* Development Profiles Are Associated With Lung Non-small Cell ...

Non-small Cell Lung Cancer Molecular Profiles Are Associated With Lung Development* Alain C. Borczuk, Adel M. Assaad, Liqun Q. Wang, Kristin L. Walter and Charles A. Powell Chest 2004;125;115S-116S DOI 10.1378/chest.125.5_suppl.115S-a The online version of this article, along with updated information and services can be found online on the World Wide Web at: http://chestjournal.chestpubs.org/content/125/5_suppl/115S.2.full.html

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13 Garber ME, Troyanskaya OG, Schluens K, et al. Diversity of gene expression in adenocarcinoma of the lung. Proc Natl Acad Sci U S A 2001; 98:13784 –13789 14 Bhattacharjee A, Richards WG, Staunton J, et al. Classification of human lung carcinomas by mRNA expression profiling reveals distinct adenocarcinoma subclasses. Proc Natl Acad Sci U S A 2001; 98:13790 –13795 15 Wigle DA, Jurisica I, Radulovich N, et al. Molecular profiling of non-small cell lung cancer and correlation with disease-free survival. Cancer Res 2002; 62:3005–3008 16 Gordon GJ, Jensen RV, Hsiao LL, et al. Translation of microarray data into clinically relevant cancer diagnostic tests using gene expression ratios in lung cancer and mesothelioma. Cancer Res 2002; 62:4963– 4967 17 Sugita M, Geraci M, Gao B, et al. Combined use of oligonucleotide and tissue microarrays identifies cancer/testis antigens as biomarkers in lung carcinoma. Cancer Res 2002; 62:3971–3979 18 Welsh JB, Sapinoso LM, Kern SG, et al. Large-scale delineation of secreted protein biomarkers overexpressed in cancer tissue and serum. Proc Natl Acad Sci U S A 2003; 100:3410–3415 19 Park IW, Wistuba II, Maitra A, et al. Multiple clonal abnormalities in the bronchial epithelium of patients with lung cancer. J Natl Cancer Inst 1999; 91:1863–1868 20 Powell CA, Spira A, Derti A, et al. Gene expression in lung adenocarcinomas of smokers and non-smokers. Am J Respir Cell Mol Biol 2003; 29:157–162 21 Johnson SK, Kerr KM, Chapman AD, et al. Immune cell infiltrates and prognosis in primary carcinoma of the lung. Lung Cancer 2000; 27:27–35 22 Moran CJ, Arenberg DA, Huang CC, et al. RANTES expression is a predictor of survival in stage I lung adenocarcinoma. Clin Cancer Res 2002; 8:3803–3812 23 Zuo F, Kaminski N, Eugui E, et al. Gene expression analysis reveals matrilysin as a key regulator of pulmonary fibrosis in mice and humans. Proc Natl Acad Sci U S A 2002; 99:6292– 6297

Impact of Cigarette Smoke on the Normal Airway Transcriptome* Avrum Spira, MD; Frank Schembri, MD; Jennifer Beane; Vishal Shah; Gang Liu, PhD; and Jerome S. Brody, MD

(CHEST 2004; 125:115S) is unclear whether all smokers respond at a molecular I tlevel to cigarette smoke in the same fashion or whether smokers destined to develop cancer display early changes

*From The Pulmonary Center, Boston University School of Medicine (Drs. Spira, Schembri, Liu, and Brody); and the Bioinformatics Program, Boston University, Boston, MA (Dr. Spira, Ms. Beane, and Mr. Shah). This research was supported in part by grants HL71771 and ES10377 from Affymetrix, and by a Doris Duke Clinical Scientist Development Award (to AS). Reproduction of this article is prohibited without written permission from the American College of Chest Physicians (e-mail: [email protected]). Correspondence to: Avrum Spira, MD, The Pulmonary Center, 715 Albany St, R3, Boston, MA 02118-2526; e-mail: aspira@lung. bumc.bu.edu www.chestjournal.org

