Inhibition of extracellular vesicle-derived miR-146a-5p decreases progression of melanoma brain metastasis via Notch pathway dysregulation in astrocytes.

Melanoma has the highest propensity of all cancers to metastasize to the brain with a large percentage of late-stage patients developing metastases in the central nervous system (CNS). It is well known that metastasis establishment, cell survival, and progression are affected by tumour-host cell interactions where changes in the host cellular compartments likely play an important role. In this context, miRNAs transferred by tumour derived extracellular vesicles (EVs) have previously been shown to create a favourable tumour microenvironment. Here, we show that miR-146a-5p is highly expressed in human melanoma brain metastasis (MBM) EVs, both in MBM cell lines as well as in biopsies, thereby modulating the brain metastatic niche. Mechanistically, miR-146a-5p was transferred to astrocytes via EV delivery and inhibited NUMB in the Notch signalling pathway. This resulted in activation of tumour-promoting cytokines (IL-6, IL-8, MCP-1 and CXCL1). Brain metastases were significantly reduced following miR-146a-5p knockdown. Corroborating these findings, miR-146a-5p inhibition led to a reduction of IL-6, IL-8, MCP-1 and CXCL1 in astrocytes. Following molecular docking analysis, deserpidine was identified as a functional miR-146a-5p inhibitor, both in vitro and in vivo. Our results highlight the pro-metastatic function of miR-146a-5p in EVs and identifies deserpidine for targeted adjuvant treatment.

A multi-omics integrative approach unravels novel genes and pathways associated with senescence escape after targeted therapy in NRAS mutant melanoma.

Therapy Induced Senescence (TIS) leads to sustained growth arrest of cancer cells. The associated cytostasis has been shown to be reversible and cells escaping senescence further enhance the aggressiveness of cancers. Chemicals specifically targeting senescent cells, so-called senolytics, constitute a promising avenue for improved cancer treatment in combination with targeted therapies. Understanding how cancer cells evade senescence is needed to optimise the clinical benefits of this therapeutic approach. Here we characterised the response of three different NRAS mutant melanoma cell lines to a combination of CDK4/6 and MEK inhibitors over 33 days. Transcriptomic data show that all cell lines trigger a senescence programme coupled with strong induction of interferons. Kinome profiling revealed the activation of Receptor Tyrosine Kinases (RTKs) and enriched downstream signaling of neurotrophin, ErbB and insulin pathways. Characterisation of the miRNA interactome associates miR-211-5p with resistant phenotypes. Finally, iCell-based integration of bulk and single-cell RNA-seq data identifies biological processes perturbed during senescence and predicts 90 new genes involved in its escape. Overall, our data associate insulin signaling with persistence of a senescent phenotype and suggest a new role for interferon gamma in senescence escape through the induction of EMT and the activation of ERK5 signaling.

miRNA-132-5p mediates a negative feedback regulation of IL-8 secretion through S100A8/A9 downregulation in neutrophil-like HL-60 cells.

Background: Neutrophils are an important source of pro-inflammatory and immunomodulatory cytokines. This makes neutrophils efficient drivers of interactions with immune and non-immune cells to maintain homeostasis and modulate the inflammatory process by notably regulating the release of cytokines. Ca2+-dependent regulatory mechanism encompassing cytokine secretion by neutrophils are not still identified. In this context, we propose to define new insights on the role of Ca2+-binding proteins S100A8/A9 and on the regulatory role of miRNA-132-5p, which was identified as a regulator of S100A8/A9 expression, on IL-8 secretion. Methods: Differentiated HL-60 cells, a human promyelocytic leukemia cell line that can be induced to differentiate into neutrophil-like cells, were used as a model of human neutrophils and treated with N- formyl-methionyl-leucyl-phenylalanine (fMLF), a bacterial peptide that activates neutrophils. shRNA knockdown was used to define the role of selected targets (S100A8/A9 and miRNA-132-5p) on IL-8 secretion. Results and discussion: Different types of cytokines engage different signaling pathways in the secretion process. IL-8 release is tightly regulated by Ca2+ binding proteins S100A8/A9. miRNA-132-5p is up-regulated over time upon fMLF stimulation and decreases S100A8/A9 expression and IL-8 secretion. Conclusion: These findings reveal a novel regulatory loop involving S100A8/A9 and miRNA-132-5p that modulates IL-8 secretion by neutrophils in inflammatory conditions. This loop could be a potential target for therapeutic intervention in inflammatory diseases.