Endothelial-to-mesenchymal transition drives circulating tumor cell cluster formation in a 3D tumor-vessel model

Circulating tumor cell (CTC) clusters, which are often found in the blood of patients with high-grade tumors, are strongly associated with metastasis and poor prognosis. However, the mechanisms by which cancer cell clusters invade blood vessel barriers remain unclear. In this study, we developed a three-dimensional (3D) in vitro model to visualize tumor intravasation by positioning tumor organoids with distinct genetic backgrounds around engineered microvessels composed of luminal structure of endothelial cells. Our system successfully captured key steps of intravasation, including collective migration toward microvessels, vessel co-option, and the release of CTC clusters—an invasion mechanism not previously reported. Moreover, we identified that elevated transforming growth factor-β (TGF-β) and activin expression in endothelial cells played a critical role in facilitating intravasation, which was associated with endothelial-to-mesenchymal transition (EndoMT). This 3D model provides a valuable platform for developing clinical intervention strategies targeting CTCs and tumor metastasis.