Soluble E-cadherin as a microenvironmental factor which enhances tumor progression

Abstract

This study aims to investigate two questions: How do tumor cells interact and alter the normal epithelial cells and whether elevated soluble E-cadherin (sE-cad) levels in the microenvironment impact normal epithelial cells as well. We use a three-dimensional (3D) cell-culture system comprising non-transformed Madin-Darby canine kidney (MDCK) epithelial cells and invasive cancer cells (MSV-MDCK) to determine the impact of tumor cells on the normal epithelial cells. Using 3D co-culture system we demonstrate that carcinoma cells sequentially induced pre-neoplastic lumen filling and EMT in normal epithelial cysts. We provide evidence that carcinoma cells secrete MMP-9 which cleaves extracellular domain of E-cadherin from normal epithelial cells to generate soluble E-cadherin (sE-cad). Addition of purified sE-cad and by immunodepletion of sE-cad from conditioned medium prevented filling up of the lumen indicating that sE-cad was necessary and sufficient to induce lumen filling. Lumen filling is mediated by the activation of EGFR and its downstream ERK and AKT pathways. Long term (72-96hrs) sE-cad treatment induced EMT in normal epithelial cysts and the AKT pathway was found to be implicated in the induction of EMT. We demonstrate our results using in vitro 3D culture system, confocal microscopy and western blotting. While some aspects of the TME such as ECM remodeling and tumor angiogenesis are well studied, the interaction between tumor cells and the adjacent epithelium is poorly understood and this study demonstrates that cancer cells can utilize the surrounding normal epithelial cells as accessories to produce factors necessary to maintain and promote tumor growth. This finding may have important clinical correlations, as our results suggest that elevated sE-cad levels observed in cancer patients' sera are derived from both tumor cells as well as adjacent normal epithelial cells. In vivo, carcinoma cells induced shedding of cell bound E-cadherin from adjacent normal tissue may result in transdifferentiation of the compact epithelial tissue into disorganized mesenchymal phenotype. Elevated sE-cad levels may also have pro-tumorigenic effects on other components of the TME such as the various stromal cell types. Thus, accumulation of sE-cad in the microenvironment may have additive or synergistic effects on the pro-oncogenic TME since it can possibly alter the different components of the TME. Our results further support the idea that effectively eliminating sE-cad may benefit current EGFR-based therapies against various carcinomas as sE-cad can induce EGFR activation even in normal epithelial cells, and blocking sE-cad reduces the phosphorylated levels of several RTKs, the absence of sE-cad in the sera may be of importance for the efficacy of EGFR inhibitor-based therapeutic intervention.