CT-2A neurospheres-derived high-grade glioma in mice: a new model to address tumor stem cells and immunosuppression

Matteo Riva,Uwe Himmelreich,Sarah Belderbos,Akila Weerasekera,Thaïs Baert,Marc Van Ranst,Roxanne Wouters,Dietmar R Thal,Willy Gsell,David Nittner,Roberto Giovannoni,An Coosemans

doi:10.1242/bio.044552

Abstract

ABSTRACTRecently, several promising treatments for high-grade gliomas (HGGs) failed to provide significant benefit when translated from the preclinical setting to patients. Improving the animal models is fundamental to overcoming this translational gap. To address this need, we developed and comprehensively characterized a new in vivo model based on the orthotopic implantation of CT-2A cells cultured in neurospheres (NS/CT-2A). Murine CT-2A methylcholanthrene-induced HGG cells (C57BL/6 background) were cultured in monolayers (ML) or NS and orthotopically inoculated in syngeneic animals. ML/CT-2A and NS/CT-2A tumors' characterization included the analysis of tumor growth, immune microenvironment, glioma stem cells (GSCs), vascularization and metabolites. The immuno-modulating properties of NS/CT-2A and ML/CT-2A cells on splenocytes were tested in vitro. Mice harboring NS/CT-2A tumors had a shorter survival than those harboring ML/CT-2A tumors (P=0.0033). Compared to standard ML/CT-2A tumors, NS/CT-2A tumors showed more abundant GSCs (P=0.0002 and 0.0770 for Nestin and CD133, respectively) and regulatory T cells (Tregs, P=0.0074), and a strong tendency towards an increased vascularization (P=0.0503). There were no significant differences in metabolites' composition between NS/ and ML/CT-2A tumors. In vitro, NS were able to drive splenocytes towards a more immunosuppressive status by reducing CD8+ T cells (P=0.0354) and by promoting Tregs (P=0.0082), macrophages (MF, P=0.0019) and their M2 subset (P=0.0536). Compared to standard ML/CT-2A tumors, NS/CT-2A tumors show a more aggressive phenotype with increased immunosuppression and GSCs proliferation. Because of these specific features, the NS/CT-2A model represents a clinically relevant platform in the search for new HGG treatments aimed at reducing immunosuppression and eliminating GSCs.

Highlights

High-grade gliomas (HGGs) are aggressive brain tumors
CT-2A NS promote glioma stem cells (GSCs) and tumor vasculature We evaluated the presence of GSCs within the tumor by staining the brains for Nestin and CD133
Several preclinical studies addressed the role of GSCs in HGGs; most of these studies were based on deriving GSCs from human samples and re-implanting them in immune-deficient xenografts (Audia et al, 2017)

Summary

Introduction

High-grade gliomas (HGGs) are aggressive brain tumors. With current standard therapies (surgery and chemo-radiation), the median survival of glioblastoma (GBM, the most malignant subtype of HGG) is only 15 months, relapse is almost universal and the large majority of patients die of the disease (Ostrom et al, 2014). During the last few decades, a vast amount of research has been carried out and the knowledge of HGG biology has tremendously improved. This has led to the development of several innovative treatments, such as immunotherapy or oncolytic virotherapy; despite very promising preclinical results, all these therapies failed to provide significant survival benefits when administered to patients (Dunn-Pirio and Vlahovic, 2017; Filley et al, 2017; Luksik et al, 2017).

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