Abstract
While subcutaneous tumor models remain the standard for studying drug efficacy in vivo, these tumors rarely metastasize and lack physiological relevance due to differences in the tumor microenvironment, vascularization, immune landscape, and physiological cues associated with the organ of interest. Orthotopic tumors, grown from the organ corresponding with the cancer type, provide a more translational approach to study disease progression and drug efficacy. Utilization of a syngeneic mouse model allows for a complete immune landscape, key for adaptive immunotherapy studies. MC38 and CT26 cells are commonly used murine colorectal cancer cell lines with clinically relevant mutations. While CT26 cells have been orthotopically implanted with high fidelity, successful engraftment of orthotopic MC38 tumors varies greatly between studies. Thus, we have developed a detailed protocol for MC38 orthotopic tumor inoculation via intracecal injection. Nine C57BL/6 mice were injected with 2 × 10(6) cells into the cecal wall and sacrificed after 7 weeks. Survival after surgery was 100%, and one mouse died before the 7-week study end point from tumor burden and metastatic spread. We observed a successful tumor engraftment rate of 67%. Half of mice presenting with tumors were found to have macroscopic metastatic lesions in clinically relevant foci, including the mesenteric lymph nodes, liver, and peritoneum. These mice also presented with very large tumors and an enlarged spleen. The other half of the mice presented with small, localized tumors that did not metastasize. Herein, we describe tips specific for the intracecal injection of MC38 cells to improve the engraftment rate consistency in this model.