Project Summary/Abstract It is critically important to establish the causes of organ-specific metastasis; without this knowledge, prevention and timely treatment of metastatic cancer progression will likely remain limited. The proposed project aims to identify how metabolic gradients in the primary tumor can maintain diverse lineages with specific metabolic adaptations, and how these adaptations contribute to organ-specific metastasis in breast cancer. The central hypothesis is 1) that metabolic adaptations are key to the match between the seed (the disseminated cell) and the soil (the distal site) in metastatic breast cancer, and 2) that the metabolic microenvironment in a primary tumor drives metabolically diverse subpopulations. The hypothesis has been formulated based on 1) published data detailing metabolic heterogeneity and that metabolic adaptations can promote metastasis, 2) preliminary data and analysis of RNA expression, metabolomics, and flux measurements, revealing different metabolic adaptations in breast tumor cells that home to different tissues, and 3) preliminary data showing that metastatic lineages respond differently to hypoxia and nutrient gradients, indicating a role for the metabolic microenvironment in maintaining diverse subpopulations within the same heterogeneous primary tumor. The proposed aims will be accomplished using a combination of experimental and computational biology. Integrated flux balance models of metastatic lineages will identify perturbed metabolic pathways in each lineage. Knocking down or overexpressing key nodes in these pathways and assessing tropism in vivo will determine the role of metabolic adaptations in organ-specific metastatic selection. The ability of spontaneous nutrient gradients in the primary tumor to affect spatial segregation of lineages with distinct metabolic adaptations will be investigated using custom MEMIC tissue mimetic plates. Mathematical models will be used to study the ecological interactions between cell lines and their microenvironment that lead to coexistence of metabolically distinct pre-metastatic subpopulations in the primary tumor. Joao Xavier will provide excellent mentorship for this project and the candidate’s career goals of leading an independent research program, with a combination of hands-on teaching and freedom for independent discovery. The current project will form the foundation of the candidate’s future independent research. The training plan also includes meetings with a co-sponsor and collaborators, facilitating a comprehensive education and ensuring a high probability of success for the project. The environment at MSKCC provides further resources important for the completion of the proposed project, as well as career development support as the candidate transitions to an independent career.