An efficient method for the transduction of primary pediatric glioma neurospheres

Meel, Michaël H.; Metselaar, Dennis S.; Waranecki, Piotr; Kaspers, Gertjan J. L.; Hulleman, Esther

doi:10.1016/j.mex.2018.02.006

Cited by 16 publications

(16 citation statements)

References 16 publications

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“…The neurosphere stem cell culture is generally accepted as the gold standard for primary glioma stem cells [ 34 , 35 ]. Recently, two studies have employed a reliable protocol for lentiviral cell transduction of primary DIPG cell lines cultured as neurospheres, by exposing the cells to FBS for a short period of time [ 36 , 37 ]. They have demonstrated that the short-term exposure of the cells to serum and a rapid return to serum-free conditions, improve the neurosphere transduction efficiency without inducing a change in their stem cell proprieties.…”

Section: Discussionmentioning

confidence: 99%

Integration of Multiple Platforms for the Analysis of Multifluorescent Marking Technology Applied to Pediatric GBM and DIPG

Pericoli

Petrini

Giorda

et al. 2020

IJMS

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The intratumor heterogeneity represents one of the most difficult challenges for the development of effective therapies to treat pediatric glioblastoma (pGBM) and diffuse intrinsic pontine glioma (DIPG). These brain tumors are composed of heterogeneous cell subpopulations that coexist and cooperate to build a functional network responsible for their aggressive phenotype. Understanding the cellular and molecular mechanisms sustaining such network will be crucial for the identification of new therapeutic strategies. To study more in-depth these mechanisms, we sought to apply the Multifluorescent Marking Technology. We generated multifluorescent pGBM and DIPG bulk cell lines randomly expressing six different fluorescent proteins and from which we derived stable optical barcoded single cell-derived clones. In this study, we focused on the application of the Multifluorescent Marking Technology in 2D and 3D in vitro/ex vivo culture systems. We discuss how we integrated different multimodal fluorescence analysis platforms, identifying their strengths and limitations, to establish the tools that will enable further studies on the intratumor heterogeneity and interclonal interactions in pGBM and DIPG.

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Section: Discussionmentioning

confidence: 99%

Integration of Multiple Platforms for the Analysis of Multifluorescent Marking Technology Applied to Pediatric GBM and DIPG

Pericoli

Petrini

Giorda

et al. 2020

IJMS

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“…VUMC-HGG-05 [16] (giant-cell GBM), VUMC-HGG-09 (IDH1 R132H) and VUMC-HGG-14 are resection-derived glioblastomas. VUMC-HGG-11 [17] is a thalamic midline glioma harbouring a H3.3 K27M mutation. All models are FANCD2 wildtype as confirmed by whole-genome sequencing.…”

Section: Methodsmentioning

confidence: 99%

“…Cells were cultured as neurospheres as previously described [15,17,18]. Cells were only used when confirmed mycoplasm negative and short-tandem repeat (STR) analysis was performed to ensure cell line identities.…”

Section: Methodsmentioning

confidence: 99%

Celastrol-induced degradation of FANCD2 sensitizes pediatric high-grade gliomas to the DNA-crosslinking agent carboplatin

