The clone counts of DCLK1 overexpressing HCCC9810 group was 61.33 ± 5.79, which was significantly higher than that of HCCC9810 control cells (23.33 ± 3.86, t = 7.72, P = 0.002) and DCLK1 overexpressing cells treated with LY294002 (5.33 ± 1.25, t = 13.37, P < 0.001).The colony formation assay also confirmed that the PI3K/AKT/mTOR pathway inhibitor could significantly reduce the proliferation of CCA cells mediated by high expression of DCLK1 (Fig. 5B). This evidence concerns the gene MTOR and cholangiocarcinoma.