A novel series of fourteen substituted selenadiazoles has been synthesized and the compounds tested for their in vitro antiproliferative and cytotoxic activities. The tests were carried out against leukemia (CCRF-CEM), colon (HT-29), lung (HTB-54), and breast (MCF-7) cancer cells. In order to assess the selectivity of the compounds under investigation the assays were also carried out on two non-tumoral lines - one mammary (184B5) and one bronchial epithelium (BEAS-2B) cell line. Assay-based antiproliferative activity studies revealed that seven derivatives (2a, 2c, 2e, 2f, 2g, 3a, and 3b) exhibited good activity against MCF-7 cells: for instance, 2c and 2f inhibited cell growth with nanomolar GI₅₀ values. Compound 2f had a better antitumoral profile than vinorelbine and paclitaxel, two drugs that are used as first-line treatments in advanced, recurrent, and/or metastatic cancer. In the other cell lines the compounds showed moderate activity or were inactive - with the exception of 2a, which was also found to have antiproliferative activity. Modulation of the cell cycle and apoptotic effects of active compounds were further evaluated in MCF-7 cells. Of these, 6-bromo[1,2,5]selenadiazolo[3,4-b]pyridine (2a) was the most active, with an apoptogenic effect 3.9 times higher than that of camptothecin, which was used as a positive control. Compound 2a also provoked cell cycle arrest with a significant decrease in the G₀/G₁ phase cell population and an increase in S and G₂/M cells, thus suggesting mitotic arrest prior to metaphase.