Compromised Blood-Brain Barrier Junctions Enhance Melanoma Cell Intercalation and Extravasation

SIMPLE SUMMARY: The worst outcome of melanoma is the formation of melanoma-brain metastasis, which depends on the successful extravasation of metastatic melanoma cells across the tight blood-brain barrier (BBB). Therefore, a detailed understanding of the role of the BBB barrier properties in melanoma cell extravasation is important for preventing brain metastasis formation. In this study, we use in vitro live cell imaging to show that melanoma cells exclusively use the junctional pathway for intercalation into the BBB. By using a broad-spectrum protease inhibitor in an experiment analysing barrier disruption by melanoma cells, we confirm the role of proteases in the process of intercalation of melanoma cells into the BBB in vitro. These observations underscore the role of the BBB junctions in the process of melanoma-brain metastasis formation. Finally, using two different in vitro model systems and one in vivo mouse model, we showed that compromised BBB barrier properties facilitate melanoma cell extravasation. Taken together, our data suggest that preserving BBB integrity is an important measure to limit the formation of melanoma-brain metastasis. ABSTRACT: Melanoma frequently metastasises to the brain, and a detailed understanding of the molecular and cellular mechanisms underlying melanoma cell extravasation across the blood-brain barrier (BBB) is important for preventing brain metastasis formation. Making use of primary mouse brain microvascular endothelial cells (pMBMECs) as an in vitro BBB model, we imaged the interaction of melanoma cells into pMBMEC monolayers. We observed exclusive junctional intercalation of melanoma cells and confirmed that melanoma-induced pMBMEC barrier disruption can be rescued by protease inhibition. Interleukin (IL)-1β stimulated pMBMECs or PECAM-1-knockout (-ko) pMBMECs were employed to model compromised BBB barrier properties in vitro and to determine increased melanoma cell intercalation compared to pMBMECs with intact junctions. The newly generated brain-homing melanoma cell line YUMM1.1-BrM4 was used to reveal increased in vivo extravasation of melanoma cells across the BBB of barrier-compromised PECAM-1-deficient mice compared to controls. Taken together, our data indicate that preserving BBB integrity is an important measure to limit the formation of melanoma-brain metastasis.