Characterisation of the p53 pathway in cell lines established from TH-MYCN transgenic mouse tumours

Chen,Lindi; Esfandiari,Arman; Reaves,William; Vu,Annette; Hogarty,Michael  D.; Lunec,John; Tweddle,Deborah  A.

doi:10.3892/ijo.2018.4261

Characterisation of the p53 pathway in cell lines established from TH-MYCN transgenic mouse tumours

Authors:
- Lindi Chen
- Arman Esfandiari
- William Reaves
- Annette Vu
- Michael D. Hogarty
- John Lunec
- Deborah A. Tweddle
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Affiliations: Wolfson Childhood Cancer Research Centre, Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne NE1 7RU, UK, Newcastle Cancer Centre, Northern Institute for Cancer Research, Newcastle University, Newcastle upon Tyne NE1 7RU, UK, The Children's Hospital of Philadelphia, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104, USA
Published online on: January 31, 2018 https://doi.org/10.3892/ijo.2018.4261
Pages: 967-977

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Abstract

Cell lines established from the TH-MYCN transgenic murine model of neuroblastoma are a valuable preclinical, immunocompetent, syngeneic model of neuroblastoma, for which knowledge of their p53 pathway status is important. In this study, the Trp53 status and functional response to Nutlin-3 and ionising radiation (IR) were determined in 6 adherent TH-MYCN transgenic cell lines using Sanger sequencing, western blot analysis and flow cytometry. Sensitivity to structurally diverse MDM2 inhibitors (Nutlin-3, MI-63, RG7388 and NDD0005) was determined using XTT proliferation assays. In total, 2/6 cell lines were Trp53 homozygous mutant (NHO2A and 844MYCN+/+) and 1/6 (282MYCN+/-) was Trp53 heterozygous mutant. For 1/6 cell lines (NHO2A), DNA from the corresponding primary tumour was found to be Trp53 wt. In all cases, the presence of a mutation was consistent with aberrant p53 signalling in response to Nutlin-3 and IR. In comparison to TP53 wt human neuroblastoma cells, Trp53 wt murine control and TH-MYCN cell lines were significantly less sensitive to growth inhibition mediated by MI-63 and RG7388. These murine Trp53 wt and mutant TH-MYCN cell lines are useful syngeneic, immunocompetent neuroblastoma models, the former to test p53-dependent therapies in combination with immunotherapies, such as anti-GD2, and the latter as models of chemoresistant relapsed neuroblastoma when aberrations in the p53 pathway are more common. The spontaneous development of Trp53 mutations in 3 cell lines from TH-MYCN mice may have arisen from MYCN oncogenic driven and/or ex vivo selection. The identified species-dependent selectivity of MI-63 and RG7388 should be considered when interpreting in vivo toxicity studies of MDM2 inhibitors.

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