Colorectal cancer (CRC) is a leading cause of cancer death worldwide and 5-year survival for metastatic CRC is only ~15%. Immunotherapy is an important new approach to treat solid tumors. However, to date it appears largely ineffective against CRC, and the immune mechanisms underlying human CRC growth and metastasis are poorly understood. In part, this is because human CRC xenograft models could only be generated in immunodeficient mouse hosts because of xeno-rejection. We recently developed chemokine targeted mouse models (CTMM), a novel experimental approach to robustly model primary CRCs in the native GI micro- environment without requiring surgery. By using the Chemokine Receptor 9/ Chemokine 25 axis, CTMM traffics human CRC cells to form orthotopic tumors in the GI tract. Here, we describe an innovative combination of mouse ES cell technology and CTMM to generate human CRC-mouse chimeras, the first robust model of human orthotopic CRC in immunocompetent mouse hosts for mechanistic studies to address underlying tumor immunology and potential therapeutic targets in vivo. The overall goal of this application is to understand mechanisms of interaction between human CRC cells, tumor infiltrating immune cells, and the intestinal microbiota to promote orthotopic tumor growth in immunocompetent hosts.