Objectives: Lung cancer is the leading cause of cancer-related death in the world. Chemotherapy has a major role to treat patients with non-small cell lung cancer (NSCLC). However, the best regimens can only give an overall response rate of 30-50% and the mechanisms involved in chemotherapy resistance remain unknown. In the present study, the role of hypoxia induced autophagy in acquiring chemoresistance was evaluated in NSCLC. Methods: To confirm the existence of hypoxia in human NSCLC, we evaluate the expression of hypoxia induced factor -1 α (HIF-1 α) on tumor and normal lung tissue. Human lung cancer cell lines(A549) were exposure to 1% oxygen up to 8 hours and control cells were cultured in standard tissue culture conditions. We analyzed autophagic proteins and HIF-1α expression by western blot analysis. Also we analyzed autophagosome by TEM and GFP-LC3 puncta under confocal microscopy. Then, cells were exposed to cisplatin at different concentrations for 8 h under normoxia or hypoxia. We measured the percentage of killed cells by MTT colorimetric assay to confirm the chemoresistance. Results: HIF-1α proteins were predominantly expressed in tumor tissue compared with normal lung tissue by immunohistochemical stains and western blot. Hypoxia increased autophagic protein and HIF-1 alpha expression by Western blots. And hypoxia increased GFP-LC3 puncta and number of autophagosomes in TEM. Under hypoxia, cisplatin induced cell death was decreased. Conclusions: Autophagy induced by hypoxia may play an important role in acquiring chemoresistance in NSCLC.