Innovative Thiosemicarbazones Down-Regulate N-Myc Expression in Neuroblastoma Cells via Transcriptional and Post-Transcriptional Mechanisms

Neuroblastoma (NB) is an aggressive solid childhood tumor. A key oncogenic effector in NB is the transcription factor, N-Myc, whose genetic amplification leads to poor prognosis. Previous studies have indicated that iron-binding ligands such as desferrioxamine (DFO) effectively down-regulate N-myc expression. However, the mechanism of how this occurs remains unknown and could provide clues to elucidating potential therapeutic interventions. Herein, we mechanistically dissect the activity of a new class of iron-binding ligands that have entered clinical trials and show potent anti-proliferative activity against NB, namely the thiosemicarbazone, DpC. This was achieved using RNA sequencing (RNA-seq), where we have examined the effect of these ligands on gene expression in 3 NB cell-types with (Kelly and BE2-C) or without (SH-SY5Y) N-myc amplification. RNA-seq revealed that DpC mimicked the effect of DFO on gene expression at a 20-fold lower concentration, up-regulating key N-Myc targets such as the metastasis suppressor NDRG1, which translocates to the nucleus upon treatment. DFO and DpC significantly reduced N-Myc mRNA in p53-mutant NB cells (Kelly, BE(2)-C). In contrast, DFO increased N-Myc mRNA in wild-type p53 SH-SY5Y NB cells, where it was demonstrated that MDM2 stabilized N-Myc mRNA.