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Extracellular vesicles (EVs) are cell-derived nanostructures that mediate intercellular communication by delivering complex signals in normal tissues and cancer. The cellular coordination required for tumor development and maintenance is mediated, in part, through EV transport of molecular cargo to resident and distant cells. Most studies on EV-mediated signaling have been performed in two-dimensional (2D) monolayer cell cultures, largely because of their simplicity and high-throughput screening capacity. Three-dimensional (3D) cell cultures can be used to study cell-to-cell and cell-to-matrix interactions, enabling the study of EV-mediated cellular communication. 3D cultures may best model the role of EVs in formation of the tumor microenvironment (TME) and cancer cell-stromal interactions that sustain tumor growth. In this review, we discuss EV biology in 3D culture correlates of the TME. This includes EV communication between cell types of the TME, differences in EV biogenesis and signaling associated with differing scaffold choices and in scaffold-free 3D cultures and cultivation of the premetastatic niche. An understanding of EV biogenesis and signaling within a 3D TME will improve culture correlates of oncogenesis, enable molecular control of the TME and aid development of drug delivery tools based on EV-mediated signaling.

Visualizing Extracellular Vesicles and Their Function in 3D Tumor Microenvironment Models