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Purpose: The purpose of this study was to determine the frequency and overall contribution of specific gene amplification events in the formation of Barrett's adenocarcinomas. The relationship of gene amplification to clinical-pathological variables and its potential usefulness as a marker for early cancer detection were also examined., Experimental Design: We used quantitative PCR and Southern blot analysis to screen 87 cases of Barrett's adenocarcinoma for the presence or absence of 13 distinct gene amplification events. Gene amplification was then examined for correlation with other amplification events and clinical variables (survival, stage, nodal involvement, tumor invasion, smoking history, and gender). Additionally, 22 specimens of Barrett's with high-grade dysplasia (HGD) were examined for the presence of gene amplification., Results: One or more amplification events were present in 50 of 87 (57%) adenocarcinomas. The ERBB2 gene was amplified in 19 of 87 (21.8%), CCNE1 in 11 of 87 (12.6%), GATA4 in 9 of 87 (10.3%), KRAS in 9 of 87 (10.3%), EGFR in 7 of 87 (8.0%), CCND1 in 6 of 87 (6.8%), HNF3alpha in 5 of 87 (5.7%), PIK3CA in 5 of 87 (5.7%), C-MYC in 4 of 87 (4.6%), DYRK2 in 2 of 87 (2.3%), and AIB1, AKT1, and IGF1R were amplified in 0 of 87 (0%) of the tumors. CCND1 amplification was found to correlate negatively with survival (P < 0.05). In addition, the ERBB2 amplicon positively correlated (P < 0.05) with GATA4 amplification. Increased copy number of the ERBB2 (1 of 22), GATA4 (1 of 22), KRAS (2 of 22), C-MYC (1 of 22), CCNE1 (2 of 22), and CCND1 (2 of 22) genes was also observed in one or more Barrett's adenocarcinomas with HGD., Conclusions: The high frequency of gene amplification in esophageal adenocarcinomas and HGD indicates the important role of these events in esophageal adenocarcinoma development. Additionally, these results underscore the possible usefulness of early detection approaches and chemotherapeutic strategies (ErbB2 and cyclin D1) targeted against amplified gene products.

Purpose: The purpose of this study was to determine whether expression of migration inhibitory factor (MIF) is increased in non-small cell lung cancer, and whether it correlates with angiogenesis and/or prognosis., Experimental Design: We measured vessel density, and levels of MIF, angiogenic CXC chemokines, and vascular-endothelial growth factor (VEGF; by ELISA) in tumor and normal lung tissue from 87 patients after resection of lung cancer. We compared vessel density with levels of MIF, VEGF, or angiogenic CXC chemokines in the corresponding tumor homogenate. Disease-free survival was analyzed in a Cox proportional hazards model., Results: Levels of MIF in lung cancer demonstrated a bimodal distribution, with some having "normal" values (relative to normal lung tissue) and a second cluster with markedly high values. The increased levels of MIF in lung cancer were statistically significant in both paired and unpaired comparisons (P < 0.05). The strongest correlation of vessel density was with the sum of angiogenic CXC chemokines. MIF correlated very strongly with levels of angiogenic CXC chemokines. Tumors in the high MIF group had a strong correlation between MIF level and vessel density. Risk of recurrence was associated with high levels of glutamic acid-leucine-arginine amino acid motif CXC chemokines, MIF, and/or VEGF in a Cox proportional hazards model., Conclusions: MIF expression in non-small cell lung cancer occurs in a bimodal distribution, and is closely associated with tumor levels of angiogenic CXC chemokines and with vessel density. High levels of tumor-associated CXC chemokines, MIF, or VEGF are associated with risk of recurrence after resection of lung cancer.

Purpose: The presence of an active lymphocytic response (ALR) in non-small cell lung cancer (NSCLC) tumors has previously been associated with a more favorable prognosis. The purpose of this study was to identify differences in global gene expression profiles between stage I NSCLC tumors with ALR (ALR+) and those without ALR (ALR-)., Experimental Design: Sixty-three stage I lung adenocarcinomas were analyzed for gene expression using Affymetrix oligonucleotide microarrays. Tumors were stratified into ALR+ and ALR- groups and compared for statistically significant differences in gene expression. Identified candidate genes were validated using both ELISA and immunohistochemistry. Follow-up data for these patients were collected and used to assess patient prognosis., Results: Of the 63 tumors studied, 27 were ALR+ and 36 were ALR-. A total of 303 genes showed significant differences in gene expression between the two populations (t test, P < 0.02). Three of the genes overexpressed by ALR+ tumors were the chemokines: small inducible cytokine A4 (MIP-1beta), RANTES, and interferon inducible protein 10 (IP-10). Immunohistochemistry analysis showed that the tumor cells expressed these cytokines. ELISA showed that MIP-1beta and RANTES were overexpressed at the protein level by ALR+ tumors. Univariate Cox proportional hazards analysis showed that RANTES was a predictor of survival in stage I lung adenocarcinomas (P = 0.002)., Conclusion: When tested in the Cox univariate proportional hazards model, RANTES expression by lung adenocarcinoma cells is a predictor of survival in stage I NSCLC patients and may be useful as a prognostic factor in lung cancer.

Purpose: The goal of this study was to identify potential protein markers in lung adenocarcinomas., Experimental Design: A series of 93 lung adenocarcinomas (64 stage I and 29 stage III) and 10 uninvolved lung samples were examined for quantitative differences in protein expression using two-dimensional PAGE. Candidate proteins were identified using matrix-assisted laser desorption/ionization mass spectrometry or peptide sequencing. The levels of the individual isoforms of nine proteins found to be overexpressed in the lung tumors were examined. Potential mechanisms for overexpression were examined by comparing mRNA expression levels, assessed using oligonucleotide arrays, to the protein values in the same samples., Results: Antioxidant enzyme AOE372, ATP synthase subunit d (ATP5D), beta1,4-galactosyltransferase, cytosolic inorganic pyrophosphatase, glucose-regulated M(r) 58,000 protein, glutathione-S-transferase M4, prolyl 4-hydroxylase beta subunit, triosephosphate isomerase, and ubiquitin thiolesterase (UCHL1) were identified as being significantly overexpressed in lung adenocarcinomas. The expression of these proteins was increased from 1.4- to 10.6-fold as compared with uninvolved lung tissue. The expression of the individual protein isoforms was correlated with 10 clinicopathological variables as well as with each gene's mRNA level in the same sample. Both isoforms of glucose-regulated M(r) 58,000 protein were found to be significantly correlated with their mRNA expression profiles (P < 0.05), indicating that increased transcription likely underlies the increased expression of these proteins., Conclusions: Two-dimensional PAGE and mass spectrometry can identify proteins showing increased expression in lung adenocarcinoma. The association of specific isoforms of these proteins with clinical variables and understanding the regulation of their expression will aid in determination of their potential use as biomarkers in this cancer.