Comparative Transcriptomic Analysis of Temozolomide Resistant Primary GBM Stem-Like Cells and Recurrent GBM Identifies Up-Regulation of the Carbonic Anhydrase CA2 Gene as Resistance Factor.

Ricarda Hannen,Till Acker,Barbara Carl,Thorsten Stiewe,Andrea Nist,Axel Pagenstecher,Martin Selmansberger,Jörg Walter Bartsch,Christopher Nimsky,Maria Hauswald

doi:10.3390/cancers11070921

Abstract

About 95% of patients with Glioblastoma (GBM) show tumor relapse, leaving them with limited therapeutic options as recurrent tumors are most often resistant to the first line chemotherapy standard Temozolomide (TMZ). To identify molecular pathways involved in TMZ resistance, primary GBM Stem-like Cells (GSCs) were isolated, characterized, and selected for TMZ resistance in vitro. Subsequently, RNA sequencing analysis was performed and revealed a total of 49 differentially expressed genes (|log2-fold change| > 0.5 and adjusted p-value < 0.1) in TMZ resistant stem-like cells compared to their matched DMSO control cells. Among up-regulated genes, we identified carbonic anhydrase 2 (CA2) as a candidate gene correlated with glioma malignancy and patient survival. Notably, we describe consistent up-regulation of CA2 not only in TMZ resistant GSCs on mRNA and protein level, but also in patient-matched clinical samples of first manifest and recurrent tumors. Co-treatment with the carbonic anhydrase inhibitor Acetazolamid (ACZ) sensitized cells to TMZ induced cell death. Cumulatively, our findings illustrate the potential of CA2 as a chemosensitizing target in recurrent GBM and provide a rationale for a therapy associated inhibition of CA2 to overcome TMZ induced chemoresistance.

Highlights

Glioblastoma (GBM) is the most lethal brain tumor with a median survival of only 15 months [1].Despite an aggressive standard of care consisting of surgical resection, adjuvant radiation and chemotherapy with temozolomide (TMZ) [2,3] virtually all GBMs recur [4].Cancers 2019, 11, 921; doi:10.3390/cancers11070921 www.mdpi.com/journal/cancersGBM is characterized by a high degree of heterogeneity on phenotypic, genetic and cellular levels [5]
Among up-regulated genes, we identified carbonic anhydrase 2 (CA2) as a candidate gene correlated with glioma malignancy and patient survival
As other solid tumors GBM are composed of various brain resident as well as transformed cell types: there are rapidly multiplying tumor cells which make up the bulk of the tumor mass and, on the other hand, there are self-renewing cell types, often regarded as Glioblastoma stem cells (GSCs) [3,5,6]

Summary

Introduction

Glioblastoma (GBM) is the most lethal brain tumor with a median survival of only 15 months [1].Despite an aggressive standard of care consisting of surgical resection, adjuvant radiation and chemotherapy with temozolomide (TMZ) [2,3] virtually all GBMs recur [4].Cancers 2019, 11, 921; doi:10.3390/cancers11070921 www.mdpi.com/journal/cancersGBM is characterized by a high degree of heterogeneity on phenotypic, genetic and cellular levels [5]. Glioblastoma (GBM) is the most lethal brain tumor with a median survival of only 15 months [1]. Despite an aggressive standard of care consisting of surgical resection, adjuvant radiation and chemotherapy with temozolomide (TMZ) [2,3] virtually all GBMs recur [4]. As other solid tumors GBM are composed of various brain resident as well as transformed cell types: there are rapidly multiplying tumor cells which make up the bulk of the tumor mass and, on the other hand, there are self-renewing cell types, often regarded as Glioblastoma stem cells (GSCs) [3,5,6]. It is well established that GSC driven recurring tumors are resistant to further treatment, but the underlying molecular changes are not fully understood [10,11]

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Conclusion