Abstract 5384: Prolonged survival of the TH-MYCN murine model of high-risk neuroblastoma, in response to the histone deacetylase inhibitor panobinostat, is the result of both apoptosis and differentiation
Authors: Waldeck Kelly , Cullinane Carleen , Shortt Jake , Martin Ben , Ardley Kerry , Tothill Richard , McArthur Grant , Wood Paul J. 
Published: August 2015

Background/Aims: Deregulated acetylation of histones plays a key role in the pathogenesis of hematological as well as solid tumors by changing the chromatin structure and consequently altering transcription of genes involved in cell cycle control, differentiation and apoptosis. Inhibitors of histone deacetylases (HDACs) can therefore result in multiple cellular responses, including cell death and differentiation. The aim of this study was to investigate the mechanism of major prolongation in survival induced by the HDAC-inhibitor panobinostat, in the TH-MYCN mouse model for neuroblastoma,

Methods: Mice homozygous for the TH-MYCN transgene underwent abdominal ultrasounds (US) from five weeks of age until neuroblastomas of between 50mm3 and 200 mm3 were detected. Mice were then randomized to receive the pan-HDAC inhibitor, panobinostat or vehicle daily for nine weeks. Tumor volume was monitored by serial US. One hundred days following cessation of therapy tumors were harvested for immunohistochemical (IHC) analysis of key markers for differentiation. For short-term experiments, tumors were harvested after 24hr for western blot and IHC analysis of key proteins of the apoptosis pathways as well as biomarkers of HDAC inhibition. RNA-seq analysis was performed on tumors harvested at baseline, 14 hr and 24 hr post panobinostat treatment. Gene set enrichment, and pathway analysis was then performed to investigate changes in the expression of genes associated with apoptosis, differentiation and NMYC expression.

Results: Treatment with panobinostat significantly improved survival with 100% of TH-MYCN mice alive at day 63 compared with vehicle (0.0%, mean survival 7 days; p&lt;0.0001). One hundred days after the withdrawal of drug 88.9% of panobinostat treated mice remained alive (p&lt;0.0001) with no evidence of disease progression on US. Panobinostat induced rapid tumor regression as determined by US, and significant apoptosis mediated through the caspase-dependent pathway via up-regulation of the BH3 only pro-apoptotic proteins, BMF and BIM. Panobinostat also induced differentiation of malignant neuroblastomas into benign ganglioneuromas as evidenced by H&E and IHC analysis of markers of differentiation including S100 and SSTR2. Correspondingly tumors had reduced expression of NMYC compared with untreated tumors. The findings confirmed up-regulation of genes associated with apoptosis and differentiation, as well as down-regulation of NMYC expression.

Conclusions: Treatment of TH-MYCN mice with continuous panobinostat significantly improved survival and reduced tumor burden, primarily via induction of apoptosis and differentiation. These preclinical findings indicate that HDAC inhibition is a promising therapeutic strategy, and supports the further study of panobinostat as a treatment option for high-risk neuroblastoma.

Citation Format: Kelly Waldeck, Carleen Cullinane, Jake Shortt, Ben Martin, Kerry Ardley, Richard Tothill, Grant McArthur, Paul J. Wood. Prolonged survival of the TH-MYCN murine model of high-risk neuroblastoma, in response to the histone deacetylase inhibitor panobinostat, is the result of both apoptosis and differentiation. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5384. doi:10.1158/1538-7445.AM2015-5384