MYCN sensitizes neuroblastoma to the MDM2-p53 antagonists Nutlin-3 and MI-63

Abstract

MYCN amplification is a major biomarker of poor prognosis, occurring in 25-30% of neuroblastomas. MYCN plays contradictory roles in promoting cell growth and sensitizing cells to apoptosis. We have recently shown that p53 is a direct transcriptional target of MYCN in neuroblastoma and that p53-mediated apoptosis may be an important mechanism of MYCN-induced apoptosis. Although p53 mutations are rare in neuroblastoma at diagnosis, the p53/MDM2/p14ARF pathway is often inactivated through MDM2 amplification or p14ARF inactivation. We hypothesised that reactivation of p53 by inhibition of its negative regulator MDM2, using the MDM2-p53 antagonists Nutlin-3 and MI-63, will result in p53-mediated growth arrest and apoptosis especially in MYCN amplified cells. Using the SHEP Tet21N MYCN regulatable system, MYCN(−) cells were more resistant to both Nutlin-3 and MI-63 mediated growth inhibition and apoptosis compared to MYCN(+) cells and siRNA mediated knockdown of MYCN in 4 MYCN amplified cell lines resulted in decreased p53 expression and activation, as well as decreased levels of apoptosis following treatment with MDM2-p53 antagonists. In a panel of 18 neuroblastoma cell lines treated with Nutlin-3 and MI-63, the sub-set amplified for MYCN had a significantly lower mean GI50 value and increased caspase 3/7 activity compared to the non MYCN amplified group of cell lines, but p53 mutant cell lines were resistant to the antagonists regardless of MYCN status. We conclude that amplification or overexpression of MYCN sensitizes neuroblastoma cell lines with wildtype p53 to MDM2-p53 antagonists and that these compounds may therefore be particularly effective in treating high risk MYCN amplified disease.

Massive parallel sequencing and digital gene expression analysis reveals potential mechanisms to overcome therapy resistance in pulmonary neuroendocrine tumors

“…Numerous environmental influences and genetic alterations induced by platin compounds are able to activate P53 by post-translational modifications, which results in cell cycle arrest, cellular senescence, or apoptosis 47. Nutlin-3A, a cis-imidazoline analogue, is a potent and selective MDM2 inhibitor 48, 49 that prevents MDM2-TP53-interaction by binding to the hydrophobic binding pocket of MDM2 leading to an immediate reactivation of P53 48, 50, 51. It is currently tested in a phase I clinical trial (NCT01143519, NCT00623870) 51.…”

p53 Loss in MYC-Driven Neuroblastoma Leads to Metabolic Adaptations Supporting Radioresistance

“…While direct mutations to p53 are a rare event in de novo cases of neuroblastoma (6), examination of the p53 pathway has underlined its role in the pathogenesis of high-risk neuroblastoma (45)(46)(47)(48)(49)(50). The role of p53 loss of function in the pathogenesis of untreated neuroblastomas has thus far not been elucidated.…”

Outcome of the p53-mediated DNA damage response in neuroblastoma is determined by morphological subtype and MYCN expression

“…We also have recent data showing that S-type cell lines are more likely to G 1 arrest following MDM2/p53 inhibitor treatment and less likely to undergo apoptosis. 35 N-type cells undergo increased apoptosis after DNA damage, while expressing high levels of the anti-apoptotic protein BCL-2. Although this appears contradictory, BCL-2 is just one of many factors in determining cellular apoptosis.…”