Melanoma can be stratified into unique subtypes based on distinct pathologies. The acral/mucosal melanoma subtype is characterized by aberrant and constitutive activation of the proto-oncogene ...receptor tyrosine kinase C-KIT, which drives tumorigenesis. Treatment of these melanoma patients with C-KIT inhibitors has proven challenging, prompting us to investigate the downstream effectors of the C-KIT receptor. We determined that C-KIT stimulates MAP kinase-interacting serine/threonine kinases 1 and 2 (MNK1/2), which phosphorylate eukaryotic translation initiation factor 4E (eIF4E) and render it oncogenic. Depletion of MNK1/2 in melanoma cells with oncogenic C-KIT inhibited cell migration and mRNA translation of the transcriptional repressor SNAI1 and the cell cycle gene CCNE1. This suggested that blocking MNK1/2 activity may inhibit tumor progression, at least in part, by blocking translation initiation of mRNAs encoding cell migration proteins. Moreover, we developed an MNK1/2 inhibitor (SEL201), and found that SEL201-treated KIT-mutant melanoma cells had lower oncogenicity and reduced metastatic ability. Clinically, tumors from melanoma patients harboring KIT mutations displayed a marked increase in MNK1 and phospho-eIF4E. Thus, our studies indicate that blocking MNK1/2 exerts potent antimelanoma effects and support blocking MNK1/2 as a potential strategy to treat patients positive for KIT mutations.
The progression of cancers from primary tumors to invasive and metastatic stages accounts for the overwhelming majority of cancer deaths. Understanding the molecular events which promote metastasis ...is thus critical in the clinic. Translational control is emerging as an important factor in tumorigenesis. The messenger RNA (mRNA) cap-binding protein eIF4E is an oncoprotein that has an important role in cancer initiation and progression. eIF4E must be phosphorylated to promote tumor development. However, the role of eIF4E phosphorylation in metastasis is not known. Here, we show that mice in which eukaryotic translation initiation factor 4E (eIF4E) cannot be phosphorylated are resistant to lung metastases in a mammary tumor model, and that cells isolated from these mice exhibit impaired invasion. We also demonstrate that transforming growth factor-beta (TGFβ) induces eIF4E phosphorylation to promote the translation of Snail and Mmp-3 mRNAs, and the induction of epithelial-to-mesenchymal transition (EMT). Furthermore, we describe a new model wherein EMT induced by TGFβ requires translational activation via the non-canonical TGFβ signaling branch acting through eIF4E phosphorylation.
The translation initiation factor eIF4E is an oncogene that is commonly overexpressed in primary breast cancers and metastases. In this article, we report that a pharmacologic inhibitor of eIF4E ...function, ribavirin, safely and potently suppresses breast tumor formation. Ribavirin administration blocked the growth of primary breast tumors in several murine models and reduced the development of lung metastases in an invasive model. Mechanistically, eIF4E silencing or blockade reduced the invasiveness and metastatic capability of breast cancer cells in a manner associated with decreased activity of matrix metalloproteinase (MMP)-3 and MMP-9. Furthermore, eIF4E silencing or ribavirin treatment suppressed features of epithelial-to-mesenchymal transition, a process crucial for metastasis. Our findings offer a preclinical rationale to explore broadening the clinical evaluation of ribavirin, currently being tested in patients with eIF4E-overexpressing leukemia, as a strategy to treat solid tumors such as metastatic breast cancer.
Melanoma can be stratified into unique subtypes based on distinct pathologies. The acral/mucosal melanoma subtype is characterized by aberrant and constitutive activation of the proto-oncogene ...receptor tyrosine kinase C-KIT, which drives tumorigenesis. Treatment of these melanoma patients with C-KIT inhibitors has proven challenging, prompting us to investigate the downstream effectors of the C-KIT receptor. We determined that C-KIT stimulates MAP kinase-interacting serine/threonine kinases 1 and 2 (MNK1/2), which phosphorylate eukaryotic translation initiation factor 4E (eIF4E) and render it oncogenic. Depletion of MNK1/2 in melanoma cells with oncogenic C-KIT inhibited cell migration and mRNA translation of the transcriptional repressor SNA/1 and the cell cycle gene CCNE1. This suggested that blocking MNK1/2 activity may inhibit tumor progression, at least in part, by blocking translation initiation of mRNAs encoding cell migration proteins. Moreover, we developed an MNK1/2 inhibitor (SEL201), and found that SEL201-treated KIT-mutant melanoma cells had lower oncogenicity and reduced metastatic ability. Clinically, tumors from melanoma patients harboring K/T mutations displayed a marked increase in MNK1 and phospho-eIF4E. Thus, our studies indicate that blocking MNK1/2 exerts potent antimelanoma effects and support blocking MNK1/2 as a potential strategy to treat patients positive for K/T mutations.
