A novel hypoxia-associated subset of FN1highMITFlowmelanoma cells: identification, characterization, and prognostic value

In many human cancers, the epithelial-to-mesenchymal transition has an important role in the induction of cancer stem-like cells, and hence, in the causation of intratumoral heterogeneity. This process, also referred to as mesenchymal mimicry, is, however, only poorly understood in melanoma and histological correlation is still lacking. In an immunohistochemical analysis of a large prospective series of 220 primary and metastatic melanomas for the well-known epithelial-to-mesenchymal transition marker FN1, we observed melanoma cells with high FN1 expression in metastases with ischemic necrosis, but rarely or not at all in samples lacking evidence of hypoxia. In a blinded, retrospective series of 82 melanoma metastases with 10-year follow-up, the presence of clusters of these FN1highmelanoma cells correlated significantly with shortened melanoma-specific survival, highlighting the prognostic value of their presence. We describe in detail the unique light- and electron-microscopic features of these FN1highmelanoma cells, enabling their identification in routinely hematoxylin-and-eosin-stained sections. In addition, by laser microdissection and subsequent gene expression analysis and immunohistochemistry, we highlight their distinctive, molecular phenotype that includes expression of various markers of the epithelial-to-mesenchymal transition (eg, ZEB1) and of melanoma stem-like cells (eg, NGFR), and lack of immunoreactivity for the melanocytic marker MITF. This phenotype could be reproduced in vitro by culturing melanoma cells under hypoxic conditions. Functionally, the hypoxic microenvironment was shown to induce a more migratory and invasive cell type. In conclusion, we identified a novel clinically relevant FN1highMITFlowcell type in melanoma associated with ischemic necrosis, and propose that these cells reside at the crossroad of the epithelial-to-mesenchymal transition and stem-like cell induction, plausibly triggered by the hypoxic environment.