SUMMARY We examined the expression of Op18 in 93 lung adenocarcinomas and 10 uninvolved lung samples using quantitative 2D-PAGE analysis with confirmation by mass spectrometry and 2D-Western blot analysis. mRNA was examined expression using oligonucleotide microarrays, and the cellular localization of the Op18 protein was examined using immunohistochemical analysis of tissue microarrays. Three phosphorylated and one unphosphorylated forms of the Op18 protein were identified and found to be overexpressed in lung adenocarcinomas as compared to normal lung. The percentage of phosphorylated to total Op18 protein isoforms increased from 3.2% in normal lung to 7.9% in lung tumors. Both the phosphorylated and unphosphorylated Op18 proteins were significantly increased in poorly differentiated tumors as compared to moderate or well-differentiated lung adenocarcinomas (P < 0.03), suggesting that upregulated expression of Op18 reflects a poor differentiation status and higher cell proliferation rates. This was further verified in A549 and SKLU1 lung adenocarcinoma cell lines by examining Op18 levels and phosphorylation status following treatment that altered either cell proliferation or differentiation. The increased expression of Op18 protein was significantly correlated with its mRNA level indicating that increased transcription likely underlies elevated expression of Op18. The overexpression of Op18 proteins in poorly differentiated lung adenocarcinomas and the elevated expression of the phosphorylated forms of Op18 may offer a new target for drug or gene directed therapy and may have potential utility as a tumor marker.