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Purpose: Autoantibodies against tumor antigens represent one type of biomarkers that could be assayed in serum for detection of cancer and monitoring disease progression. In this study, we applied the proteomics-based approach to identify candidate tumor antigens in non-small cell lung cancer (NSCLC). Experimental Design: Proteins from NSCLC were separated on 1-DE or 2-DE gels and transferred onto membranes that are then incubated with sera from patients with NSCLC or non-cancer control subjects. Proteins that react specifically with antibodies in sera from NSCLC patients were identified by mass spectrometry and further evaluated by western blotting and ELISA using recombinant proteins. Results: By proteomics-based screening, we firstly identified some proteins such as alpha-enolase and gp96 that may elicit humoral response in NSCLC patients. Then we confirmed that autoantibodies against alpha-enolase were present in a subset of NSCLC patient sera by western blotting using recombinant protein. Moreover, to evaluate autoantibodies against alpha-enolase as a diagnostic marker, the frequencies of the autoantibodies in sera were determined by ELISA using recombinant protein. We tested 68 sera from patients with NSCLC, 8 sera from patients with small cell lung cancer (SCLC), 8 sera from patients with gastrointestinal cancer, 9 sera from patients with mycobacterium avium complex (MAC) and 60 sera from non-cancer control subjects. When “Mean ODnon-cancer control sera+3SDnon-cancer control sera” was used as cutoff point, the prevalence of autoantibodies against alpha-enolase was 32.4% in patients with NSCLC (22 of 68), 1.7% in non-cancer control subjects (1 of 60), while none of sera from 8 patients with SCLC, 8 patients with gastrointestinal cancer and 9 patients with MAC. The prevalence of autoantibodies was 42.8% in advanced NSCLC (stage III/IV), which is significantly higher than stage I/II (15.4%). Immunohistochemical analysis revealed that alpha-enolase is overexpressed in malignant lung tissue from NSCLC patient who showed the presence of autoantibodies against alpha-enolase. Thus immunogenicity of alpha-enolase in NSCLC is likely to be related to over-expression of alpha-enolase in malignant lung tissue. Conclusions: Autoantibodies against alpha-enolase may be a promising biomarker of NSCLC and may be a key molecule in immunotherapy against NSCLC.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]