Trimethoxyl stilbene (TMS) is a derivative of resveratrol, a compound shown to inhibit development of a variety of tumor types. We aimed to evaluate the effect of TMS on cell proliferation and apoptosis in the A549 non-small cell lung cancer cell line. Growth inhibition rate and colony formation was measured and apoptosis was determined with Hoechst 33258 staining. Protein expression levels of caspase-3, STAT3, STAT5b, JAK2, NF-κB, and IκB were examined by Western blotting. Furthermore, localization of NF-κB protein was also explored. TMS inhibited proliferation (IC50 8.6 μmol/L) and induced apoptosis of the cells in a concentration-dependent manner., also inducing apoptosis accompanied by up-regulated expression and cleavage activation of caspase-3, up-regulation of IκB and down-regulation of NFκB, STAT3, STAT5b, and JAK2 signal transduction. TMS has potential as a new drug for treatment of non-small cell lung cancer patients with anti-proliferation and apoptosis inducing effect of TMS to A549 cells apparently related to its inhibitory effect on STATs and NF-κB signal transduction. Up-regulation of caspase-3 further supports the potential clinical use of TMS for the treatment of non-small cell lung adenocarcinoma.