Abstract 2869: Activation of AMP-activated protein kinase(AMPK) enhances Radiosensitization via inhibition of mTOR pathway in Rapamycin-resistant non-small cell lung cancer cells

Abstract

Abstract Purpose: Most drug resistance cancer cells have resisted to radiation therapy. In this study, we hypothesized that inhibition of mammalian target of rapamycin(mTOR) via activation of AMPK could potentially enhance radiosensitization in Rapamycin-resistant non- small cell lung cancer stem cells. Methods: Stem cell marker was detected with CD133 and ALDH antibodies. P-AKT, P-mTOR and P-S6R were detected by Western Blotting. Clonogenic assay was used to determine radiosensitivity. Apoptosis was measured using the Annexin V-fluorescent isothiocyante apoptosis detection kit I (Pharmingen) with flow cytometry in vitro, and cleaved capase 3 by Western botting. Autophagy was determined by punctuate localization of GFP-LC3 fusion protein, which observed under a confocal fluorescence microscope in vitro. ATP levels were measured using Bioluminescence Assay kit CLS II from Roche Scientific (Indianpolis, IN, USA) as per the manufacturer's protocol. Results: Level of CD133 and ALDH expression which are markers of stem cell, P-Akt, p-mTOR and P-S6R proteins were enhanced in Rapamycin-resistant non- small cell lung cancer stem cells (CSNSCLCSTC) when compared to parental non-small cell lung cancer (H460) cells. Rapamycin-resistant non- small cell lung cancer stem cells mediate less radiation sensitivity when compared with parental cells in clonogenic assay(DER=0.82, p=0.02). However Metformin, activator of AMPK, obviously enhanced sensitivity of Rapamycin-selected non- small cell lung cancer stem cells to radiation (DER=1.2, p=0.01). Therefore, this enhanced radiosensitization was associated with a downregulation of mTOR signaling pathway and an increase in characteristic punctuate localization of GFP-LC3 as a marker for autophagy. Conclusions: These findings may be utilized as a novel strategy to enhance radiation therapy in drug-selected non-small cell lung cancer stem cells. 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 2869. doi:10.1158/1538-7445.AM2011-2869