Abstract
Radiotherapy is based on the induction of lethal DNA damage, primarily DNA double-strand breaks (DSB). Efficient DSB repair via Non-Homologous End Joining or Homologous Recombination can therefore undermine the efficacy of radiotherapy. By suppressing DNA-DSB repair with hyperthermia (HT) and DNA-PKcs inhibitor NU7441 (DNA-PKcsi), we aim to enhance the effect of radiation.The sensitizing effect of HT for 1 hour at 42°C and DNA-PKcsi [1 μM] to radiation treatment was investigated in cervical and breast cancer cells, primary breast cancer sphere cells (BCSCs) enriched for cancer stem cells, and in an in vivo human tumor model. A significant radio-enhancement effect was observed for all cell types when DNA-PKcsi and HT were applied separately, and when both were combined, HT and DNA-PKcsi enhanced radio-sensitivity to an even greater extent. Strikingly, combined treatment resulted in significantly lower survival rates, 2 to 2.5 fold increase in apoptosis, more residual DNA-DSB 6 h post treatment and a G2-phase arrest. In addition, tumor growth analysis in vivo showed significant reduction in tumor growth and elevated caspase-3 activity when radiation was combined with HT and DNA-PKcsi compared to radiation alone. Importantly, no toxic side effects of HT or DNA-PKcsi were found.In conclusion, inhibiting DNA-DSB repair using HT and DNA-PKcsi before radiotherapy leads to enhanced cytotoxicity in cancer cells. This effect was even noticed in the more radio-resistant BCSCs, which are clearly sensitized by combined treatment. Therefore, the addition of HT and DNA-PKcsi to conventional radiotherapy is promising and might contribute to more efficient tumor control and patient outcome.
Highlights
The underlying mechanism of many anti-cancer treatments, including ionizing radiation, is the induction of lethal DNA double strand breaks (DSB) [1, 2]
Limiting dilution analysis with breast cancer sphere cells (BCSCs) that are enriched for cancer stem cells (Figure 1E) showed a significant decrease in clonogenic growth after radiation, and this decrease is further enhanced by HT or DNAPKcsi
This study investigated the additional effects of DNA-double-strand breaks (DSB) repair inhibitors when added to conventional radiotherapy
Summary
The underlying mechanism of many anti-cancer treatments, including ionizing radiation, is the induction of lethal DNA double strand breaks (DSB) [1, 2]. The more rapidly dividing tumor cells are thought to be more sensitive to ionizing radiation healthy cells, and their subsequent DNA damage response less efficient [2]. Some tumor cells are thought to be less sensitive to radiation treatment [3], i.e. cancer stem cells, which might resist therapy or repair DNA breaks more efficiently [4]. Other DSB repair proteins, including MDC1, 53BP1 and RAD51 are attracted to the break ends and, accompanied by γ-H2AX, form ionizing radiation induced foci (IRIF) [12, 13]. The higher the number of induced γ-H2AX foci or slower disappearance rate, the more sensitive tumor cells are to radiation treatment. Persisting γ-H2AX IRIF in normal cells 24 h after radiation are associated with the development of late severe side effects
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