A tissue-engineered model of the blood-tumor barrier during metastatic breast cancer.

Metastatic brain cancer has poor prognosis due to challenges in both detection and treatment. One contributor to poor prognosis is the blood-brain barrier (BBB), which severely limits the transport of therapeutic agents to intracranial tumors. During the development of brain metastases from primary breast cancer, the BBB is modified and is termed the 'blood-tumor barrier' (BTB). A better understanding of the differences between the BBB and BTB across cancer types and stages may assist in identifying new therapeutic targets. Here, we utilize a tissue-engineered microvessel model with induced pluripotent stem cell (iPSC)-derived brain microvascular endothelial-like cells (iBMECs) and surrounded by human breast metastatic cancer spheroids with brain tropism. We directly compare BBB and BTB in vitro microvessels to unravel both physical and chemical interactions occurring during perivascular cancer growth. We determine the dynamics of vascular co-option by cancer cells, modes of vascular degeneration, and quantify the endothelial barrier to antibody transport. Additionally, using bulk RNA sequencing, ELISA of microvessel perfusates, and related functional assays, we probe early brain endothelial changes in the presence of cancer cells. We find that immune cell adhesion and endothelial turnover are elevated within the metastatic BTB, and that macrophages exert a unique influence on BTB identity. Our model provides a novel three-dimensional system to study mechanisms of cancer-vascular-immune interactions and drug delivery occurring within the BTB.

Three-dimensional induced pluripotent stem-cell models of human brain angiogenesis.

During brain development, chemical cues released by developing neurons, cellular signaling with pericytes, and mechanical cues within the brain extracellular matrix (ECM) promote angiogenesis of brain microvascular endothelial cells (BMECs). Angiogenesis is also associated with diseases of the brain due to pathological chemical, cellular, and mechanical signaling. Existing in vitro and in vivo models of brain angiogenesis have key limitations. Here, we develop a high-throughput in vitro blood-brain barrier (BBB) bead assay of brain angiogenesis utilizing 150 µm diameter beads coated with induced pluripotent stem-cell (iPSC)-derived human BMECs (dhBMECs). After embedding the beads within a 3D matrix, we introduce various chemical cues and extracellular matrix components to explore their effects on angiogenic behavior. Based on the results from the bead assay, we generate a multi-scale model of the human cerebrovasculature within perfusable three-dimensional tissue-engineered blood-brain barrier microvessels. A sprouting phenotype is optimized in confluent monolayers of dhBMECs using chemical treatment with vascular endothelial growth factor (VEGF) and wnt ligands, and the inclusion of pro-angiogenic ECM components. As a proof-of-principle that the bead angiogenesis assay can be applied to study pathological angiogenesis, we show that oxidative stress can exert concentration-dependent effects on angiogenesis. Finally, we demonstrate the formation of a hierarchical microvascular model of the human blood-brain barrier displaying key structural hallmarks. We develop two in vitro models of brain angiogenesis: the BBB bead assay and the tissue-engineered BBB microvessel model. These platforms provide a tool kit for studies of physiological and pathological brain angiogenesis, with key advantages over existing two-dimensional models.