Reciprocal Interactions between Melanoma and Microglia Cells Reprogram Melanoma Malignancy Phenotype

Introduction: Melanoma has one of the highest propensities to metastasize to the brain. Once disseminated in the brain, melanoma cells communicate with microglia (MG), the main immunocompetent cells in the CNS. The present study is aimed to elucidate the role of MG in the formation and survival of melanoma brain metastasis (MBM).

Methods: Human and mouse brain specimens bearing MBM were stained by IHC for the MG marker Iba1. Melanoma cells sorted from melanoma-MG co-cultures as well as naïve melanoma cells were inoculated subcutaneously into nude mice to determine the effect of MG on melanoma tumorigenicity. Human MBM cells from different patients were exposed to human MG cells or MG-derived soluble factors. Human MG cells were similarly exposed to MBM cells or their soluble factors. RNA-seq, RPPA, cytokine arrays and phospho-protein arrays were used to determine the alterations in each of these cell types following the exposure to the counter cell type. Treated cells were tested for alterations in migration ability, matrix metalloproteinase-2 (MMP2) activity, proliferation, morphology, and spheroid formation. Melanoma cells were also tested for transendothelial migration. Similar in vitro assays were performed using melanoma cells with high or low Aldolase C (ALDOC) expression to demonstrate its role in the formation and maintenance of MBM.

Results and discussion: IHC demonstrated infiltration of MG cells to MBM lesions and a close spatial distance between the two cell types. Subcutaneous inoculation of melanoma cells sorted from melanoma-MG co-cultures resulted in increased tumor size, compared to naïve melanoma cells. Reprograming of gene expression, cell signaling and cytokine secretion occurred in MBM and MG cells following the exposure of each cell type to supernatants of the counter cell type. Melanoma cells exerted morphological changes on MG cells, enhanced their proliferation, MMP2 activity and migration. MG cells enhanced melanoma malignant phenotype: increased melanoma proliferation, migration, MMP2 activity, brain endothelial penetration and spheroid formation. One of the consequences of melanoma-MG interactions was ALDOC upregulation in melanoma cells. Melanoma cells overexpressing ALDOC demonstrated enhanced migration, adhesion to brain endothelial cells, brain endothelial penetration, viability and spontaneous brain micrometastasis formation. Soluble factors from ALDOC overexpressing melanoma cells increased MG proliferation and migration, which may induce MG involvement in the metastatic process and act as a positive feedback loop, further enhancing MBM progression. Inter-tumor heterogeneity was demonstrated with respect to the effect of ALDOC on melanoma malignancy.

Conclusion: Melanoma and MG cells interact in a bi-directional, reciprocal manner. Such interactions may promote melanoma progression by several mechanisms.