Recent studies have shown that genetically engineered stem cells (GESTECs) produce suicide enzymes that convert non-toxic pro-drugs to toxic metabolites which selectively migrate toward tumor sites and reduce tumor growth. In the present study, we evaluated whether these GESTECs are capable of migrating to lung cancer cells and examined the potential therapeutic efficacy of gene-directed enzyme pro-drug therapy against lung cancer cells in vitro. A modified transwell migration assay was performed to determine the migratory capacity of GESTECs to lung cancer cells. GESTECs [i.e., HB1.F3.CD or HB1.F3.CD.interferon-β (IFN-β)] engineered to express a suicide gene, cytosine deaminase (CD), selectively migrated toward lung cancer cells. Treatment of a human non-small cell lung carcinoma cell line (A549, a lung carcinoma derived from human lung epithelial cells) with the pro-drug 5-fluorocytosine (5-FC) in the presence of HB1.F3.CD or HB1.F3.CD.IFN-β cells resulted in the inhibition of lung cancer cell growth. Based on the data presented herein, we suggest that GESTECs expressing CD may have a potent advantage for selective treatment of lung cancers. Furthermore, GESTECs expressing fusion genes (i.e., CD and IFN-β) may have a synergic antitumor effect on lung cancer cells.