Scanning Proton FLASH Irradiation Using a Synchrotron Accelerator: Effects on Cultured Cells and Differences by Irradiation Positions

Abstract

An ultra-high dose rate radiation (FLASH) is now attracting great attention. Although FLASH radiation with a cyclotron-type proton therapy machine has been reported, production of the ultra-high dose rate using other machines has scarcely been reported. In this study, we established a FLASH radiation system using a synchrotron proton beam accelerator and compared the effects of FLASH and conventional dose-rate radiation on cultured cells. Both types of radiation were delivered at the 1-cm spread-out Bragg peak (SOBP) and plateau region before SOBP. Protons were accelerated by adjusting the application pattern of high-frequency power. Other incident conditions for passive irradiation were also applied to increase proton accumulation to approximately twice the normal scanning conditions. Ultra-high dose rates were achieved by applying a high-frequency power dozens of times higher than usual to extract all protons within approximately 50 ms, thereby monitoring with a Faraday cup immediately after synchrotron extraction and adjusting the waveform. The dose and dose rates were measured using a float glass, gafchromic film, and plane parallel ionization chamber. Four cultured tumor cell lines and two normal cell lines were used. FLASH radiation at 6 and 10 Gy (200 Gy/s, 1 shot) and conventional radiation at 6 and 10 Gy (3 Gy/s) were delivered to the SOBP and plateau region before SOBP. After radiation, appropriate amounts of reagents to stain caspase-3/7 were added, and differences in cell growth rates and cell cycle were examined using live-cell fluorescence time-lapse imaging. The cell confluence ratio and apoptotic change were also quantified. V79 spheroids were subjected to FLASH (300 Gy/s, 10 and 18 Gy, 1 shot) and conventional radiation (3 Gy/s, 10 and 18 Gy) at SOBP. Subsequently, they were disaggregated to single cells and assayed for cell survival by colony assay. A 1-cm SOBP was formed by a mini-ledge filter was prepared with a high-definition 3D printer. FLASH radiation could be achieved at 40-800 Gy/s. In vitro, similar cell-killing effects were observed for FLASH and conventional radiation in the cancer cells, with cell-cycle delay. Significant apoptotic effects and G2 arrest during the cell cycle were observed for FLASH peak irradiation, probably due to the higher LET at the Bragg peak. Decreased proliferation and cell-killing effects were also observed in the normal cells for FLASH, but the apoptotic effect was not different between FLASH and conventional irradiation. Stronger cell-killing effects were observed with the V79 spheroids for FLASH peak than conventional peak irradiation. Scanning proton FLASH radiation with a synchrotron accelerator was established for biological studies. FLASH seemed to be equally effective for cancer and normal cells in vitro. The effects were higher with FLASH peak irradiation owing to the high LET at the Bragg peak, which is a characteristic of synchrotron accelerators.