Introduction: We previously developed a murine triple knockout (Trp53-/-, Pten-/-, and Rb1-/-, TKO) organoids that are transplantable into immunocompetent C57 BL/6J mice. We sought to characterize the treatment response of TKO tumor metastases to anti-PD1 immunotherapy. Methods: The TKO cells were labeled with a lentiviral luciferase and GFP double-expressing reporter (pFUGW-Pol2-ffLuc2-eGFP, Addgene #71394). The TKO cells were injected into the tail veins of C57 BL/6J mice. Tumor bioluminescence (In Vivo Imaging System Imaging, Perkin Elmer) was quantified by integrating the photonic flux (photons/second) through regions encircling each tumor. Upon tumor detection, the mice were randomly treated with an anti-PD-1 (BioXcell, RMP1-14, 200 µg, intraperitoneally twice weekly) (n = 8) or a control IgG2a (BioXcell, 2A3) (n = 6). The mice were monitored for overall survival, serial imaging, and analyses of tumor metastasis at the endpoint. Results: All mice developed lung and/or bone metastases (n = 14). Kaplan-Meier analysis showed no difference in median survival between the control group and anti-PD-1 group (14.5 days vs 17.5 days, p = 0.54). There was no difference in tumor bioluminescence between the control group and anti-PD-1 groups (median p/s 6.94E8 vs 4.32E8, p = 0.85). Histomorphological features and immunohistochemical profile (positive for CK7, p63, GATA3) of lung and bone metastases revealed high-grade urothelial carcinoma. Conclusions: The TKO metastatic model represents an ideal experimental model for studying tumor cell dissemination and treatment resistance to immunotherapy. SOURCE OF Funding: This research work was supported in part by NIH Grants, K08CA252161(Li, Q), P30CA016056 (NCI Cancer Center Core Support Grant), Roswell Park Alliance Foundation, and The Friends of Urology Foundation. We acknowledge the shared instrumentation (S10) grant (S10OD16450).
