Aberrant expression of embryonic epithelial-mesenchymal transition-inducing transcription factors (EMT-TFs) in epithelial cells triggers EMT, neoplastic transformation, stemness, and metastatic dissemination. We found that regulation and functions of EMT-TFs are different in malignant melanoma. SNAIL2 and ZEB2 transcription factors are expressed in normal melanocytes and behave as tumor-suppressor proteins by activating an MITF-dependent melanocyte differentiation program. In response to NRAS/BRAF activation, EMT-TF network undergoes a profound reorganization in favor of TWIST1 and ZEB1. This reversible switch cooperates with BRAF in promoting dedifferentiation and neoplastic transformation of melanocytes. We detected EMT-TF reprogramming in late-stage melanoma in association with enhanced phospho-ERK levels. This switch results in E-cadherin loss, enhanced invasion, and constitutes an independent factor of poor prognosis in melanoma patients.

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The aims of this study were to investigate, in cutaneous malignant melanoma (MM), the integrity of nuclear vitamin D receptor (VDR) signalling, as implied by VDR subcellular location; to investigate the relationship between VDR and tumour progression and the inhibitory effect on VDR by mitogen-activated protein kinase (MAPK) overactivity. Archived tissue from 34 benign melanocytic naevi, 149 MMs and 44 matched metastases were stained by immunohistochemistry for VDR and a subset of primary MMs were stained for phosphorylated-extracellular signal-regulated kinase as a marker of MAPK activity. MM cell lines were investigated to show the subcellular location of VDR and cell viability in response to ligand±MAPK inhibitor. Benign melanocytic naevi showed mainly a strong nuclear VDR staining in contrast to MM where decreased nuclear and emergent cytoplasmic VDRs were associated with malignant progression in terms of dermal invasion and metastasis. MMs that retained exclusive nuclear VDR at the tumour base did not metastasize, a potentially important prognostic indicator. Decreased nuclear VDR correlated with increased cytoplasmic staining, suggesting the failure of nuclear entry as a primary cause of defective VDR signalling in MM. The histological subset analysis and MM cell line studies confirmed the inhibitory effect of MAPK activity on VDR signalling, but the pattern of VDR subcellular localization suggested failure of VDR nuclear entry as a primary effect of MAPK activity rather than direct inhibition of VDR-regulated transcription. Furthermore, high MAPK activity in tumours expressing cytoplasmic VDR was associated with worsened prognosis.

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Embryonic transcription factors (ETFs) of the Snail, Zeb and Twist families are essential mediators of EMT programs (Epithelial to Mesenchymal Transition) and play central roles in cell determination and cell plasticity. The aberrant expression of ETFs is frequently observed in various cancer types, particularly in carcinomas, and is associated with poor prognosis and high risk of metastasis. We and others have demonstrated that their oncogenic potential relies on their ability to enable escape from oncogene-induced failsafe programs, senescence and apoptosis. Therefore, EMT-resembling processes initiated by ETFs contribute to neoplastic transformation in vitro and accelerate tumor progression in vivo in an epithelial context. We now unveil a completely different scenario in melanoma, a highly metastatic neural crest-derived cancer. Consistent with their role in melanoblasts determination, SNAIL2 and ZEB2 are already expressed in melanocytes and naevi, but their expression is down-regulated in primary melanoma. In contrast, TWIST1 and ZEB1 are aberrantly reactivated in primary and metastatic melanoma. Interestingly, this switch in ETFs expression represents a novel independent factor of poor prognosis in patients with malignant melanoma. We further demonstrate that BRAF activation, the most frequent driving event in melanoma, induces a drastic upregulation of TWIST1 and ZEB1 at the expense of SNAIL2 and ZEB2 in murine and human melanocytes. This reversible switch, orchestrated by the FRA1 transcription factor, is determinant for BRAF-driven melanocyte transformation in vitro and in a xenograft model. As a whole, while TWIST1 and ZEB1 cooperate with oncogenic BRAF to induce neoplastic transformation, SNAIL2 and ZEB2 rather behave as oncosuppressive proteins in melanoma. Gene expression profiles analyses further show that this oncogenic reprogramming of ETFs regulates the expression of the MITF master regulator, with an inhibition of MITF-regulated differentiation program and an activation of TGFβ- and invasion-associated gene expression signatures. Collectively, these data highlight yet unexpected antagonistic properties of ETFs in tumor progression, and place the ETF network as a master regulator of MITF-dependent phenotype switching during melanomagenesis. Citation Format: Julie Caramel, Eftychios Papadogeorgakis, Louise Hill, Gareth J. Browne, Geoffrey Richard, Anne Wierinckx, Gerald Saldanha, Peter Hutchinson, James H. Pringle, Alain Puisieux, Eugene Tulchinsky, Stephane Ansieau. Antagonistic functions of EMT-inducing transcription factors during melanoma development. [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 285. doi:10.1158/1538-7445.AM2013-285