Abstract 2362: 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 both early development and cancer whereby an epithelial cell transforms to a mesenchymal phenotype resulting in enhanced migration, invasiveness, and resistance to apoptosis. The transcription factor ZEB1 promotes EMT by repressing E-cadherin, and has been shown to promote tumor metastasis and increase resistance to standard chemotherapies in several human cancers. Its role in the tumorigenic progression of lung cancer however is less understood. Methods and Results: We analyzed genome-wide mRNA expression profiles across a cohort of 77 non small cell lung cancer (NSCLC) lines representing all common NSCLC histologies and oncogenotypes using Affymetrix GeneChip U133 Plus 2.0 arrays. Stratification of NSCLC lines by high/low ZEB1 expression identified 640 genes with significantly (p<0.001) different expression between the two groups. E-cadherin (CDH1) was significantly higher in cell lines with low ZEB1 expression, as was family member CDH3 and ERBB3. Mesenchymal markers vimentin, N-cadherin, (CDH2) as well as neuroendocrine markers (RTN1, NCAM1 and NEUROD1) were significantly up-regulated in cell lines with high ZEB1 expression. E-cadherin (r=-0.75) and Claudin 7 (r=-0.74) demonstrated the most significant indirect correlation with ZEB1 expression. To investigate the ability of ZEB1 to transform ‘normal’ epithelial cells we analyzed ZEB1 expression in an isogenic series of human bronchial epithelial cells (HBECs) - immortalized without viral oncoproteins - genetically manipulated to have single and multiple oncogenic changes common in lung cancer such as those for KRAS, p53, EGFR, and c-MYC. ZEB1 mRNA and protein expression was significantly increased in HBECs with combined oncogenic KRASV12 and p53 knockdown that had also been clonally selected for growth in soft agar. These transformed HBEC(KRASP53), which are tumorigenic in NOD/SCID mice in vivo, exhibit a decrease in epithelial markers and an increase in mesenchymal markers when compared to parental cells, indicating the cells have undergone EMT. To determine whether the observed increase in ZEB1 expression was causal to tumorigenicity or a bystander effect, ZEB1 expression was knocked down in HBEC(KRASP53) with RNA interference resulting in a significant decrease in colony formation, invasiveness and cell motility (p<0.005). Conclusion: These findings indicate that in NSCLC, ZEB1 expression is highly variable, is associated with an mRNA gene expression signature and is inversely correlated with E-cadherin expression, a known negative target of ZEB1. analyzing an isogenic series of progressively transformed HBECs we found ZEB1 increased with tumorigenic progression and an invasive phenotype. This phenotype could in turn be reversed by knockdown of ZEB1 suggesting ZEB1 plays an important role in tumorigenic transformation of human bronchial epithelial cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2362.