Stem cell cultures derived from pediatric brain tumors accurately model the originating tumors

Abstract

Brain tumors are the leading cause of cancer-related death in children but high-grade gliomas in children and adolescents have remained a relatively under-investigated disease despite this. A better understanding of the cellular and molecular pathogenesis of the diseases is required in order to improve the outcome for these children. In vitro-cultured primary tumor cells from patients are indispensable tools for this purpose by enabling functional analyses and development of new therapies. However, relevant well-characterized in vitro cultures from pediatric gliomas cultured under serum-free conditions have been lacking. We have therefore established patient-derived in vitro cultures and performed thorough characterization of the cells using large-scale analyses of DNA methylation, copy-number alterations and investigated their stability during prolonged time in culture. We show that the cells were stable during prolonged culture in serum-free stem cell media without apparent alterations in morphology or growth rate. The cells were proliferative, positive for stem cell markers, able to respond to differentiation cues and initiated tumors in zebrafish and mice suggesting that the cells are cancer stem cells or progenitor cells. The cells accurately mirrored the tumor they were derived from in terms of methylation pattern, copy number alterations and DNA mutations. These unique primary in vitro cultures can thus be used as a relevant and robust model system for functional studies on pediatric brain tumors.

Accumulation of DNA methylation alterations in paediatric glioma stem cells following fractionated dose irradiation

“…In this study, we profiled DNA methylation alterations that follow after few (n = 3) and many (n = 15 or 15 + 3) fractions of radiation. For this, we used previously generated GSC cultures [12] established from patients before (n = 4) and after treatment (n = 1). These wellcharacterised cells have been shown to be stable during culturing and maintain their DNA methylation profiles and DNA copy number alterations (CNA).…”

“…All patients received conventional doses of 1.8 Gy in daily fractions (5 days a week) and a total dose of 54 Gy as part of their clinical management. Patients, tumours and cell cultures have been fully described previously [12]. Samples were collected with written informed consent according to the ethical approval (Dnr 604-12).…”

“…Identifying targets and dynamics of DNA methylation alterations following radiation has the potential to reveal biological consequences. For this, we used patientderived paediatric high-grade glioma stem cells (GSCs) grown under conditions that preserve the signatures of the originating tumour and maintain the tumourinitiating capacity of the cells [12]. Genome-wide DNA methylation maps of pre-and post-irradiated GSC cultures were generated to determine the effects radiation has on epigenomic plasticity and stability.…”

Hypoxic Environment and Paired Hierarchical 3D and 2D Models of Pediatric H3.3-Mutated Gliomas Recreate the Patient Tumor Complexity

“…In addition, for drug testing, most of the studies in the literature focused on 3D/NS PDCLs, which were considered the gold standard cultures to preserve stem-like features [31] and resistance to therapy. Some other publications supported an equivalence of 2D and 3D cultures for modeling drug response [17,32].…”

Zebrafish in Translational Cancer Research: Insight into Leukemia, Melanoma, Glioma and Endocrine Tumor Biology

“…To investigate the presence of tumor-initiating cells in stem cells cultures derived from high-grade pediatric brain tumors, cells were transplanted into zebrafish and mice [ 129 ]. Implanted cells were able to initiate tumors showing stemness properties.…”