Recent studies demonstrated that microRNA-145 is downregulated in human cancer cells and may function as a tumor suppressor. However, its role in lung cancer tumorigenesis remains unclear. Here, we demonstrated that upregulation of miR-145 reduced the proliferation and invasion as well as the ratio of CD133-positive initiating cells and the tumorosphere growth capacity of the human lung adenocarcinoma A549 cell line. The direct targeting of miR-145 to OCT4 mRNA was predicted by bioinformatic analysis and validated by a luciferase reporter system. We, therefore, confirmed that miR-145 can impair the proliferation of human lung adenocarcinoma-initiating cells by targeting OCT4 and leads to inhibition of lung cancer development.