Abstract 2603: Cancer stem cells and dedifferentiation: the stromal match-point.
Authors: Rodrigues Carlos F. , Val Mariana , Rodrigues Inês P. , Ferreira Susana , Botelho Filomena , Ramalho José , Zhitkovich Anatoly , Carreira Isabel M. , Alpoim Carmen 
Published: August 2014

Abstract
Bypassing all the research advances in the last decades, cancer remains as a major public health problem affecting more than 1.5 million (M) new individuals each year just in the USA, and killing more than 0.5 M. Recent research emphasized the major role of cancer stem cells (CSCs) in the metastatic disease, the main cause of cancer patients mortality. CSCs drive tumorigenesis and differentiation, contributing to tumors’ heterogeneity and to their relative chemo- and radiotherapy resistance and eventually relapse. Following CSCs identification, targeted therapeutic approaches have been developed to abolish them. However, CSCs can reemerge through dedifferentiation of tumor-committed stromal cells condemning this therapeutics. The mechanisms behind dedifferentiation are still unclear and are the main focus of our investigation.
Lung cancer is one of the main causes of cancer-related deaths worldwide. Its prevalence is increasing due to the widespread smoking habits and increasing accumulation of atmosphere pollutants. In this work hexavalent chromium [Cr(VI)] was selected as a model of cancer carcinogenesis mainly due to is increasing occupational relevance. The non-malignant human bronchial epithelial cell line BEAS-2B was malignantly transformed into the RenG2 system using low density culture in the presence of Cr(VI). A parallel control cell line (Cont1) was produced under the same conditions, though, in the absence of Cr(VI). Two additional cell lines (DRenG2 and DDRenG2) were attained following serial rounds of injection in nude mice. Metabolic studies using [18F]fluoro-2-deoxyglucose) and nuclear magnetic resonance spectroscopy performed in all the cell lines revealed a more glycolytic phenotype for the derivatives (DRenG2 and DDRenG2), compatible with a quiescent phenotype. Subsequent karyotype and real time PCR-based cellular characterization identified different cellular sub-populations within each cell line, strengthening the hypothesis on the CSCs presence. The sphere-formation assay, used to search for CSCs presence, revealed the presence of CSCs only DRenG2 and in DDRenG2 cell lines. This suggested that a dedifferentiation process featured the formation of CSCs during RenG2 derivation in nude mice. The involvement of mice stroma in this process was uncovered by surgical isolation of mouse stromal cells of the subcutaneous compartment and subsequent co-culture with RenG2 cells for 30-60 days (time needed to induce tumors in mice with RenG2), which resulted in the emergence of a CSCs sub-population. A cytokine multiplex array analysis performed in the conditioned medium of the co-cultured cells in parallel with comparative genome hybridization array (aCGH) analysis performed in all the cell lines under study revealed a panel of potential paracrine orchestrators of this stromal-induced dedifferentiation process.
Citation Format: Carlos F. Rodrigues, Mariana Val, Inês P. Rodrigues, Susana Ferreira, Filomena Botelho, José Ramalho, Anatoly Zhitkovich, Isabel M. Carreira, Carmen Alpoim. Cancer stem cells and dedifferentiation: the stromal match-point. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2603. doi:10.1158/1538-7445.AM2013-2603