The early diagnosis of lung cancer is an effective approach to reduce the mortality caused by malignancy.To explore serum biomarkers of lung cancer at early stage, M-BE, a SV40T-transformed human bronchialepithelial cell line with the phenotypic features of early tumorigenesis at high passage, was cultured inthe conditioned media to collect its secretory proteins. The proteins secreted from different passageM-BE cells were extracted and then separated by two-dimensional electrophoresis (2-DE). MALDI-TOF/TOF mass spectrometry was adopted to identify the passage-dependent 2-DE spots. Totally, 47 proteinswere identified, including 23 that were up-regulated and 24 that were down-regulated. Of these proteins,cathepsin D was a typical secretory protein that exhibited the increased abundance either in culturemedia or in cells during passaging. Furthermore, the proteomic conclusions were validated in the clinicalsamples of lung cancer patients. When sandwich ELISA was used, the concentrations of cathepsin Din plasma showed significant differences between lung squamous cell carcinomas (SCC, 104 cases)and normal donors (36 cases,
p 0.015). When tissue microarray (TMA) was used, cathepsin Dexpression levels in SCC tissues (178 cases) were significantly higher than those in normal donors (40cases,
p < 0.001). The present study has revealed that M-BE cells at different passages could secreteor release some proteins into the living environment, which might serve as the potential resource forexploring the biomarkers of lung cancer.