Hsp90 inhibitor is a good candidate for effective combination therapy with carbon ions

Abstract

Purpose/Background: To further improve the effectiveness of heavy ion radio-therapy, we studied the biology of a combined treatment using Hsp 90 inhibitor and carbon ion irradiation. We have previously reported that an Hsp90 inhibitor 17AAG can be an effective radio-sensitizer with X-rays for certain human tumor cells, while normal cells were not sensitized by this drug [ 1]. The underlying mechanism for this was demonstrated to be inhibition of DNA double-strand break (DSB) repair by 17AAG; particularly, homologous recombination repair (HRR) pathway was shown to be affected by this agent. In vivo mouse xenograft study also indicated a better tumor control with the combined treatment when compared with X-ray treatment alone.Materials and Methods: Cell lines used were SQ5 human lung cancer cells and HFL III normal human fibroblasts as control. For irradiation, X-rays and 290 MeV/n carbon ions were used at 50–70 keV/µm LET setting. As Hsp90 inhibitors, 17AAG and PU-H71 were used for pre-irradiation treatment for 24 h. Colony formation assay was used for radiation sensitivity studies. Repair of DNA DSBs was measured by constant field gel electrophoresis, and the appearance/disappearance of Rad51 foci was analyzed for HRR efficiency. For IRB-approved mouse xenograft study, BALB/c nu/nu mice were used to implant SQ5 cells for local irradiation.Results/Discussion: SQ5 tumor cells were better controlled with the combination of 17 AAG and carbon ion irradiation in vitro and in vivo xenograft model when compared with carbon irradiation alone. The cause of this radio-sensitizing effect seems to come from inhibition of repair of DNA DSBs by 17AAG as in X-rays. Likewise, HRR pathway was affected by addition of 17AAG in carbon irradiated tumor cells. The effect of 17AAG pre-treatment is shown below where the appearance of an HRR key protein Rad51 was significantly delayed after carbon ion irradiation in the drug-treated samples when compared with samples with carbon alone (Fig. 1).We also started to investigate PU-H71 with carbon ion irradiation in vitro and in vivo. PU-H71 alone is currently under phase I clinical trial. Our data indicate that PU-H71 pre-treatment also showed a significant radio-sensitization in SQ5 human lung tumor cells exposed to carbon ions as well as to X-rays. As shown with 17AAG, normal human cells were not significantly affected with this drug. PU-H71 also seems to affect repair of radiation-induced DSBs. Further mechanism studies and in vivo experiments are currently underway.Conclusion: Hsp90 inhibitor would be a good candidate for the effective combined treatment with carbon ion radio-therapy. Fig. 1.Post-irradiation Rad51 foci appearance kinetics in SQ5 cells irradiated with 2 Gy carbon ions compared with the combined treatment of 17AAG and carbon radiation (2 Gy).