Abstract 3419: Defining the role of ZEB1 in the molecular pathogenesis of lung cancer

Abstract

Abstract Background: Epithelial-to-mesenchymal transition (EMT) is an important biologic process in cancer resulting in enhanced migration, invasiveness, and therapeutic resistance. The transcription factor ZEB1 promotes EMT and tumor metastasis in several human cancers yet its role in the pathogenesis of lung cancer is less well defined. In addition to a large panel of lung cancer lines, our laboratory has developed an isogenic series of human bronchial epithelial cells (HBECs) – immortalized without viral oncoproteins – genetically manipulated with single and multiple oncogenic changes common in lung cancer (sh-p53, KRASV12, mutant EGFR, and c-MYC), providing an invaluable resource to study the progression of bronchial epithelial cells towards malignancy. Methods and Results: ZEB1 expression was analyzed by qRT-PCR across a panel of 77 non small cell lung cancer (NSCLC) lines to test for a direct correlation with ‘M’ status and indirect correlation with CDH1 expression and ‘E’ status. Genome-wide mRNA expression profiling of the NSCLC cell line cohort using Affymetrix U133 Plus 2.0 arrays was used to identify novel ZEB1-associated genes. Expression of the ephrin receptor EPHA1 significantly inversely correlated with ZEB1 expression. This was confirmed in independent siRNA experiments where knockdown of ZEB1 resulted in an increase in EPHA1 in both NSCLC cell lines and oncogenically transformed HBECs. To characterize the role of ZEB1 in the oncogenic progression of bronchial epithelial cells, ZEB1 was analyzed across our series of genetically manipulated HBECs. Expression of ZEB1 was generally absent, but was highly up-regulated with the combination of p53-knockdown, KRASV12, and c-MYC over-expression. This onco-genetic combination led to the most tumorigenic phenotype in vivo accompanied with a switch from ‘E’ to ‘M’ status. The functional importance of ZEB1 was seen with siRNA knockdown of ZEB1 resulting in a significant decrease in soft agar colonies, transwell invasiveness, and motility suggesting ZEB1 is a crucial driver of tumorigenicity in sh-p53 + KRASV12 + c-MYC transformed HBECs. To confirm the oncogenic role of ZEB1 in lung cancer, shRNA-mediated knockdown of ZEB1 was performed in several NSCLC cell lines, leading to a significant decrease in soft agar colonies, transwell invasiveness, motility, and 3D growth. Subcutaneous injection into NOD/SCID mice found inhibition of tumor growth in shZEB1 knockdown lines. Finally, loss of ZEB1 expression in NSCLCs and transformed HBECs resulted in a decrease in a “cancer stem cell” like population detected by ALDH activity. Conclusion: These findings involving mechanistic studies in NSCLC lines and oncogenically manipulated HBECs demonstrate the prominent role of ZEB1 function in EPHA1 expression, EMT, tumorigenicity, and the invasive phenotype of lung cancer, supporting the important role of ZEB1 in the pathogenesis of lung cancer making it an important new therapeutic target. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3419. doi:10.1158/1538-7445.AM2011-3419