Abstract
The eukaryotic translation initiation factor 4E (eIF4E) is an oncogene that drives cellular transformation, tumorigenesis and metastasis by facilitating nuclear export and translation of ...specific mRNAs, including cyclins, c-myc, matrix metalloproteinases (MMPs), VEGF and others. eIF4E is commonly overexpressed and/or activated in cancer, and we and others have shown that high eIF4E expression in primary and metastatic breast cancers correlates with poor prognosis. Recent studies have shown that Ribavirin, an old antiviral drug, can inhibit oncogenic transformation mediated by eIF4E and exert antitumor activity in cancer cell lines, animal models, and patients with acute myeloid leukemia (AML) that express elevated eIF4E. Dramatic clinical responses in AML correlated with reduced eIF4E level and activity. Ribavirin is thus the first drug to date with antitumor activity linked to direct inhibition of eIF4E in patients.
In this study, we have evaluated the potential anti-metastatic activity of ribavirin. We first assessed the role of eIF4E in the epithelial-to-mesenchymal transition (EMT), a process enabling metastasis. We showed that either knockdown of eIF4E or Ribavirin treatment attenuate EMT induced by TGFβ in normal mammary epithelial cells, blocking the increase of mesenchymal markers as well as MMP-9 secretion and decreasing cell migration and invasion.
We then studied the effects of eIF4E targeting on triple negative breast cancer cells in vitro and in vivo. We have previously shown that ribavirin suppresses breast cancer cell proliferation and this is associated with a decrease in known eIF4E targets. Using human and murine cell lines, we further observed a reduction in several mesenchymal markers, including N-cadherin, fibronectin and snail, following knockdown or chemical inhibition of eIF4E. Moreover, ribavirin significantly reduced tumor cell motility and invasion through Matrigel. Interestingly, overexpression of snail in MDA-MB-231 cells blunts their response to ribavirin, suggesting its involvement downstream of eIF4E.
The activity of ribavirin against mammary tumors in vivo was assessed in two syngeneic mouse models, MT2186 and 66cl4, which form tumors in FVB and Balb/c mice, respectively. A non-toxic and clinically relevant dose of Ribavirin caused a striking reduction in MT2186 tumor growth, which correlated with reduced Ki-67 staining and reduced clonogenic potential of cells isolated from ribavirin treated tumors. Growth of the more aggressive and highly metastatic 66cl4 tumors was modestly affected; with only a slight delay in primary tumor growth. However, preliminary data suggest that ribavirin significantly suppresses the formation of 66cl4 lung metastases. Importantly, we measured MMP levels in plasma and found a selective increase in MMP-9 with growth of tumors over time, which was suppressed in the ribavirin treated mice. Future studies will further define the mechanism responsible for the anti-metastatic activity of ribavirin in this model.
Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A154.
Citation Format: Audrey Emond, Sonia V. Del Rincon, Bonnie Huor, Filippa Pettersson, Wilson H. Miller. Inhibition of eIF4E with Ribavirin suppresses EMT and breast cancer invasiveness. abstract. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A154.
Abstract
The epithelial-mesenchymal-like transition (EMT) is a process enabling epithelial cells to gain the motile characteristics of mesenchymal cells, in a manner resembling metastasis. TGF-beta, ...via well-defined transcriptional mechanisms, is considered a master regulator of EMT. However, the idea that TGF-beta can regulate the translational machinery to drive EMT remains largely unexplored. The eukaryotic translation initiation factor eIF4E is known to be overexpressed in breast cancer, has been linked to increased invasiveness, and is a promising target for the treatment of breast cancer. Our hypothesis is that phosphorylation of eIF4E stimulated by TGF-beta is required for inducing EMT and metastasis in breast cancer. Our novel preliminary data show that TGF-beta can stimulate eIF4E phosphorylation as normal mammary epithelial cells become mesenchymal. Silencing of eIF4E attenuates molecular and behavioral changes associated with EMT. Moreover, decreasing eIF4E levels can impair TGF-beta induced migration and invasion of ErbB2-expressing breast cancer cells. In keeping with a role of phosphorylated eIF4E in driving the metastatic phenotype, we show that chemically and genetically inhibiting the eIF4E kinase MNK1 attenuates TGF-beta-stimulated EMT. We hypothesized that the expression of master regulators of EMT could be restricted when the eIF4E/MNK pathway is functionally impaired, which prompted us to look at the expression of Twist and Snail in TGF-beta treated eIF4E- and MNK- silenced cells. Our data shows that TGF-beta-induced Snail protein expression, but not mRNA level, is repressed when the MNK/eIF4E pathway is functionally impaired. Our results indicate that MNK and eIF4E are essential for EMT and suggest that therapeutic inhibition of MNK/eIF4E pathway may be a useful strategy for the control of tumor invasion and metastasis.
Citation Format: Sonia V. del Rincon, Bonnie Huor, Elaine Ngan, Luca Petruccelli, Peter Siegel, Wilson H. Miller, Jr. TGF-beta induced epithelial-to-mesenchymal transition is attenuated when the MNK/eIF4E pathway is functionally impaired. abstract. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 288. doi:10.1158/1538-7445.AM2013-288
PDF
