Abstract A54: Mithramycin inhibits cigarette smoke-mediated induction of ABCG2 in lung and esophageal cancer cells: Implications for targeting cancer stem cells in thoracic malignancies.

Abstract

Abstract Limited information is available regarding epigenetic mechanisms by which cigarette smoke mediates initiation and progression of thoracic malignancies. In order to examine this issue, lung and esophageal cancer cells (A549, Calu-6, NCI-SB-ESC1 and NCI-SB-ESC2), were cultured in normal media (NM) with or without cigarette smoke condensate (CSC) under clinically relevant exposure conditions. Microarray analysis revealed that five day CSC exposure significantly up-regulated ABCG2, encoding a xenobiotic pump highly expressed in cancer stem cells. Quantitative reverse transcription-PCR (qRT-PCR), western blot, and immunohistochemistry experiments confirmed up-regulation of ABCG2 in cultured cancer lines, but not normal small airway epithelial cells (SAEC) or immortalized esophageal squamous cells (HET1A) exposed to CSC. Flow cytometry experiments demonstrated that CSC increased the side population (SP) of cultured cancer cells. Transient transfection experiments using ABCG2 promoter reporter constructs revealed that deletion of xenobiotic response elements as well as SP-1 sites markedly attenuated ABCG2 induction by CSC. Chromatin immunoprecipitation (ChIP) experiments revealed that CSC-mediated induction of ABCG2 coincided with increased occupancy of aryl hydrocarbon receptor (AHR), SP-1, and Nrf2, as well as increased levels of RNA pol II and H3K9Ac within the ABCG2 promoter. Under conditions potentially achievable in clinical settings, mithramycin diminished basal as well as CSC-mediated increases in AHR, SP-1, and Nrf2 levels within the ABCG2 promoter, down-regulated ABCG2 expression, decreased S P, inhibited in-vitro proliferation of lung and esophageal cancer cells, and markedly diminished growth of established tumor xenografts. Microarray analysis revealed significant down-regulation of numerous stem-cell related pathways in lung cancer cells following mithramycin exposure. Collectively, these findings provide a potential mechanistic link between smoking status and outcome of patients with lung and esophageal cancers, and support clinical evaluation of mithramycin for targeting cancer stem cells in thoracic malignancies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr A54.