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In this work, we combined three-dimensional (3D) scaffolds with flow perfusion bioreactors to evaluate the gradient effects of scaffold architecture and mechanical stimulation, respectively, on tumor cell phenotype. As cancer biologists elucidate the relevance of 3D in vitro tumor models within the drug discovery pipeline, it has become more compelling to model the tumor microenvironment and its impact on tumor cells. In particular, permeability gradients within solid tumors are inherently complex and difficult to accurately model in vitro. However, 3D printing can be used to design scaffolds with complex architecture, and flow perfusion can simulate mechanical stimulation within the tumor microenvironment. By modeling these gradients in vitro with 3D printed scaffolds and flow perfusion, we can identify potential diffusional limitations of drug delivery within a tumor. Ewing sarcoma (ES), a pediatric bone tumor, is a …