Lung adenocarcinoma is the most common type of lung cancer and found in both smokers and non-smokers, but the treatment of lung cancer is limited. ITR-284 has been shown to be a potent carboxamide-derived anticancer agent and to induce apoptosis in leukemia and colon cancer cells. However, little is known whether ITR-284 has anticancer activity in human lung adenocarcinoma cells through induction of apoptosis and suppression of migration in vitro. We showed that ITR-284 inhibited human lung cancer A549 cells using the thiazolyl blue tetrazolium bromide (MTT) assay and evoked apoptosis via the cell cycle distribution at S phase arrest. After treatment with 20 nM ITR-284 for 24 h, apoptotic cells were induced and detected by Annexin V-FITC/PI staining. The production of reactive oxygen species (ROS) was dose-dependently increased in A549 cells caused by ITR-284. The results from immunoblotting analysis showed an elevation of protein levels of p53 and phosphorylation of p53 in A549 cells prior to ITR-284 exposure. Additionally, apoptosis-associated proteins such as Bax, cleaved caspase-3 and cleaved PARP were upregulated after ITR-284 treatment. By wound healing assay, low concentrations (1-5 nM) of ITR-284 exerted a greater effect on inhibition of A549 cell migration. The protein levels of E-cadherin and vimentin, which are the epithelial-mesenchymal transition (EMT) markers, were modulated in ITR-284-treated cells assessed by western blot analysis. Taken together, our data suggest that ITR-284 may be an effective anticancer agent for treating lung adenocarcinoma.