EXPLORING THE ROLE OF HYPOXIC EXTRACELLULAR VESICLES IN NEUROBLASTOMA METASTASIS USING AN IN VIVO MODEL AND MICROFLUIDIC CHIPS

Neuroblastoma (NB) is the most common cancer in infants, accounting for 7-10% of childhood cancers. While the overall 5-year survival rate is approximately 80% in Western countries, the prognosis varies by disease stage, with nearly half of NB cases presenting with metastases affecting the bone, bone marrow (BM), liver, and other organs. Metastasis, the leading cause of cancer-related deaths, involves complex processes, including tumor cell dissemination and angiogenesis. Extracellular vesicles (EVs), which are membrane-enclosed particles secreted by cells, play a key role in this process by facilitating both short- and long-range communication between cells at different stages of metastasis. This study explores the role of EVs in NB metastasis using zebrafish embryos and microfluidic models. EVs derived from NB cells grown under hypoxic conditions (hEVs) injected in the zebrafish embryos were found to promote angiogenesis, increase macrophage mobilization, enhance the expression of metastasis-related genes, and increase NB cell proliferation, suggesting their role in metastasis. Additionally, we developed a metastasis-on-a-chip (MOC) model and a simplified metastasis-on-a-chip (sMOC) model to simulate NB metastasis to the BM. Surprisingly, perfusion of hEVs reduced NB cell invasion in these systems, indicating that they may first affect endothelial barrier integrity and inflammatory responses. Gene expression analysis confirmed that hEVs influence pathways related to epithelial-mesenchymal transition (EMT), further supporting their role in metastasis. These findings highlight the potential of hEVs in shaping the metastatic niche and provide new insights into the mechanisms of NB progression. The combination of in vivo and microfluidic chips offers a valuable platform for studying NB metastasis and testing therapeutic interventions targeting the metastatic process.