Three series of new benzimidazole hybrids were designed and synthesized as promising human TopoII inhibitors. They were characterized by different spectroscopic techniques (H-1, C-13 NMR, ESI-MS and IR). All hybrids (<bold>6</bold>-<bold>23</bold>) were screened for their in vitro antiproliferative activity against five human cancer cell lines namely; HepG-2, MCF-7, PC-3, HCT-116 and Hela. Compound <bold>21</bold> showed the most potent anticancer activity against all cancer cell lines, with IC50 range of 2.82 to 12.59 mu M, while proving safe towards normal cells WI-38 (IC50 = 31.89 mu M) compared to the reference drug doxorubicin (IC50 = 6.72 mu M). The most active candidates <bold>13</bold>, <bold>20</bold>, <bold>21</bold>, <bold>22</bold> and <bold>23</bold> were further assessed for their human TopoII inhibition. The best of which, compounds <bold>13</bold> and <bold>20</bold> showed IC50 of 6.72 and 8.18 mu M respectively compared to staurosporine (IC50 = 4.64 mu M). Further mechanistic studies for compound <bold>13</bold> showed cell cycle arrest at S-phase by 51.29 % and a significant increase in the total apoptosis by 62.5 folds. Furthermore, apoptosis study proved that it induced apoptosis by decreasing both IAP and Bcl-2, activating caspases 3, 8 and 9, and increasing accumulation of ROS in HepG-2 cells. Besides, it decreased transcription factors' binding activity to DNA. Comparative molecular docking study was performed between the most potent TopoII inhibitors <bold>13</bold> and <bold>20</bold>, and the least potent one <bold>23</bold> to relate the binding pattern with TopoII catalytic active site to the biological activity, where all results came in agreement with the biological results. Additional molecular modeling studies including surface mapping and contact preferences were performed to emphasize the importance of hydrophobicity. Physicochemical calculations were assessed where compounds <bold>13</bold> and <bold>20</bold> represented very promising orally active drug candidates.