Mdm2 binds to p53 and promotes its degradation. A class of small molecule Mdm2 inhibitors (MI) was recently discovered that binds to Mdm2 and disrupts Mdm2-p53 binding. The efficacy of MI-43 was tested against two pairs of human lung cancer lines differing in p53 status: adenocarcinoma A549 (p53 wild-type, wt) and H522 (p53-null) and non-small cell lung carcinoma H460 (p53wt) and H1299 (p53-null). MI-43 induced the accumulation of p53 and its downstream target genes, Mdm2, p21, Noxa and Puma only in wt p53-containing cells, indicating that disruption of Mdm2-p53 binding increases p53, which is transcriptionally active. MI-43 preferentially inhibited the growth of wt p53-containing cells in a p53 dependent manner, but was much less effective in p53-null cells. Mechanistically, MI-43 induced G(1) or G(2) arrest at low concentration as result of p21 induction, and apoptosis at high concentration due to Puma/Noxa induction. Importantly, MI-43 is much less toxic to normal fetal lung fibroblast, MRC5 cells. Finally, when used in combination, MI-43 sensitized chemo-resistant A549 cells to etoposide-induced apoptosis. Thus, MI-43 or its analogues could be further developed as a novel class of anticancer drug for lung cancer cells harboring wt p53 as a single agent or in combination with chemo-drugs.