PROJECT SUMMARY (Characterization Unit) Metastasis involves the complex process of spreading of cancer cells beyond the primary tumor site, and is the cause of the majority of cancer deaths. The complexity arises from a dynamic interplay between heterogeneous and evolving cancer cells and the tumor microenvironment, during the progression from locally invasive to metastatic cancer. Understanding this progression, which is closely linked to immunity and development, calls for characterization of the various cell states and their interaction with the local niche during the progression. However, constructing such an atlas of the cellular states poses challenges due to the lack of a priori knowledge of all cell types present, the range of cell states to expect, or the rates of transition. To overcome these challenges we propose three iterative aims. First, we will establish a robust, cost-effective, high quality pipeline for single-cell genomics (primarily RNA) characterization of clinical samples provided by the Biospecimen Unit. Through this effort we will build a taxonomy of cell types, cell states and temporal lineage tracing connecting clones from the primary tumor to metastasis. Next, we will establish a robust, cost- effective, high quality pipeline for spatial mapping at single-cell resolution of clinical samples provided by the Biospecimen Unit with the goal systematically mapping the spatial structure of tumors/metastasis and their associated microenvironment. Finally, we will evaluate and assimilate the best new emerging technologies as these reach production level capacity. This is important to do as the field of single-cell technologies is moving forward at a staggering pace, thus poising the characterization unit will remain up to date on these technologies.