In this work, we developed imidazole nucleoside derivatives with anti-dengue virus (DENV) activity was examined. First, compounds in a nucleosides library were screened to find lead compounds which inhibit replication of DENV. As a result, 5-ethynyl-(1-β-d-ribofuranosyl)imidazole-4-carboxamide (1; EICAR) and its 4-carbonitrile derivative EICNR (2) were selected as promising antiviral compounds. However, both of them also exhibited cytotoxicity. In order to develop an effective and less toxic compound, 4'-thio and 4'-seleno derivatives of EICAR and EICNR 3-6 were prepared. The resulting 4'-thioEICAR and 4'-thioEICNR showed inhibitory effect on DENV replication without cytotoxicity as potent as ribavirin, a positive control.