Abstract A11: Airway gene expression indicates an increase in oxidative phosphorylation in the field of injury of individuals with premalignant lesions
Authors: Mazzilli Sarah A. , Liu Gang , Zhang Xiaohui , Si Huiqing , Lam Stephen , Lenburg Marc , Spira Avrum , Beane Jennifer E. 
Published: October 2015

Lung squamous cell carcinoma (SCC) claims the lives of approximately 50,000 individuals in the United States annually due to late stage diagnosis and lack of effective treatment options. Lung SCC arises in the epithelial layer of the bronchial airways and is preceded by the development of premalignant lesions (PMLs). The molecular events involved in the progression of PMLs to lung SCC are not clearly understood as not all PMLs that develop go on to form carcinoma. Our group is now using high-throughput genomic techniques to characterize the process of premalignant progression by examining PMLs and non-lesion areas in the airway of individuals with lesions (i.e. the “field of injury”,) to identify events that lead to the development of SCC.

mRNA-Seq was conducted on cytologically-normal proximal airway epithelium collected by brushing the mainstem bronchus from current and former smokers with (n=43) and without (n=32) PMLs (metaplatic lesions excluded). Linear modeling strategies were used to identify genes altered between subjects with and without PMLs (n=801 out of 13,916 genes at FDR&lt;0.01). Genes were significantly concordantly enriched (FDR&lt;0.01) in an overlapping sample set profiled by microarray (n=57 out of the 75 samples, 33/57 with PMLs) and enriched among up-regulated genes in the independent sample set (n=158, 38/158 with PMLs). Pathway analysis by GSEA revealed enrichment (FDR&lt;0.05) of oxidative phosphorylation (OXPHOS) /respiratory electron transport among genes up-regulated in the airways of subjects with PMLs.

OXPHOS is the most efficient metabolic pathway that generates energy in the form of ATP by utilizing the structures and enzymes of the mitochondria and OXPHOS is often elevated during repair epithelial tissue. However, OXPHOS metabolism is often superseded in the development of cancer by glycolysis. To validate these computational findings related to the OXPHOS pathway, we examined oxygen consumption rates (OCR) and extracellular acidification rates (ECAR) in lung cell lines. These cell lines have either high OXPHOS and low glycolytic gene expression (H1299) or low OXPHOS and high glycolytic gene expression (HCC4006). Consistent with the gene expression pattern in these cell lines, The OCR for H1299s and HBECs were 10% higher than HCC4006 cells reflecting higher OXPHOS activity (p = 0.06). Whereas the ECAR was 39% higher in HCC4006 than H1299 cells and 46 % higher than HBECs (p&lt;0.001and p&lt;0.001 respectively), reflecting higher glycolytic metabolism corresponding to the gene expression. Next, we measured protein expression of genes that were elevated in the field of injury of PMLs, which included: mitochondrial membrane protein, translocase of the outer membrane (TOMM 22) and a subunit of complex IV of the electron transport chain, cytochrome C oxidase (COX-IV), in FFPE sections of lung tissue from the N-nitroso-tris-chloroethylurea (NTCU) mouse model of lung SCC. The NTCU model develops PMLs in the bronchial epithelium that progress to lung SCC in a histologically similar pattern to that observed in humans; it is important to note the carcinogen is administered cutaneously and not inhaled thus the field of injury may differ from inhaled carcinogens such as tobacco smoke. Protein levels of TOMM 22, and COX-IV were found to be elevated in dysplastic lesions compared to untreated controls. Together these data suggest that metabolism-associated gene expression is correlated with cellular metabolism and furthermore there is an increase in OXPHOS associated with the development of PMLs. While we are currently in the process of validating these findings in human normal and PMLs, our results suggest that there is potential that therapeutically increasing or maintaining OXPHOS in premalignant lesions or the field of injury may be a mechanism of prevention for lung cancer.

Citation Format: Sarah A. Mazzilli, Gang Liu, Xiaohui Zhang, Huiqing Si, Stephen Lam, Marc Lenburg, Avrum Spira, Jennifer E. Beane. Airway gene expression indicates an increase in oxidative phosphorylation in the field of injury of individuals with premalignant lesions. [abstract]. In: Proceedings of the Thirteenth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2014 Sep 27-Oct 1; New Orleans, LA. Philadelphia (PA): AACR; Can Prev Res 2015;8(10 Suppl): Abstract nr A11.