Treatment of various types of cells with the mitochondrial ATP-sensitive K(+) channel opener, diazoxide, preconditions cells to subsequent injuries and inhibits apoptosis. However, the role and mechanism(s) of diazoxide in solid tumor cell growth are largely unknown. Here we demonstrate that diazoxide inhibited the proliferation of lung cancer cells as well as the transcription of cell cycle-related protein Cyclin D1. Cyclin D1 overexpression inhibited the negative role of diazoxide in cell cycle progression. We further explored the mechanisms by which diazoxide affected Cyclin D1 transcription and found that the beta-catenin transcription factor was downregulated by diazoxide, with a corresponding decrease in Cyclin D1 promoter activity. Taken together, these results suggest that diazoxide inhibits lung cancer cell proliferation via downregulation of Cyclin D1 transcription, which may have important therapeutic implications in lung cancer patients.