Targeting The Polyamine-Hypusine Nexus For The Treatment Of Neuroblastoma

Abstract The highly conserved and unique post-translational hypusine modification is essential for the activation of a single cellular protein called eukaryotic initiation factor 5A (eIF5A). Hypusination of eIF5A represents an essential mechanism in the control of cell proliferation and has been linked to cancer. The hypusine modification of eIF5A requires the enzymes deoxyhypusine synthase (DHPS) and deoxyhypusine hydroxylase (DOHH) and, importantly, uniquely depends on the polyamine spermidine as a selective substrate. Polyamines including spermidine are frequently dysregulated in neoplastic disease and are critical for cell proliferation. Ornithine decarboxylase (ODC) is the sentinel enzyme in polyamine biosynthesis. We pioneered the repurposing of the irreversible ODC inhibitor alpha-difluoromethylornithine (DFMO) for neuroblastoma (NB), a devastating pediatric cancer of the sympathetic nervous system. Our preclinical work was confirmed by other groups and spurred two consortia (NMTRC and NANT) to test DFMO in independent phase I/II NB clinical trials. The oncogene MYCN is commonly amplified in high-risk NB patients and predicts poor prognosis. MYCN directly activates ODC (and eIF5A) and drives NB tumor progression, presumably through the activation of polyamines, but the precise molecular mechanisms are not fully understood. Since spermidine is the only molecule that can activate eI5FA through hypusine modification, we hypothesized that the polyamine-hypusine nexus provides a rational target site to identify new drug combinations that trigger synergistic anti-tumor effects. In this study we found that DFMO combined with DHPS inhibitor GC7 induced synergistic drug reactions in NB at concentrations that are not lethal on their own. While each drug alone at higher concentrations induced p27/Rb-mediated G1/S cell cycle arrest, the combination of both drugs resulted in the induction of programmed cell death (apoptosis). The intracellular free polyamine levels (putrescine, spermidine, and spermine) were quantified by C-18 RP-HPLC and responded directly to drug treatments, suggesting specific drug targeting effects. This dual-pronged drug combination approach that specifically targets the blockade of the unique polyamine/spermidine-dependent hypusine-eIF5A signaling circuit known to be involved in cell proliferation might lead to more effective DFMO treatment options by reducing high DFMO doses currently needed while achieving better therapeutic outcomes in NB. Citation Format: Andre S. Bachmann, Raid El-Khawaja, Chad Schultz. Targeting the polyamine-hypusine nexus for the treatment of neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5828. doi:10.1158/1538-7445.AM2017-5828