 pathway, mediated primarily by the extracellular signal-regulated kinase (ERK) component of the MAPK family, was observed. Inactive drug analogs and other cytotoxic compounds that do not target microtubules failed to induce similar levels of activation, thereby indicating that an interaction between these drugs and the microtubule is essential for the activation of MAPKs. Evaluation of the endogenous levels of MAPK expression revealed cell-dependent expression of the ERK, c-jun N-terminal kinase, and p38 pathways. In the case of HeLa cells, time-dependent activation of ERK coincided with increased poly(ADP-ribose) polymerase (PARP) cleavage, phosphatidylserine externalization, and increased accumulation of cells in G 2 M. In both cell lines, inhibition of ERK activity potentiated paclitaxel-induced PARP cleavage and phosphatidylserine externalization, suggesting that ERK activity coincided with, but did not mediate, the cytotoxic effects of paclitaxel. We evaluated the nature of the interaction between paclitaxel and the MAPK kinase inhibitor U0126 in three cell lines, on the basis of a potential chemotherapeutic advantage of paclitaxel plus ERK inhibition. Our data confirmed additivity in those cells lines that undergo paclitaxel-induced ERK activation, and antagonism in cells with low ERK activity, suggesting that in tumors with high ERK activity, there may be an application for this strategy in therapy

Exploiting MEK inhibitor-mediated activation of ERα for therapeutic intervention in ER-positive ovarian carcinoma.

While the clinical benefit of MEK inhibitor (MEKi)-based therapy is well established in Raf mutant malignancies, its utility as a suppressor of hyperactive MAPK signaling in the absence of mutated Raf or Ras, is an area of ongoing research. MAPK activation is associated with loss of ERα expression and hormonal resistance in numerous malignancies. Herein, we demonstrate that MEKi induces a feedback response that results in ERα overexpression, phosphorylation and transcriptional activation of ER-regulated genes. Mechanistically, MEKi-mediated ERα overexpression is largely independent of erbB2 and AKT feedback activation, but is ERK-dependent. We subsequently exploit this phenomenon therapeutically by combining the ER-antagonist, fulvestrant with MEKi. This results in synergistic suppression of tumor growth, in vitro and potentiation of single agent activity in vivo in nude mice bearing xenografts. Thus, we demonstrate that exploiting adaptive feedback after MEKi can be used to sensitize ERα-positive tumors to hormonal therapy, and propose that this strategy may have broader clinical utility in ERα-positive ovarian carcinoma

MEK Inhibition Increases ERα Expression in Human Ovarian Carcinoma Cells.

<p>(A) Expression of ERα protein in human ovarian cancer cell lines. MCF-7, a breast cancer cell line was used as positive control. All cell lines were treated with MEKi at 1 uM for 24 h. (B) The effect of estrogen deprivation on cell cycle. Cells were grown in phenol red-free charcoal stripped RPMI for 48 h to simulate ES-free conditions, and subsequently analyzed for cell cycle distribution and doubling time, as described in Materials and Methods. (C) The effect of MEK inhibition for 24 h on ERα expression and MAPK pathway activation in ovarian cancer cells. DMSO was used as the vehicle-only control. (D) Dose-dependent increase in ERα expression in SKOV3 cells by MEKi (24 h); and densitometric quantification relative to GAPDH. (E) Flow cytometric analysis of cell-cycle distribution at various time points indicates G1 arrest 24 h post MEKi treatment in SKOV3 cells.</p