New nucleoside derivatives with nitrogen substitution at the C-6 position were prepared and screened initially for their in vitro anticancer bioactivity against human epithelial cancer cells (liver Huh7, colon HCT116, breast MCF7) by the NCI-sulforhodamine B assay. N6-(4-trifluoromethylphenyl)piperazine analog (27) exhibited promising cytotoxic activity. The compound 27 was more cytotoxic (IC50 = 1-4 μM) than 5-FU, fludarabine on Huh7, HCT116 and MCF7 cell lines. The most potent nucleosides (11, 13, 16, 18, 19, 21, 27, 28) were further screened for their cytotoxicity in hepatocellular cancer cell lines. The compound 27 demonstrated the highest cytotoxic activity against Huh7, Mahlavu and FOCUS cells (IC50 = 1, 3 and 1 μM respectively). Physicochemical properties, drug-likeness, and drug score profiles of the molecules showed that they are estimated to be orally bioavailable. The results pointed that the novel derivatives would be potential drug candidates.