Oncotarget

Research Papers:

Low-dose Actinomycin-D treatment re-establishes the tumoursuppressive function of P53 in RELA-positive ependymoma

Theophilos Tzaridis, Till Milde, Kristian W. Pajtler, Sebastian Bender, David T. W. Jones, Simone Müller, Andrea Wittmann, Magdalena Schlotter, Andreas E. Kulozik, Peter Lichter, V. Peter Collins, Olaf Witt, Marcel Kool, Andrey Korshunov, Stefan M. Pfister and Hendrik Witt _

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Oncotarget. 2016; 7:61860-61873. https://doi.org/10.18632/oncotarget.11452

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Abstract

Theophilos Tzaridis1, Till Milde 2,3,4, Kristian W. Pajtler1,2,4, Sebastian Bender1, David T. W. Jones1, Simone Müller5, Andrea Wittmann1, Magdalena Schlotter6, Andreas E. Kulozik2, Peter Lichter6, V. Peter Collins7, Olaf Witt2,3,4, Marcel Kool1, Andrey Korshunov8,9, Stefan M. Pfister1,2,4, Hendrik Witt1,2,4

1Division of Pediatric Neurooncology (B062), German Cancer Consortium (DKTK), German Cancer Research Center (DKFZ), Heidelberg, Germany

2Department of Pediatric Oncology, Hematology and Immunology, Children’s Hospital, University of Heidelberg, Heidelberg, Germany

3Clinical Cooperation Unit Pediatric Oncology (G340), German Cancer Research Center (DKFZ), Heidelberg, Germany

4National Center for Tumor diseases (NCT), Clinical Trial Center, Heidelberg, Germany

5Department of Cardiology, Angiology and Pulmology, University of Heidelberg, Heidelberg, Germany

6Division of Molecular Genetics (B060), German Cancer Research Center (DKFZ) and German Cancer Consortium (DKTK), Heidelberg, Germany

7Division of Molecular Histopathology, Department of Pathology, University of Cambridge, Cambridge UK

8Clinical Cooperation Unit Neuropathology (G380), German Cancer Research Center (DKFZ) and German Cancer Consortium (DKTK), Heidelberg, Germany

9Department of Neuropathology, Heidelberg University Hospital, Heidelberg, Germany

Correspondence to:

Hendrik Witt, email: [email protected]

Stefan M. Pfister, email: [email protected]

Keywords: ependymoma, p53, RELA, Actinomycin-D

Received: February 02, 2016     Accepted: July 27, 2016     Published: August 20, 2016

ABSTRACT

Ependymomas in children can arise throughout all compartments of the central nervous system (CNS). Highly malignant paediatric ependymoma subtypes are Group A tumours of the posterior fossa (PF-EPN-A) and RELA-fusion positive (ST-EPN-RELA) tumours in the supratentorial compartment. It was repeatedly reported in smaller series that accumulation of p53 is frequently observed in ependymomas and that immunohistochemical staining correlates with poor clinical outcome, while TP53 mutations are rare. Our TP53 mutation analysis of 130 primary ependymomas identified a mutation rate of only 3%. Immunohistochemical analysis of 398 ependymomas confirmed previous results correlating the accumulation of p53 with inferior outcome. Among the p53-positive ependymomas, the vast majority exhibited a RELA fusion leading to the hypothesis that p53 inactivation might be linked to RELA positivity.

In order to assess the potential of p53 reactivation through MDM2 inhibition in ependymoma, we evaluated the effects of Actinomycin-D and Nutlin-3 treatment in two preclinical ependymoma models representing the high-risk subtypes PF-EPN-A and ST-EPN-RELA. The IC-50 of the agent as determined by metabolic activity assays was in the lower nano-molar range (0.2–0.7 nM). Transcriptome analyses of high-dose (100 nM), low-dose (5 nM) and non-treated cells revealed re-expression of p53 dependent genes including p53 upregulated modulator of apoptosis (PUMA) after low-dose treatment. At the protein level, we validated the Actinomycin-D induced upregulation of PUMA, and of p53 interaction partners MDM2 and p21. Proapoptotic effects of low-dose application of the agent were confirmed by flow cytometry. Thus, Actinomycin-D could constitute a promising therapeutic option for ST-EPN-RELA ependymoma patients, whose tumours frequently exhibit p53 inactivation.


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