in gene expression that are premalignant in nature. Several studies1,2 have shown that smoking produces a “field defect” in the lung and its airways, such that molecular changes occur throughout the respiratory tract. To determine whether the molecular field defect is similar in all smokers, we have begun a study of gene expression profiles of airway epithelial cells obtained during bronchoscopy from normal nonsmokers and compared the airway transcriptome to that in smokers without cancer. Fiberoptic bronchoscopy was performed in 15 normal nonsmokers and in 15 smokers without cancer. The right upper lobe carina was brushed, and RNA was extracted and processed (U133A Genechip; Affymetrix; Santa Clara, CA) [approximately 22,500 genes]. Filter criteria for the quality of a sample were developed, and data were analyzed using a variety of computational algorithms. In nonsmokers, the variation in the expression level of genes among subjects was small. Multiple linear regression revealed that age, sex, and race had little effect on the nonsmoker transcriptome, as ⬍ 2% of the gene expression levels were associated with any of these three variables at the p ⬍ 0.01 level. Using a t test, 206 genes (4%) differed between smokers and nonsmokers at p ⬍ 0.01, and 610 genes differed at p ⬍ 0.05. The greatest increases in smokers occurred in genes associated with cell adhesion, detoxification, and secretion. The greatest decreases were in immune-regulatory genes and cytokeratins, with the decrease in the latter suggesting a change in epithelial cell differentiation. Linear regression analysis showed that the expression of 95 genes correlated (p ⬍ 0.01, R ⱖ 0.6) with the number of pack-years of smoking in smokers. These preliminary results have begun to define the gene expression profile of the normal human airway and to characterize the impact of cigarette smoking on the airway transcriptome. Our ultimate goal is to determine whether the transcriptome will identify early markers in those smokers who develop lung cancer.

References 1 Wistuba II, Lam S, Behrens C, et al. Molecular damage in the bronchial epithelium of current and former smokers. J Natl Cancer Inst 1997; 89:1366 –1373 2 Powell CA, Spira A, Derti A, et al. Gene expression in lung adenocarcinomas of smokers and nonsmokers. Am J Respir Cell Mol Biol 2003; 29:157–162

Non-small Cell Lung Cancer Molecular Profiles Are Associated With Lung Development* Alain C. Borczuk, MD; Adel M. Assaad, MD; Liqun Q. Wang, MS; Kristin L. Walter, MD; and Charles A. Powell, MD

(CHEST 2004; 125:115S–116S) Abbreviation: NSCLC ⫽ non-small cell lung cancer CHEST / 125 / 5 / MAY, 2004 SUPPLEMENT


115S

paradigms hold that lung carcinomas arise from C aurrent pleuripotent stem cell that is capable of differentiation into one or several histologic cell types. These paradigms suggest that lung tumor cell ontology is determined by the consequences of gene transcriptional activation and/or repression that recapitulates events that are important in embryonic lung development. We examined the relationship of non-small cell lung cancer (NSCLC) gene profiles with lung development.

Materials and Methods Using a DNA array (model U95Av2; Affymetrix; Santa Clara, CA), we acquired gene profiles from 7 nonmalignant lung specimens and 32 microdissected NSCLC tumors. Because we were interested in the genes that distinguished lung cancer histology, we focused our analysis only on tumors, and we utilized microdissection to minimize the confounding of profiles by genes expressed by adjacent nonmalignant structures. Neoplastic cells examined microscopically at ⫻40 magnification were microdissected with a 20-gauge needle, which enriched specimens for tumors and adjacent stroma. Unsupervised clustering demonstrated the fact that tumors were segregated tightly by histology. We determined the top 100 marker genes for adenocarcinoma, squamous cell carcinoma, large cell cancer, and carcinoid tumors using nearest-neighbor permutation analysis. All class assignments were statistically significant after 500 random permutations. The results were validated by immunostaining for 11 selected proteins using a tissue microarray that contained cylinders from 24 of 32 tumor specimens in addition to 56 other NSCLCs. The data of gene expression during lung development were obtained from accessing a publicly available data set that had been generated with a Mu11k mouse genome microarray (AJRCMB 26:541). Murine orthologs of human NSCLC histology gene markers were identified using computer software (Resourcerer, version 4.0; The Institute for Genomic Research; Rockville, MD). Self-organized mapping showed murine orthologs segregated into two temporally distinct clusters.

