(A) NOTCH1 protein expression in 13 CCA cell lines treated with two different GSi (RO4929097 and YO-01027) for 3- and 7-days. GSi were tested over the dilution series (0.1-10μM). βA: β-actin. N1: NOTCH1. (B) Mice at time of sacrifice (maximum tumor size) after subcutaneous injection of with HuCCT-1 and WITT cell lines, each with and without prior treatment with GSi. (C, D) Effect of GSi pre-treatment on tumor growth initiated by HuCCT-1 (GSi sensitive) (C) and WITT (GSi resistant) cells (D). *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.