Human Endogenous Retrovirus-K affects cellular plasticity and generation of stem-like CD133+ cells in melanoma cancer cells

Background: Malignant melanoma is one of the most aggressive types of skin cancers and its exact etiology is not yet clear. Tumor cell plasticity and the putative cancer stem cell subpopulations that express stem cell markers such as CD133 have been associated with melanoma tumor initiation and progression. Likewise, human endogenous retrovirus-K (HERV-K) has been linked to aggressiveness and immune evasion of metastatic melanoma. The exact mechanism leading to abnormal HERV-K gene expression in melanoma is not yet clearly elucidated. However, environmental factors such as stress conditions seem to be involved along with mechanisms interfering with the immune system. Protein mutations are no longer considered as sole drivers of melanoma tumors, which implies studying the role of HERV-K activation in melanoma development and progression is important to understand melanomagenesis and find a possible therapeutic target. Objective of the study: To investigate the potential role of HERV-K activation in cellular plasticity and stemness features of melanoma cells upon the modification of the microenvironment conditions. Materials and methods: Four metastatic and one primary melanoma cell lines were used in this study; cell lines TVM-A12 and TVM-A12-CD133+ were established in our laboratory. Different cell culture media were used to assess the cellular modification of cells upon the modification of the microenvironment. Flow cytometry, RT-PCR analysis, RNA interference assay, sphere-forming assay and migration/invasion assays were used to assess cell phenotypic modifications, expression of HERVK, stemness and metastatic features of the cell lines, respectively. SPSS software version 17 was used for the statistical analysis and statistical significance was set at P< 0.050. Results: The peculiar plasticity features of TVM-A12 cells were indicated by their ability to show specific differentiated and non-adherent grape-like cellular aggregate phenotypes in specific differentiation and serum-free stem cell media, respectively. The grape-like cellular aggregates were also accompanied by increased activation of HERV-K expression and generation of the stem-like CD133+ subpopulations melanoma cells. Interestingly, silencing of HERV-K expression in TVM-A12 cells by RNA interference significantly abolished the generation of the stem-like CD133+ subpopulations, dysregulated the grape-like cellular aggregates phenotype and suppressed their proliferation. Correspondingly, TVM-A12-CD133+ cell line was able to show cellular plasticity upon the modification of the microenvironments and displayed a significantly higher self-renewing, migration and invasion capacity than the heterogeneous TVM-A12 cell lines. More interestingly, treatment of TVM-A12 and TVM-A12-CD133+ with the NNRTIs, nevirapine and efavirenz, inhibited the expression of HERV-K and significantly induced high levels of apoptosis in TVM-A12-CD133+ cells. Conclusion: Therefore, taking these evidences together, this study demonstrated for the first time that the plasticity of melanoma cancer cells and their potential to generate the stem-like CD133+ cells under stress conditions are dependent upon HERV-K expression. Hence, given the relevance of the putative CD133+ cancer stem cells in melanoma progression we suggest HERV-K expression as a potential target of melanoma therapy in order to enhance the anti-metastatic effects and improve patient survival.