Some novel fused heterocyclic compounds of 2,5-disubstituted-benzoxazole derivatives, which were previously synthesized by our group, were investigated for their mutagenic properties on Salmonella typhimurium TA 98 and TA 100 strains, cytotoxic activity in L929 and HeLa cell lines by Sulforhodamine B (SRB) cytotoxicity test, and genotoxic potentials in the comet assay. By using Ames/Salmonella assay in the presence of S9 fraction, B22 (5-nitro-2-(p-nitrobenzyl) benzoxazole) was found to be mutagenic in both S. typhimurium TA98 and TA100 strains at all tested doses. IC50 values which were evaluated by SRB cytotoxicity assay revealed that B11 (2-(p-nitrobenzyl) benzoxazole) (IC50 = 99.16 mu M) was the most anti-proliferative compound on HeLa cancer cells. Compounds were also tested for their genotoxicity by using comet assay, and it was found that all the compounds had DNA-damaging genotoxic activity on HeLa cells. The comet assay results showed that B11 produced DNA damage at lower concentrations than the other compounds tested on HeLa cancer cells. The results obtained from all the tests suggest that B11 could be a good candidate as a new anticancer agent.