Apoptosis Induction in Human Melanoma Cells by Inhibition of MEK Is Caspase-Independent and Mediated by the Bcl-2 Family Members PUMA, Bim, and Mcl-1

Abstract

Purpose: Given that inhibitors of mitogen-activated protein (MAP)/extracellular signalregulated kinase (ERK) kinase (MEK) are being introduced into treatment for melanoma, the present study was carried out to better understand the mechanism by which they may induce apoptosis of melanoma cells. Experimental Design: A panel of human melanoma cell lines and fresh melanoma isolates was assessed for their sensitivity to apoptosis induced by the MEK inhibitor U0126. The apoptotic pathways and regulatory mechanisms involved were examined by use of the inhibitor and small interfering RNA (siRNA) techniques. Results: Inhibition of MEK induced apoptosis in the majority of melanoma cell lines through a mitochondrial pathway that was associated with the activation of Bax and Bak, release of mitochondrial apoptogenic proteins, and activation of caspase-3. However, apoptosis was independent of caspases and instead was associated with mitochondrial release of AIF as shown by the inhibition of apoptosis when AIF was knocked down by siRNA. Inhibition of MEK resulted in the up-regulation of the BH3-only proteins PUMA and Bim and down-regulation of the antiapoptotic protein Mcl-1. These changes were critical for the induction of apoptosis by U0126 as siRNA knockdown of PUMA or Bim inhibited apoptosis, whereas siRNA knockdown of Mcl-1 increased apoptosis particularly in the apoptosis-resistant cell lines. Conclusions: Apoptosis of melanoma cells induced by the inhibition of the MEK/ERK pathway is mediated by the up-regulation/activation of PUMA and Bim and down-regulation of Mcl-1. Release of AIF rather than the activation of caspases seems to be the mediator of apoptosis. Our results suggest that cotargeting Mcl-1 and the MEK/ERK pathway may further improve treatment results in melanoma.

Tumour cell survival signalling by the ERK1/2 pathway

“…63 In a separate study, melanoma cell lines with high levels of ERK1/2 signalling underwent apoptosis when treated with U0126 and the increase in BIM and PUMA and the loss of MCL-1 appeared to play the critical role in cell death. 64 ERK1/2 signalling can also provide protection against chemotherapeutic cytotoxic drugs. In BRAF V600E -positive melanoma cells ERK1/2 signalling protects against apoptosis induced by cisplatin, actinomycin D or daunorubicin.…”

A naturally occurring cancer with molecular connectivity to human diseases

“…[12][13][14][15] Wild type p53 exists as a dimer or oligmer at the mitochondrial membrane and binds via its DNA binding domains with Bcl2 forming an inhibitory complex. Such binding disrupts the anti-apoptotic Bcl2/BAX ratio and results in outer mitochondrial membrane permeability and the release of toxic peptides from the outer compartment of the mitochondrion resulting in apoptosis.…”

The multikinase inhibitor sorafenib induces caspase-dependent apoptosis in PC-3 prostate cancer cells

“…This signalling pathway appears to be involved in prostate cancer drug resistance [7,8]. Furthermore, previous research has shown that inhibition of just one of the signalling proteins in this pathway can induce ap-npg optosis through down-regulation of myeloid cell leukaemia 1 (MCL-1), an anti-apoptotic member of the B-cell lymphoma 2 (BCL-2) gene family [9,10]. MCL-1 is expressed in a fairly high percentage of prostate tumours [11,12], and the inhibition of ERK pathway-mediated signals, and consequently expression of MCL-1, might be a key target for treatment of advanced prostate cancer cells, as suggested by Cavarretta et al [13].…”