Randomly Distributed K 14 þ Breast Tumor Cells Polarize to the Leading Edge andGuide Collective Migration inResponse toChemical andMechanical Environmental Cues

Collective cell migration is an adaptive, coordinated interactive process involving cell–cell and cell–extracellular matrix (ECM) microenvironmental interactions. A critical aspect of collective migration is the sensing and establishment of directionalmovement. It has beenproposed that a subgroupof cells known as leader cells localize at the front edge of a collectively migrating cluster and are responsible for directing migration. However, it is unknownhowandwhen leader cells arrive at the front edge and what environmental cues dictate leader cell development and behavior. Here, we addressed these questions by combining a microfluidic device design that mimics multiple tumor microenvironmental cues concurrently with biologically relevant primary, heterogeneous tumor cell organoids. Prior to migration, breast tumor leader cells (K14þ) were present throughout a tumor organoid and migrated (polarized) to the leading edge in response to biochemical and biomechanical cues. Impairment of either CXCR4 (biochemical responsive) or the collagen receptor DDR2 (biomechanical responsive) abrogated polarization of leader cells and directed collective migration. This work demonstrates that K14þ leader cells utilize both chemical and mechanical cues from the microenvironment to polarize to the leading edge of collectively migrating tumors. Significance: These findings demonstrate that pre-existing, randomly distributed leader cells within primary tumor organoids use CXCR4 and DDR2 to polarize to the leading edge and direct migration.