Abstract 1108: A novel, selective inhibitor of DNA-dependent protein kinase (DNA-PK) potentiates the effects of DNA-damaging therapies in hepatocellular carcinoma
Authors: Willoughby Catherine E. , Thomas Huw D. , Rennison Tommy , Cano Celine , Reeves Helen L. , Wedge Stephen R. 
Published: July 2017

DNA-dependent protein kinase (DNA-PK) is a key component in the repair of DNA double-strand breaks via non-homologous end-joining. Studies have found elevated DNA-PK expression and activity in hepatocellular carcinoma (HCC) to be strongly correlated with increased tumor grade, resistance to DNA-damaging therapies and poor survival (1,2).

We have explored the selective inhibition of DNA-PK in combination with DNA-damaging agents as a potential therapeutic approach in HCC using NDD0004 - a novel, orally-bioavailable small molecule inhibitor of DNA-PK (in vitro IC50 = 8 nM).

NDD0004 was evaluated in a panel of DNA-PK overexpressing human HCC cell lines (Hep3B, HepG2, Huh7) in combination with ionizing radiation or the topoisomerase II poison doxorubicin. DNA-PK activity was determined by Ser2056 phosphorylation status, DNA damage quantified by γH2AX levels, cell proliferation determined by SRB assays and cell survival assessed using clonogenic assays. In vivo efficacy was evaluated using a novel murine model of localized and sustained doxorubicin therapy, involving intra-tumoral injection of doxorubicin-loaded DC M1 polymer beads into established Huh7 human HCC xenografts in CD1 nude mice. Oral twice-daily treatment with 30 mg/kg NDD0004 or vehicle control was commenced 1 hour following bead implantation and continued for up to 20 days (6 mice per group).

NDD0004 dose-dependently inhibited activation of DNA-PK in HCC cell lines in vitro in response to ionizing radiation, and significantly increased and sustained DNA damage following treatment. Furthermore, NDD0004 sensitized DNA-PK overexpressing HCC cell lines to doxorubicin and ionizing radiation in proliferation and survival assays by ≥ 5-fold. Combining NDD0004 with doxorubicin-loaded beads in vivo significantly inhibited the rapid growth of HCC tumors when compared to treatment with doxorubicin-loaded bead monotherapy, with the time taken for tumor volumes to quadruple (RTV4) being extended from 11 to 18 days (P&lt;0.01, Mann-Whitney). Individual and combination treatments were well tolerated throughout. Immuno-histochemical analysis revealed γH2AX levels in hepatocytes surrounding doxorubicin-loaded beads to be significantly increased by NDD0004 co-treatment for 72-hours (P&lt;0.02, 2-way ANOVA).

In conclusion, selective inhibition of DNA-PK catalytic activity sensitized DNA-PK overexpressing HCC cell lines to doxorubicin chemotherapy and ionizing radiation in vitro, and augmented the anti-tumor activity of localized chemotherapy in HCC xenografts in vivo. These data support the concept of combining a DNA-PK inhibitor with localized DNA-damaging therapies in the treatment of HCC.

1. Cornell et al. (2015) Clin Cancer Res. 21: 925-33

2. Evert et al. (2013) Br J Cancer. 109: 2654-64

Citation Format: Catherine E. Willoughby, Huw D. Thomas, Tommy Rennison, Celine Cano, Helen L. Reeves, Stephen R. Wedge. A novel, selective inhibitor of DNA-dependent protein kinase (DNA-PK) potentiates the effects of DNA-damaging therapies in hepatocellular carcinoma [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 1108. doi:10.1158/1538-7445.AM2017-1108