High throughput screening to assess urothelial cancer lines and their metastatic derivatives for assessing changes in sensitivity in therapeutics.

Abstract

407 Background: Chemotherapy for urothelial cancer (UC) is less effective in patients with large volume metastatic disease, likely due to acquired driver mutations. Driver mutation may lead to differences in sensitivity between primary cell lines and metastatic clones. Quantitative high throughput screening (qHTS) assesses large drug libraries to evaluate in vitro effectiveness. We utilized this modality to compare effectiveness of cell inhibition in two bladder cancer cell lines with their metastatic lines. Methods: We screened T24 and UMUC-3 bladder cancer cell lines against 1,912 oncology-focused drugs using a 48 hr cell proliferation assay with an ATP-based readout (CellTiterGlo), and determined the activity and potency of the compounds in a dose response manner. We compared the results of their activity with known metastatic lines for reach of these primary lines: T24T, SLT3 and FL3 for T24 and LUL2 UMUC3 and UMUC3-luc (LUC3). We identified candidate drugs based on two parameters: 1) more than 70% inhibition at 48 hours and 2) a curve class of -1.1 or -1.2 indicating curve class with good fit (r2 > 0.9). We used a custom capture next generation DNA sequencing chip for exonic regions of 229 known cancer related genes to compare primary and metastatic tumor cell lines. Results: The qHTS compounds, are demonstrated in figure 1a for T24 cell lines and figure 1b for UMUC cell lines. Amongst 1912 drugs tested, only 141 (7.4%) met inclusion criteria for T24. However, only 79 of these drugs (56%) met criteria for all cell lines derived from T24. 160 (8.4%) compounds were found to be active in the UMUC line. Only 32 (20%) of the UMUC3 detected compounds remained active in all UMUC3 derived cell lines. Genomic comparison between cell lines identified several mutations which were found only in the derived cell lines (RAC1 and CDH1 in UMUC3) and (AR, CDK12, GATA1, GUCY1A2 in T24). Conclusions: qHTS of a library of potential therapeutic interventions can produce a list of targets and therapies for bladder cancer. This data reinforces the differences between primary and metastatic tumors as compounds identified active in the primary cell lines were inactive in known metastatic cancer cell lines .