Lung cancer is the leading cause of cancer mortality worldwide. Despite advances in anti-cancer therapies such as chemotherapy, radiotherapy and targeted therapies, five-year survival rates remain poor (<15%). Inherent and acquired resistance has been identified as a key factor in reducing the efficacy of current cytotoxic therapies in the management of non-small cell lung cancer (NSCLC). There is growing evidence suggesting that cancer stem cells (CSCs) play a critical role in tumor progression, metastasis and drug resistance. Similar to normal tissue stem cells, CSCs exhibit significant phenotypic and functional heterogeneity. While CSCs have been reported in a wide spectrum of human tumors, the biology of CSCs in NSCLC remain elusive. Current anti-cancer therapies fail to eradicate CSC clones and instead, favor the expansion of the CSC pool and select for resistant CSC clones thereby resulting in treatment resistance and subsequent relapse in these patients. The identification of CSC-specific marker subsets and the targeted therapeutic destruction of CSCs remains a significant challenge. Strategies aimed at efficient targeting of CSCs are becoming increasingly important for monitoring the progress of cancer therapy and for evaluating new therapeutic approaches. This review focuses on the current knowledge of cancer stem cell markers in treatment-resistant lung cancer cells and the signaling cascades activated by these cells to maintain their stem-like properties. Recent progress in CSC-targeted drug development and the current status of novel agents in clinical trials are also reviewed.