Standard chemotherapy and personalized target therapies are commonly used in patients with advanced non-small cell lung cancer (NSCLC). However, multidrug resistance (MDR) and tumor metastasis lead to the decline of therapeutic efficacy, which are closely related to epithelial-mesenchymal transition (EMT). Twist1, an EMT transcription factor, plays an essential role in promoting EMT, MDR and tumor metastasis. In view of the essential role of Twist1 in the tumorigenesis of NSCLC, developing antitumor small molecules that can suppress the expression of Twist1 is of far-reaching significance for the treatment of NSCLC. A series of novel benzo[d]imidazo[2,1-b]thiazole derivatives possessing 1,3,4-oxadiazole moiety were designed based on the structure of the first-in-class Twist1 inhibitor harmine. Among the synthetic twenty-two compounds, the compound containing 2-(piperidine-1-yl) ethyl exhibited remarkable anti-proliferative activity with IC<sub>50</sub> value of 2.03 μM and 9.80 μM against A549 and H2228 cell lines superior to harmine (IC<sub>50</sub> = 17.12 μM against A549, IC<sub>50</sub> = 31.06 μM against H2228). Meanwhile, western blot assay showed that the optimal compound significantly down-regulated Twist1 protein expression in a dose-dependent manner and reduced Twist1 level better than harmine. Collectively, the promising compound was identified a potential antineoplastic lead with the ability of down-regulating Twist1 level.