Studies have highlighted molecular and cell biological mechanisms of the NKG2D-NKG2D ligand system in the activation of natural killer (NK) cell and T cell functions. In this study we explore the potential of genetic overexpression of human major histocompatibility complex class I chain-related gene A (MICA), a powerful NKG2D ligand, for the induction of NK cell-mediated antitumor immunity in a humanized murine model of non-small cell lung cancer. Using tissue microarray technology we detected expression of MICA in only 30% of the samples from patients with lung cancer. Staining was always weak and focal, indicating that expression of MICA is detectable but limited in human lung cancer. Genetic overexpression of MICA by means of adenoviral transduction or transfection of expression plasmids was feasible in cell lines in vitro, primary human cancer tissue ex vivo, and in experimental human cancers in vivo. The presence of MICA on the surface of largely NK cell-resistant human lung cancer cells reestablished NK cell susceptibility and provoked NK cell-mediated antitumor immunity by murine and human NK cells in two different experimental therapy models. In this study we analyze the interaction of human NK cells with MICA-positive human cancer cells in an in vivo setting. Our data demonstrate that MICA overexpression can function as NK cell-mediated immunotherapy in experimental lung cancer.