Non-small cell lung cancer cells survived ionizing radiation treatment display cancer stem cell and epithelial-mesenchymal transition phenotypes

Roberto Gomez-Casal,Michael Gibson,Lisa Bailey,Chitralekha Bhattacharya,Michael Epperly,Per Basse,Nandita Ganesh,Vera Levina

doi:10.1186/1476-4598-12-94

Abstract

Ionizing radiation (IR) is used for patients diagnosed with unresectable non small cell lung cancer (NSCLC), however radiotherapy remains largely palliative due to radioresistance. Cancer stem cells (CSCs), as well as epithelial-mesenchymal transition (EMT), may contribute to drug and radiation resistance mechanisms in solid tumors. Here we investigated the molecular phenotype of A549 and H460 NSCLC cells that survived treatment with IR (5Gy) and are growing as floating tumor spheres and cells that are maintained in a monolayer after irradiation.Non-irradiated and irradiated cells were collected after one week, seeded onto ultra low attachment plates and propagated as tumor spheres. Bulk NSCLC cells which survived radiation and grew in spheres express cancer stem cell surface and embryonic stem cell markers and are able to self-renew, and generate differentiated progeny. These cells also have a mesenchymal phenotype. Particularly, the radiation survived sphere cells express significantly higher levels of CSC markers (CD24 and CD44), nuclear β-catenin and EMT markers (Snail1, Vimentin, and N-cadherin) than non-irradiated lung tumor sphere cells. Upregulated levels of Oct-4, Sox2 and beta-catenin were detected in H460 cells maintained in a monolayer after irradiation, but not in radiation survived adherent A459 cells.PDGFR-beta was upregulated in radiation survived sphere cells and in radiation survived adherent cells in both A549 and H460 cell lines. Combining IR treatment with axitinib or dasatinib, inhibitors with anti-PDFGR activity, potentiates the efficacy of NSCLC radiotherapy in vitro.Our findings suggest that radiation survived cells have a complex phenotype combining the properties of CSCs and EMT. CD44, SNAIL and PDGFR-beta are dramatically upregulated in radiation survived cells and might be considered as markers of radiotherapy response in NSCLC.

Highlights

IntroductionLung cancer ranks highest in terms of both incidence and mortality [1]. Despite decades of research, systemic therapies fail to cure most lung cancers
Worldwide, lung cancer ranks highest in terms of both incidence and mortality [1]
Generation of non small cell lung cancer (NSCLC) cells survived ionizing radiation and propagated as tumor spheres A549 and H460 bulk NSCLC cells were treated with clinically relevant doses of Ionizing radiation (IR) (5.0 Gy) and cultured for one week

Summary

Introduction

Lung cancer ranks highest in terms of both incidence and mortality [1]. Despite decades of research, systemic therapies fail to cure most lung cancers. Radiotherapy used for patients with unresectable NSCLC tumors remains largely palliative due to radioresistance [1,3] which is possibly due to tumor heterogeneity in terms of cell of origin, pathology, etiology and molecular/genetic pathogenesis [4]. The existence of cancer stem/progenitor cells (CSCs) or tumor-initiating cells (TICs) reflects the cellular heterogeneity within solid tumors [5,6,7]. These cells are the undifferentiated cells with a high tumorigenic and self-renewal capacity, which have been identified in various human malignancies including breast, brain, prostate, pancreatic, colon and lung cancer [8,9,10,11,12,13]. Evidence suggests that the failure of the treatment may be due to the existence of CSCs which are resistant to chemo- and radiotherapy [14,15,16]

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