EMT-Inducing Transcription Factors, Drivers of Melanoma Phenotype Switching, and Resistance to Treatment

Abstract

Transcription factors, extensively described for their role in epithelial-mesenchymal transition (EMT-TFs) in epithelial cells, also display essential functions in the melanocyte lineage. Recent evidence has shown specific expression patterns and functions of these EMT-TFs in neural crest-derived melanoma compared to carcinoma. Herein, we present an update of the specific roles of EMT-TFs in melanocyte differentiation and melanoma progression. As major regulators of phenotype switching between differentiated/proliferative and neural crest stem cell-like/invasive states, these factors appear as major drivers of intra-tumor heterogeneity and resistance to treatment in melanoma, which opens new avenues in terms of therapeutic targeting.

Keywords: EMT; ZEB2/ZEB1 switch; intra-tumor heterogeneity; melanocyte differentiation; melanoma development; neural-crest stem cells; phenotype switching; resistance to treatment; transcription factors.

Figure 1

Expression and function of epithelial–mesenchymal transition transcription factors (EMT-TFs) in the embryonic neural crest and the melanocyte lineage. (A) Schematic representation of ZEB1 and ZEB2 opposite expression patterns in the mouse embryo at embryonic day E8.5. (B) Expression of EMT-TFs during the formation of the melanocyte lineage from the embryonic neural crest. The transcription factor SOX10 is expressed in a bipotent melanoblast/glial progenitor. SNAIL2 and ZEB2 are required for neural crest cell delamination and melanoblast specification and migration through a dorsolateral pathway. SNAIL2 and ZEB2 are also required for the homeostasis of differentiated melanocytes in the epidermis, through the positive regulation of MITF expression. ZEB1 in contrast is expressed in dedifferentiated melanocyte stem cells, which have lost MITF expression. MITF: microphthalmia-associated transcription factor. DCT: dopachrome tautomerase. TYR: tyrosinase.

Figure 2

EMT-TF switch during melanoma development. ZEB2 and SNAIL2 are expressed in differentiated melanocytes within the epidermis of normal adult skin. Their expression is maintained in senescent benign nevi. During the conversion to primary melanoma, SNAIL2 and ZEB2 expression is progressively lost in favor of TWIST1 and ZEB1. Moreover, intra-tumoral heterogeneity of EMT-TFs is observed within melanoma lesions, the EMT-TF switch being generally observed as a gradient from upper part to deeper part of invasive melanoma. Gain of invasive capacity upon ZEB1 reactivation promotes melanoma cell entry into the bloodstream, with clusters of circulating melanoma cells, which may display different differentiation expression patterns. The differentiated pattern of EMT-TFs is reproduced in the secondary site following extravasation, allowing metastatic growth.

Figure 3

EMT-TFs regulate melanoma phenotype plasticity, intra-tumor heterogeneity and resistance to treatment. (A) Schematic model of putative melanoma intra-tumor heterogeneity according to a classification in five cell states, bearing various differentiation capacities, in accordance with their respective expression of MITF. Expected ZEB1 and ZEB2 expressions in the respective phenotypes are indicated. Expression of the main proposed markers of the different cell states is indicated. The invasive and stem-like phenotype actually represents two different states, both expressing high levels of ZEB1. SOX10 expression is present in all but the dedifferentiated state where it would be replaced by SOX9. The intermediate state should co-express ZEB1, ZEB2, and MITF. The intermediate state is believed to be highly metastable and to give rise to the invasive, neural crest stem cell (NCSC) and hyper-differentiated states, especially upon targeted therapy treatment-induced adaptation. Two-way arrows indicate reversible switching between cell states. Dotted arrows indicate putative transitions. The two main therapeutic strategies under investigation are also indicated. (B) Evolution of intra-tumor heterogeneity of melanoma during treatment with BRAF/MEK inhibitors-targeted therapy. The ZEB2/MITF melanocytic population is eliminated upon treatment and the tumor size decreases. Only resistant phenotypes remain with an increased proportion of NCSC-like and undifferentiated states. ZEB1-increased expression contributes to drug-induced NCSC reprogramming. Tumor adaptation ultimately leads to resistance with a regain of tumor growth and increased intra-tumor heterogeneity compared to the therapy naive tumor. BRAF/MEK resistant tumors may display high ZEB1 expression in both MITF^low and MITF^high clones. ZEB1 targeting can induce cell death in BRAFi-resistant melanoma cells, independently of MITF expression level.