Selective DNA-PKcs inhibition extends the therapeutic index of localized radiotherapy and chemotherapy
Catherine E. Willoughby, … , Anderson J. Ryan, Stephen R. Wedge
Catherine E. Willoughby, … , Anderson J. Ryan, Stephen R. Wedge
Published October 3, 2019
Citation Information: J Clin Invest. 2020;130(1):258-271. https://doi.org/10.1172/JCI127483.
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Research Article Oncology Therapeutics

Selective DNA-PKcs inhibition extends the therapeutic index of localized radiotherapy and chemotherapy

Abstract

Potentiating radiotherapy and chemotherapy by inhibiting DNA damage repair is proposed as a therapeutic strategy to improve outcomes for patients with solid tumors. However, this approach risks enhancing normal tissue toxicity as much as tumor toxicity, thereby limiting its translational impact. Using NU5455, a newly identified highly selective oral inhibitor of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) activity, we found that it was indeed possible to preferentially augment the effect of targeted radiotherapy on human orthotopic lung tumors without influencing acute DNA damage or a late radiation-induced toxicity (fibrosis) to normal mouse lung. Furthermore, while NU5455 administration increased both the efficacy and the toxicity of a parenterally administered topoisomerase inhibitor, it enhanced the activity of doxorubicin released locally in liver tumor xenografts without inducing any adverse effect. This strategy is particularly relevant to hepatocellular cancer, which is treated clinically with localized drug-eluting beads and for which DNA-PKcs activity is reported to confer resistance to treatment. We conclude that transient pharmacological inhibition of DNA-PKcs activity is effective and tolerable when combined with localized DNA-damaging therapies and thus has promising clinical potential.

Authors

Catherine E. Willoughby, Yanyan Jiang, Huw D. Thomas, Elaine Willmore, Suzanne Kyle, Anita Wittner, Nicole Phillips, Yan Zhao, Susan J. Tudhope, Lisa Prendergast, Gesa Junge, Luiza Madia Lourenco, M. Raymond V. Finlay, Paul Turner, Joanne M. Munck, Roger J. Griffin, Tommy Rennison, James Pickles, Celine Cano, David R. Newell, Helen L. Reeves, Anderson J. Ryan, Stephen R. Wedge

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Figure 7

NU5455 augments localized doxorubicin chemotherapy in HCC tumor xenografts without increased toxicity.

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NU5455 augments localized doxorubicin chemotherapy in HCC tumor xenograf...
Exponentially growing Huh7 cells in 6-well plates were treated with NU5455 (1 μM) or DMSO for 1 hour before the addition of a single doxorubicin-loaded 70- to 150-μm DC M1 bead (25 mg/mL) to the center of each well. (A and B) Cell density was determined after 96 hours of continuous treatment using sulforhodamine B staining (A), and protein extracts were taken for the detection of γH2AX via Western blotting after 24–96 hours of continuous treatment (n = 3) (B). (C) Mice bearing established Huh7 subcutaneous xenografts (180–220 mm3) were treated with 5 μL DC M1 beads loaded with 25 mg/mL doxorubicin, or unloaded beads. Beads were suspended in 30 μL saline and implanted via an intratumoral injection. Twice-daily dosing with NU5455 (30 mg/kg orally, 9-hour dosing interval) or vehicle control was commenced 60 minutes after bead implantation. (D) Mean Huh7 tumor volume over time (6 mice per group). (E) Percentage body weight at day 8 relative to pretreatment body weight on day 1. (F) Percentage phospho–DNA-PKcs Ser2056 nuclear positivity in Huh7 tumor sections, and high (grade III) γH2AX nuclear positivity in a 400-μm radius surrounding individually embedded beads (5 mice per group). Representative images of phospho–DNA-PKcs Ser2056 and γH2AX staining in Huh7 tumor xenografts treated with unloaded or doxorubicin-loaded DC M1 beads. Scale bars: 200 μm. All graphs represent the mean ± SEM. Statistical significance was assessed using 1-way ANOVA (A, E, and F) and unpaired t test (D). **P < 0.01, ***P < 0.001, ****P < 0.0001.