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Inhibition of Gli1 and Gli2 as a targeted therapy for bladder cancer

University of British Columbia

The sonic hedgehog (SHH) signaling pathway has been shown to play an integral role in the maintenance and progression of bladder cancer (BCa). Smoothened inhibitors are currently used in the clinic for treatment of some skin cancers, however they have not been evaluated in BCa and SHH inhibition may be an efficacious strategy for BCa treatment. I assessed an in-house human BCa tissue microarray comprising non-invasive, invasive and lymph node metastasized transitional cell carcinoma and found that the transcription factors downstream of SHH, Gli1 and Gli2, were increased in more aggressive tumors. A panel of BCa cell lines show that two invasive lines, UM-UC-3 and 253J-BV, both express these transcription factors but differ in other parameters in the SHH pathway. UM-UC-3 produces greater quantities of SHH ligand, is less responsive in viability to pathway stimulation by recombinant human SHH or SAG, and less responsive to inhibition by a variety of molecules including the Smoothened inhibitors cyclopamine and SANT-1. 253J-BV, on the other hand, was highly responsive to these manipulations and appears more representative of canonical SHH signaling while UM-UC-3 resembles non-canonical autocrine signaling. To overcome this variability I utilized a Gli1 and Gli2 antisense oligonucleotide (ASO) to bypass pathway mechanics and target the transcription factors directly. UM-UC-3 decreased in viability due to both ASOs but 253J-BV was only affected by Gli2 ASO. IC50s were in the nanomolar range. To evaluate in vivo efficacy I developed a murine intravesical orthotopic human bladder cancer (mio-hBC) model for the establishment of non-invasive urothelial cell carcinomas. In this model I pre-treat the bladder with poly-L-lysine for 15 minutes, followed by intravesical instillation of luciferase–transfected human UM-UC-3 cells. Cancer cells are quantified by bioluminescent imaging. Tumors grew to 541.6±0.75 fold (Mean±SE) initial size after 40 days and were confirmed to reflect patient samples by a response to mitomycin C. Treatment of these tumors with Gli2 ASO resulted in decreased tumor size, growth rate and Gli2 mRNA and protein expression. These results validate this model and support the conclusion that Gli2 ASO may be a promising new targeted therapy for BCa.

Text

2019-06-30

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/66407

Experimental Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/