Hec1 (highly expressed in cancer) plays an important role in chromosome segregation by interacting with a subset of checkpoint proteins that survey proper chromosome alignment and bipolar spindle attachment. In order to disrupt mitotic progression of tumor cell lines, we have used retrovirus and adenovirus vectors that inhibit Hec1 synthesis. Vector-expressed short hairpin RNAs (shRNAs) caused very efficient depletion of the target protein, cellular arrest and considerable mitotic catastrophe induction 96 h post infection in human cervix-adenocarcinoma (HeLa) and glioblastoma (U-373-MG) cell lines. Furthermore, adenocarcinomas induced in the flanks of nude mice show significant reduction in size compared with control when treated with either Hec1-shRNA retroviruses or adenoviruses. These results indicate that depletion of Hec1 could be used as a new strategy to block the dividing cell, and therefore against cancer.