Selective Calcium Sensitivity in Immature Glioma Cancer Stem Cells

Maria Niklasson,Karin Forsberg-Nilsson,Anna Segerman,Sten Linnarsson,Sven Nelander,Linnéa Schmidt,Voichita Dana Marinescu,Bengt Westermark,Peter Dirks,Lene Uhrbom,Michael Andäng,Shimei Wee,Annika Hermansson

doi:10.1371/journal.pone.0115698

Abstract

Tumor-initiating cells are a subpopulation in aggressive cancers that exhibit traits shared with stem cells, including the ability to self-renew and differentiate, commonly referred to as stemness. In addition, such cells are resistant to chemo- and radiation therapy posing a therapeutic challenge. To uncover stemness-associated functions in glioma-initiating cells (GICs), transcriptome profiles were compared to neural stem cells (NSCs) and gene ontology analysis identified an enrichment of Ca2+ signaling genes in NSCs and the more stem-like (NSC-proximal) GICs. Functional analysis in a set of different GIC lines regarding sensitivity to disturbed homeostasis using A23187 and Thapsigargin, revealed that NSC-proximal GICs were more sensitive, corroborating the transcriptome data. Furthermore, Ca2+ drug sensitivity was reduced in GICs after differentiation, with most potent effect in the NSC-proximal GIC, supporting a stemness-associated Ca2+ sensitivity. NSCs and the NSC-proximal GIC line expressed a larger number of ion channels permeable to potassium, sodium and Ca2+. Conversely, a higher number of and higher expression levels of Ca2+ binding genes that may buffer Ca2+, were expressed in NSC-distal GICs. In particular, expression of the AMPA glutamate receptor subunit GRIA1, was found to associate with Ca2+ sensitive NSC-proximal GICs, and decreased as GICs differentiated along with reduced Ca2+ drug sensitivity. The correlation between high expression of Ca2+ channels (such as GRIA1) and sensitivity to Ca2+ drugs was confirmed in an additional nine novel GIC lines. Calcium drug sensitivity also correlated with expression of the NSC markers nestin (NES) and FABP7 (BLBP, brain lipid-binding protein) in this extended analysis. In summary, NSC-associated NES+/FABP7+/GRIA1+ GICs were selectively sensitive to disturbances in Ca2+ homeostasis, providing a potential target mechanism for eradication of an immature population of malignant cells.

Highlights

Glioblastoma multiforme (GBM) is a highly malignant form of brain cancer with poor prognosis for affected individuals
The stemness-associated gene SOX2, the neural stem cells (NSCs) marker BLBP as well as the neuronal marker TUBB3 (Tubulin beta III), which may reflect high potency for concomitant neuronal differentiation, were expressed the highest in NSCs, and expression levels decreased in the order of the GliNS1 group
Our study identified a stemness-associated sensitivity in glioma initiating cells (GICs) related to Ca2+ homeostasis and signaling

Summary

Introduction

Glioblastoma multiforme (GBM) is a highly malignant form of brain cancer with poor prognosis for affected individuals. Malignant glioma tumors are highly heterogenous, a subpopulation of immature cells, termed glioma initiating cells (GICs) [2,3,4,5,6] coexist with more differentiated cell populations. Reflecting the immaturity of GICs and their ability to differentiate [8], these cells have been shown to share a stem cell (stemness)-associated gene expression with stem cell populations, such as teratoma-forming normal embryonic stem cells [9,10,11], and it is proposed that GICs continuously resupply the bulk tumor cells through self-renewal and differentiation [11, 12]. Much of the drug development research for GBM treatment has focused on targeting bulk cells, most of which lack tumor-initiating capacity. A major challenge that remains is increasing the efficacy of cancer treatment targeting GICs as these cells exhibit resistance to chemo- and radiotherapy using current strategies

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