Leader cell mechanics as primary… (R56CA316340) – Project Grant to Virginia Commonwealth University

Abstract: Metastasis continues to be the primary cause of mortality in breast cancer patients. Despite new therapies and diagnostics, median overall survival has only increased by 1.5 years over the past 3 decades. Over 90% of patients with metastatic disease have circulating tumor cells that invade as clusters, yet most metastatic studies focus on understanding how circulating tumor cells invade as individual cells. Collective invasion or metastasis is a type of cell migration where groups of tumor cells move together in a coordinated fashion. During collective invasion, a few cells at the front (i.e. leader cells) define the leading edge, integrate signals from the extracellular matrix (ECM) environment and send signals to neighboring (follower) cells. Our work demonstrate leader cells must arrive (or polarize) at the front to begin invasion. The tumor microenvironment contains many mechanical cues including matrix architecture and fluid flow gradients, yet how these cues impact invasion nor leader cell response is not fully understood. Our objective is to experimentally test how mechanical ECM cues link to leader cell polarization, and elucidate the leader cell mechanics responsible for sensing, interpreting, and coordinating collective invasion. Our hypothesis is that leader cell biophysics are the primary sensors of mechanical matrix cues, making leader cells the primary drivers of collective invasion. We will answer our hypothesis with the following aims: (1): we will investigate how fluid flow and matrix architecture result in directional collective invasion, mediated by leader cell polarization (2): we will interrogate mechanical response of the collective cluster and the leader cells separately (3): we will elucidate leader cell mechanosensors and associated pathways governing invasion. In our 3D microphysiological models, we will incorporate decellularized matrix (decm) from primary breast cancer tumors which provides an accurate presentation of native ECM composition. For cell source, we will culture primary tumor organoid clusters. We will induce dynamic mechanical cues fluid flow and matrix architecture, and perform live-cell imaging to quantify changes in invasion. Also, we will quantify changes in leader cell mechanical response by measuring matrix deformation rates and traction forces. We will interrogate vinculin focal adhesions as a cell-matrix linker connecting leaders to ECM. Based on our single cell sequencing work, we will probe top differentially expressed marker, desmoplakin, as a potential leader cell mechanosensor to direct invasion. We will perform loss-of-function studies with syngeneic orthotopic breast transplant models. Vertebrate animal models are essential for this project because they enable in vivo investigation of complex interactions of tumor leader cell mechanics with the native tumor microenvironment that closely recapitulate key features of human breast cancer. By investigating collective invasion mechanics, we will contribute new perspectives which can bring new approaches for therapy design. Further, we will identify new mechanical signatures of collectively invading tumor clusters, which can add to more universal biomarkers that can diagnose or predict cluster-based metastasis in patients.

Leader cell mechanics as primary regulators of collective tumor invasion

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