ARTICLE ABSTRACTPurpose: A crucial event in lung adenocarcinoma progression is the switch from an aerogenous spread toward an infiltrating tumor. Loss of RhoB expression has been suggested to be critical for lung cancer invasion. Here, we tested RhoB expression as a prognostic biomarker in non–small cell lung cancer (NSCLC) with a special focus on lepidic pattern.Experimental Design: We analyzed RhoB expression using both IHC and RT-qPCR in two series of operated patients (n = 100 and 48, respectively) and in a series of advanced lepidic adenocarcinoma (n = 31) from different hospitals. Next, we examined the role of RhoB in lung cancer progression in transgenic mice that express inducible EGFRL858R crossed with Rhob null mice.Results: We identified that loss of RhoB expression was strongly associated with worse survival (P = 0.0001) and progression-free survival (P < 0.001) in the first series. We then confirmed these results after multivariate analyses of the second series. In the series of adenocarcinoma with lepidic features issued from a clinical trial (IFCT-0401), we showed that loss of RhoB expression was associated with higher aggressiveness of stage IV. Finally, we showed that EGFRL858R/Rhob+/+ mice developed mainly diffuse lung tumors with a lepidic pattern, whereas EGFRL858R/Rhob+/− and EGFRL858R/Rhob−/− developed a greater number of tumors, and aggressive adenocarcinomas with invasive properties.Conclusions: We showed that RhoB is not only a strong prognostic factor in NSCLC but it is also critical for the acquisition of an aggressive phenotype of adenocarcinoma. Clin Cancer Res; 20(24); 6541–50. ©2014 AACR.