Results Large cell marker gene orthologs exclusively resided in a cluster expressed in pseudoglandular and canalicular stages, while adenocarcinoma orthologs were predominantly in the cluster expressed later in the terminal sac and alveolar stages of murine lung development. Representative large cell genes (eg, E2F3, MYBL2, HDAC2, CDK4, and PCNA) are expressed in the nucleus, and are associated with transcription regulation, cell cycle, and replication. Adenocarcinoma genes (eg, SFTPB, TTF-1, VEGFC, DOK1, and MUC1) are associated with lung-specific transcription, cell adhesion, and signal transduction. *From the Columbia University College of Physicians & Surgeons, New York, NY. Reproduction of this article is prohibited without written permission from the American College of Chest Physicians (e-mail: [email protected]). Correspondence to: Charles Powell, Assistant Professor of Clinical Medicine, Columbia University College of Physicians & Surgeons, Department of Medicine, Division of Pulmonary, Allergy & Critical Care Medicine, 630 W 168th St, New York, NY 10032; e-mail: [email protected] 116S

Conclusion Gene profiles of lung cancer histology subtypes are associated with lung development and clinical course. For example, large cell cancer genes are associated with proliferation, differentiation arrest, and poor prognosis, while adenocarcinoma genes are associated with glandular formation and improved outcomes relative to large cell carcinoma. In summary, NSCLC histology gene signatures recapitulate lung development pathways and provide insights into lung tumorigenesis.

Elevated Hepatocyte Growth Factor Level Correlates With Poor Outcome in Early-Stage and Late-Stage Adenocarcinoma of the Lung* Jill M. Siegfried, PhD; James D. Luketich, MD; Laura P. Stabile, PhD; Neil Christie, MD; and Stephanie R. Land, PhD

(CHEST 2004; 125:116S–119S) Abbreviations: HGF ⫽ hepatocyte growth factor; NED ⫽ no evidence of disease

he 5-year survival rate for patients with all stages of T lung cancer combined is only 15%. The survival rate is

48% for patients in whom disease is localized when detected, demonstrating that even when diagnosis occurs at an early stage of disease, relapse and death are common. Thus, identifying prognostic markers is critical for predicting patient survival in early disease and for determining proper therapeutic strategies. Hepatocyte growth factor (HGF) is a pleiotropic protein that induces cell proliferation and cell movement,1,2 and is a powerful angiogenic factor.3 It is primarily a paracrine factor that is produced by mesenchymal cells,4 although carcinoma cells may secrete HGF.5,6 The receptor for HGF, c-Met, is expressed by both epithelial and endothelial cells. The HGF/c-Met signaling pathway plays a significant role in the pathogenesis of many human cancers and is an attractive target for lung cancer therapy.7 The overexpres*From the Departments of Pharmacology (Drs. Siegfried and Stabile), Surgery (Drs. Luketich and Christie), and Biostatistics (Dr. Land), University of Pittsburgh Cancer Institute, Pittsburgh, PA. This research was supported by grants RO1 CA 79882 and P50 CA090440 (Specialized Program of Research Excellence in Lung Cancer) awarded to Dr. Siegfried from the National Cancer Institute. Dr. Stabile was supported by a fellowship from the American Lung Association. Reproduction of this article is prohibited without written permission from the American College of Chest Physicians (e-mail: [email protected]). Correspondence to: Jill M. Siegfried, PhD, The Hillman Cancer Center, UPCI Research Pavilion, Suite 2.18, Pittsburgh, PA 15213-1863; e-mail: [email protected]

Thomas L. Petty 46th Annual Aspen Lung Conference; Lung Cancer: Early Events, Early Interventions


Non-small Cell Lung Cancer Molecular Profiles Are Associated With Lung Development * Alain C. Borczuk, Adel M. Assaad, Liqun Q. Wang, Kristin L. Walter and Charles A. Powell Chest 2004;125; 115S-116S DOI 10.1378/chest.125.5_suppl.115S-a This information is current as of October 17, 2011 Updated Information & Services Updated Information and services can be found at: http://chestjournal.chestpubs.org/content/125/5_suppl/115S.2.full.html Permissions & Licensing Information about reproducing this article in parts (figures, tables) or in its entirety can be found online at: http://www.chestpubs.org/site/misc/reprints.xhtml Reprints Information about ordering reprints can be found online: http://www.chestpubs.org/site/misc/reprints.xhtml Citation Alerts Receive free e-mail alerts when new articles cite this article. To sign up, select the "Services" link to the right of the online article. Images in PowerPoint format Figures that appear in CHEST articles can be downloaded for teaching purposes in PowerPoint slide format. See any online figure for directions.
