One woman in eight is destined to develop breast cancer. All women that lose their battle against breast cancer vividly realize that its most deadly feature is not the primary lesion, but its ability to spread to distant sites and become an incurable, metastatic disease. In this proposal, we aim to address what is arguably the most ambitious challenge in the cancer field: metastasis prevention.<br/><br/> Metastatic spread of cancer occurs through the generation and hematogenous dissemination of circulating tumor cells (CTCs). We recently discovered that CTCs are heterogeneous, and that CTC clusters – multicellular aggregates of cancer cells and immune cells in circulation – represent the most powerful mediators of disease spread (Szczerba et al., Nature, 2019; Gkountela et al., Cell, 2019). Post-intravasation biological and clinical features of CTCs are extensively interrogated, yet, surprisingly, the mechanisms that dictate their generation from a solid tumor lesion are unknown. Only a minimal fraction of cancer cells is capable to intravasate in functional blood vessels, while the vast majority of tumor cells will always remain in situ. Understanding the fundamental biological principles that govern CTC generation is of paramount importance, as it could lead to the development of new therapeutic agents that lock tumor cells in their primary site, thereby preventing metastasis.<br/><br/> Our preliminary data strongly suggest that CTC generation is not occurring in a stochastic fashion, but it is dictated by very precise and previously-unappreciated signals. Based on these very exciting findings, the two predominant goals of this proposal are: first, to gain insights into those fundamental cell-autonomous and non-cell-autonomous mechanisms that govern CTC intravasation in vivo. Second, to identify actionable targets and direct treatment opportunities to lock tumor cells in situ and blunt CTC generation, with the long-term ambition to prevent the metastatic spread of breast cancer.