Tumour initiation in the TH-MYCN transgenic mouse model of neuroblastoma is characterised by transient repression of p53 stress responses in embryonal ganglionic neuroblasts, causing them to persist postnatally as cancer-prone lesions 1, 2. p53 re-activation may be a potential therapeutic strategy for removal of these lesions before they later transform in response to oncogenic stimuli. One potential target for p53 activation is the FACT protein complex which is known to inhibit p53 signalling. Here we show that high mRNA expression levels of the two FACT subunits, SSRP1 and SPT16, in human primary neuroblastoma tissue were significantly associated with poor patient outcome (p<0.001) on Kaplan Meier analysis of 650 neuroblastoma patients. Both SSRP1 and SPT16 levels had a strong correlation with the level of MYCN expression, particularly in high risk MYCN-amplified neuroblastoma tissues (p<0.05). Inhibition of MYCN expression by a specific siRNA, lowered SSRP1 and SPT16 expression in two neuroblastoma tumor cell lines (BE(2)C and Kelly). Alamar blue assays revealed that a chemical inhibitor of FACT, Curaxin or CBL137, potently and selectively promoted cytotoxicity in neuroblastoma cell lines as compared to non-malignant MRC5 fibroblasts (p<0.001). To examine the role of FACT in MYCN-driven tumourigenesis we evaluated the effect of Curaxin on tumor initiation in TH-MYCN mice. Low micromolar Curaxin doses effectively restored normal in vitro, p53-dependent death responses to primary ganglion cells deprived of serum from neonatal TH-MYCN mice, while ganglion cultures from wild-type littermate mice were significantly more resistant to Curaxin treatment, with or without serum deprivation (p<0.001). Most importantly, prophylactic Curaxin treatment of 6 day old TH-MYCN mice had a marked inhibitory effect on postnatal ganglion neuroblast persistence in vivo for both hemizygote and homozygote TH-MYCN mice, as compared to vehicle-only treated animals (p=0.01, p=0.035, respectively). Taken together, our data for the first time identifies FACT as a novel effector of MYCN-driven neuroblastoma tumorigenesis. Inhibitors of the FACT complex may be a novel treatment for high risk MYCN-amplified neuroblastoma and a potential strategy for neuroblastoma prevention.
1. Hansford, L. et al. Mechanisms of embryonal tumor initiation: Distinct roles for MycN expression and MYCN amplification. Proceedings of the National Academy of Sciences of the United States of America 101, 12664-12669 (2004).
2. Calao, M. et al. Direct effects of Bmi1 on p53 protein stability inactivates oncoprotein stress responses in embryonal cancer precursor cells at tumor initiation. Oncogene [ahead of print]
Citation Format: Daniel R. Carter, Belamy B. Cheung, Tao Liu, Carol Au, Murray D. Norris, Michelle Haber, Jayne Murray, Katerina V. Gurova, Andrei V. Gudkov, Glenn M. Marshall. The facilitates chromatin transcription (FACT) protein complex promotes neuroblastoma tumor initiation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5043. doi:10.1158/1538-7445.AM2013-5043