Background: Cyclo-oxygenase 2 (Cox-2) is expressed in many types of tumors, but typically undetectable in normal tissues. However, Cox-2 is known to be induced following infection by many microbial agents, which might threaten the tumor selectivity of the Cox-2 promoter in the context of virotherapy or viral gene delivery. Cox-2 expression is regulated in part post-transcriptionally by stimulation or inhibition of mRNA degradation by 3′-untranslated region (3′-UTR) AU-rich elements. In the present study, we investigated the induction of the Cox-2 promoter both in normal and tumor cells after adenovirus infection and explored the utility of AU-rich elements for regaining promoter selectivity. Methods: Nontumor and tumor cells were transfected in vitro and in vivo with plasmids containing the Cox-2 or cytomegalovirus immediate early promoter driving luciferase (with or without 3′-UTR elements) followed by adenoviral infection. Selectivity and activity of the promoters and 3′-UTR elements were analysed by luciferase assay and in-vivo imaging. Results: The Cox-2 promoter was induced in both normal and tumor cells following infection with E1 containing replicative adenoviruses but not in the absence of E1. Utilization of AU-rich elements counteracted promoter induction in vitro and in vivo in nonmalignant cells but not in cancer cells, thus increasing the selectivity of the approach ten-fold without loss of potency. Conclusions: Adenoviral infection induces the Cox-2 promoter in normal and tumor cells, which might compromise specificity of the promoter. Utilization of AU-rich destabilization elements can rescue the tumor selectivity of the promoter. Copyright © 2008 John Wiley & Sons, Ltd.