Abstract 4409: Assessment of sonic hedgehog pathway inhibition in a novel orthotopic xenograft bladder cancer murine model

Abstract

Abstract Purpose of the study: To develop a reliable and reproducible bladder cancer murine model for the evaluation of the SHH pathway. Introduction: The sonic hedgehog (SHH) signaling pathway has been shown to play an integral role in the maintenance and progression of bladder cancer. Smoothened inhibitors are currently used in the clinic for treatment of some skin cancers, however they have failed to be effective in bladder cancers in vitro. Blocking alternative downstream molecules of the SHH pathway may be an efficacious strategy for bladder cancer treatment. In vitro assays possess a number of limitations such as environmental differences, loss of heterogenicity and vascularization and artificial levels of growth factors and cytokines within the cell culture media. To overcome this animal models are needed to facilitate the study of carcinogenesis mechanisms. A new bladder cancer model is now required to evaluate experimental therapeutics as previous animal models have utilized the mistakenly identified cervical cancer cell line KU7. Methods: A novel orthotopic murine bladder cancer model was developed to assess the role of the SHH signalling pathway in tumor growth and disease progression. The model comprises the instillation of luciferase-expressing UM-UC3 human transitional cell carcinoma cells into the bladder of athymic nude mice following a poly-L-lysine irrigation of the bladder. Tumors are measured by bioluminescent imaging after intra-peritoneal injection of luciferin. Antisense oligonucleotides (ASO) targeted to the SHH pathway transcription factors, Gli family zinc fingers 1 and 2 (Gli1, Gli2), were assessed in this model. Immunohistochemistry (IHC) of proteins in the SHH pathway were evaluated and compared to our in-house tissue microarray (TMA) of clinical bladder cancer samples. Results: Tumors in this model grew in an average period of time of 40 days with high rates of engraftment and reproducibility. In vitro analysis of SHH pathway inhibition showed a decrease in bladder cancer cell viability and increase in apoptosis following Gli ASO treatment. This effect bypasses smoothened protein regulation which was found to be ineffective in some cell lines. Gli ASO treatment reduced tumor size, and prevented further growth, of UM-UC3 bladder cancers in this mouse model. IHC confirmed Gli2 knock-down and showed increased apoptosis and decreased proliferation in the in vivo tumors. Comparison of modelled tumors to clinical samples showed an increased and diffused Gli2 staining throughout the tissue in both cases. Conclusions: A murine intravesical model consisting of human tumors provides a robust and effective in vivo system for the assessment of cancer inhibiting treatments. Gli2 ASO proved to be a promising treatment for bladder cancer by inhibiting tumor growth and disease progression. Citation Format: Peter A. Raven, Sebastian Frees, Betty Zhou, Claudia Chavez-Munoz, Michael Cox, Alan So. Assessment of sonic hedgehog pathway inhibition in a novel orthotopic xenograft bladder cancer murine model. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4409.