et al. 2019

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BackgroundPediatric high-grade gliomas (pHGG) are the leading cause of cancer-related death during childhood. Due to their diffuse growth characteristics, chemoresistance and location behind the blood-brain barrier (BBB), the prognosis of pHGG has barely improved in the past decades. As such, there is a dire need for new therapies that circumvent those difficulties. Since aberrant expression of DNA damage-response associated Fanconi anemia proteins play a central role in the onset and therapy resistance of many cancers, we here investigated if FANCD2 depletion could sensitize pHGG to additional DNA damage.MethodsWe determined the capacity of celastrol, a BBB-penetrable compound that degrades FANCD2, to sensitize glioma cells to the archetypical DNA-crosslinking agent carboplatin in vitro in seven patient-derived pHGG models. In addition, we tested this drug combination in vivo in a patient-derived orthotopic pHGG xenograft model. Underlying mechanisms to drug response were investigated using mRNA expression profiling, western blotting, immunofluorescence, FANCD2 knockdown and DNA fiber assays.FindingsFANCD2 is overexpressed in HGGs and depletion of FANCD2 by celastrol synergises with carboplatin to induce cytotoxicity. Combination therapy prolongs survival of pHGG-bearing mice over monotherapy and control groups in vivo (P<0.05). In addition, our results suggest that celastrol treatment stalls ongoing replication forks, causing sensitivity to DNA-crosslinking in FANCD2-dependent glioma cells.InterpretationOur results show that depletion of FANCD2 acts as a chemo-sensitizing strategy in pHGG. Combination therapy using celastrol and carboplatin might serve as a clinically relevant strategy for the treatment of pHGG.FundingThis study was funded by a grant from the Children Cancer-Free Foundation (KIKA, project 210). The disclosed funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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“…For the pediatric cells, there was an Institutional Review Board-approved protocol M-1608 from the Hospital Sant Joan de Deu Barcelona. Primary pediatric DIPG cells (HSJD-DIPG-007) were derived from autopsy of a 6-year-old female and were transferred as neurospheres, exponentially growing in suspension in tumorsphere media (TSM) supplemented with corresponding growth factors (15,16), and checked for mycoplasma contamination.…”

Section: Brain Tumor Specimenmentioning

confidence: 99%

OKlahoma Nitrone-007: Novel Treatment for Diffuse Intrinsic Pontine Glioma

Thomas

Smith

Saunders

et al. 2020

Preprint

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Background: Diffuse intrinsic pontine glioma (DIPG) is the most common brainstem cancer in childhood. This rapidly progressing brainstem glioma holds a very dismal prognosis with median survival of less than 1 year. Despite extensive research, no significant therapeutic advancements have been made to improve overall survival in DIPG patients. Methods: Here, we used an orthotopic xenograft pediatric DIPG (HSJD-DIPG-007) mouse model to monitor the effects of anti-cancer agent, OKlahoma Nitrone-007 (OKN-007), as an inhibitor of tumor growth after 28 days of treatment. Using magnetic resonance imaging (MRI), we confirmed the previously described efficacy of LDN-193189, a known activin A receptor, type I (ACVR1) inhibitor, in decreasing tumor burden and found that OKN-007 was equally efficacious.Results: After 28 days of treatment, the tumor volumes were significantly decreased in OKN-007 treated mice (p<0.01). The apparent diffusion coefficient (ADC), as a measure of tissue structural alterations, was significantly decreased in OKN-007 treated tumor-bearing mice (p<0.0001). Histological analysis also showed a significant decrease in CD34 expression, essential for angiogenesis, of OKN-007 treated mice (p<0.05) compared to LDN-193189 treated mice. OKN-007-treated mice also significantly decreased protein expression of the human nuclear antigen (HNA) (p<0.001), ACVR1 (p<0.0001), and c-MET (p<0.05), as well as significantly increased expression of cleaved caspase 3 (p<0.001) and histone H3 K27-trimethylation (p<0.01), compared to untreated mouse tumors.Conclusions: With the dismal prognosis and limited effective chemotherapy available for DIPG, there is significant room for continued research studies, and OKN-007 merits further exploration as a therapeutic agent.

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An efficient method for the transduction of primary pediatric glioma neurospheres

Abstract: Graphical abstract

Cited by 16 publications

References 16 publications

Integration of Multiple Platforms for the Analysis of Multifluorescent Marking Technology Applied to Pediatric GBM and DIPG

Integration of Multiple Platforms for the Analysis of Multifluorescent Marking Technology Applied to Pediatric GBM and DIPG

Celastrol-induced degradation of FANCD2 sensitizes pediatric high-grade gliomas to the DNA-crosslinking agent carboplatin

OKlahoma Nitrone-007: Novel Treatment for Diffuse Intrinsic Pontine Glioma

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