Cancer invasion involves the growth of the tumor and the invasion of cancer cells. During this process, the tumor typically elongates protrusion-like structures, which triggers the invasion of cells into the blood vessel, and thereby, initiate metastasis. Recent evidences using individual cells have identified the key role of membrane protrusion fluctuations in the physicochemical mechanism of directed cell motion]. The exact roles of protrusion fluctuations during tumor growth and invasion have yet not been reported. Different in vitro and in vivo models have been developed to monitor cancer invasion and its mechanistic determinants. These models display certain limitations, which threaten the relevancy of the obtained data. In contrast, organs-on-chip devices can replicate all the key cellular, structural, and rheological properties of solid tumors, contributing to assess the mechanism at work of the disease. In this work, an organ-on-chip approach is used to analyze the important role of protrusion fluctuations in the invasiveness capability of lung tumor cells. To this aim, we have developed a tumor blood-vessel-on-chip platform containing human endothelial cells to mimic the blood vessel, and lung tumor m-organoids embedded into a 3D collagen matrix
to mimic the tumor microenvironment. We found that lung tumor m-organoids extended protrusions mainly towards the blood vessel channel causing a bias in protrusion distribution. The fluctuating protrusions displayed a continuous elongation and enhanced lifetime, which have been correlated with the growth of the tumor. Finally, the perturbation of protrusion dynamics using typical anti-cancer drugs (doxorubicin) and C3 Rho inhibitor affected the invasion capability of cancer cells. Altogether, the results demonstrate that fluctuations of protrusions are key players in the physicochemical mechanism of tumor invasion, and may have important implications for the development of targeted therapeutic approaches.