Abstract 2089: The role of oxidative stress in the survival of ECM-detached mammary epithelial cells

Abstract

Abstract Tumorigenic mammary epithelial cells have been found to gain the ability to survive without the critical signals provided from the extracellular matrix (ECM), which are vital in normal mammary epithelial cells. However, the mechanisms underlying how cancerous epithelial cells gain the ability to survive without attachment to the ECM are poorly understood. Furthermore, the mechanisms in place to prevent survival of cells detached from the ECM are also not well characterized. Previous publications and work from our laboratory have found that there is an increase in ROS when mammary epithelial cells are detached from the ECM. In addition, it has been found that increased levels of ROS result in inhibition of fatty acid oxidation and subsequently in cell death. Neutralization of ROS in detached mammary epithelial cells through Trolox (a Vitamin E analog and antioxidant compound) treatment have been found to increase fatty acid oxidation efficacy and thereby rescue a detachment-induced loss of ATP. This suggests that detachment-induced ROS may be important in the elimination of detached cells. We are currently working on further investigating this phenomenon. Currently, we have found that other antioxidant compounds such as Vitamin C, and Epigallocatechin gallate (EGCG), also rescue the detachment-induced ATP deficiency. In addition, treating MCF-10A cells (primed for soft agar growth through the expression of HPV-E7 and Bcl-2) with these antioxidant compounds results in a significant enhancement of colony formation in soft agar. In addition to the antioxidant compound studies, we have engineered MCF-10A cells to overexpress catalase, an enzyme that efficiently neutralizes oxidative stress. We have found that directing expression of catalase to the peroxisomes results in a significant rescue of ATP levels, suggesting that the peroxisomes may be involved in this mechanism of inhibition of fatty acid oxidation. When grown in Matrigel to induce the formation of 3-dimensional structures, these newly engineered MCF-10A cells will form structures with filled lumen suggesting that catalase expression promotes the survival of ECM-detached cells. Currently, we are working to understand the source of this detachment-induced production of ROS as well as to understand the mechanism by which ROS inhibits fatty acid oxidation. Furthermore, we are working to determine the importance of neutralizing oxidative stress in tumor cell survival. In order to study the location of ROS production as well as the mechanism of fatty acid oxidation inhibition, we are utilizing siRNA/shRNA techniques, pharmacological inhibitors, and mass spectrometry. In the near future, we will use mouse xenograft studies to examine the importance of detachment-induced oxidative stress in tumor formation. 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 2089. doi:10.1158/1538-7445.AM2011-2089