Abstract 3915: An orthotopic model for murine urothelial carcinoma with response to standard of care checkpoint inhibitor treatment
Authors: Elizabeth Rainbolt, Andrew Wong, Nisha Ansari, Isaac Mick, Evelyn Dubis, Joette Crews, Jessica Neil, Patrick Fadden
Journal: Cancer Research (2025)

Abstract MB49 is a commercially available cell line used to evaluate bladder cancer for syngeneic growth and potential therapeutic response in preclinical studies. Subcutaneous implantation provides a convenient and accessible location to monitor tumor growth and response to therapy; however, an orthotopic approach may provide a more clinically relevant model. Bladder cancer is the fourth most common cancer in men and current research is focusing on immunotherapy to reduce toxicity while improving efficacy. Current orthotopic bladder implant models include time consuming intralumenal installation methods, however we will focus on a faster more direct implant approach in the intramural space (for implant characterization and evaluation see Bitaraf 2022). To establish the orthotopic model, we implanted cells directly into the bladder wall or intramural space in syngeneic C57BL/6 mice. Two cell inoculums were evaluated: 2.5x105 based on subcutaneous implant standards and 1x105 after initial testing proved rapid tumor progression. Individual animals were evaluated for disease progression based on body weight and bladder palpation performed at regular intervals with the frequency increased as needed based on clinical observations. If enlarged, bladders were expressed to confirm bladder function and urine color. Humane endpoint was met if the bladder was unable to be expressed or if pigmenturia was present. Necropsies were performed to evaluate bladders for tumor and any abnormal observations were recorded. Both cell inoculums had fast growth with median time to endpoint at nine to twelve days post implant. The lower cell inoculum (1x105) was chosen to proceed with efficacy data. Responses to the checkpoint inhibitors anti-CTLA-4 and anti-PD-1 RMP1-14 are described in the poster figures and tables. For comparison, the anti-CTLA-4 and anti-PD-1 response data is provided for the MB49 subcutaneous model. In summary, the development of the MB49 bladder orthotopic model will provide a sensible alternative that recapitulates the difficulties of treating bladder tumors in addition to establishing a more clinically relevant model to evaluate cancer therapies compared to standard subcutaneous implanted models. Future testing with a luciferase labeled cell line will provide imaging capabilities to monitor tumor growth via in vivo bioluminescence imaging (BLI). Bitaraf , M., Muhammadnejad, S., Azimzadeh, A., Tanourlouee, S.B., Amini, E., Zolbin, M.M., Kajbafzadeh, A.M. Evaluation of direct intramural injection to the bladder wall as a method for developing orthotopic tumor models, Animal Model Exp Med (2022) Nov30; 5(6): 575-581. Citation Format: Elizabeth Rainbolt, Andrew Wong, Nisha Ansari, Isaac Mick, Evelyn Dubis, Joette Crews, Jessica Neil, Patrick Fadden. An orthotopic model for murine urothelial carcinoma with response to standard of care checkpoint inhibitor treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3915.